Protein Intake PTC8 PDF

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PROTEIN INTAKE
Before we can get to the good stuff, it is important to understand some basic protein biology.
What is protein?
Proteins are found throughout the body, with over 40% of body protein found in skeletal
muscle, over 25% found in body organs, and the rest found mostly in the skin and blood.
Proteins are essential nutrients because of their constituent amino acids, which the body needs
to create its own variety of proteins and nitrogen-containing molecules that make life possible.
Each body protein is unique in the characteristics and sequence pattern of the amino acids that
comprise its structure.
Amino acids may be classified in a variety of ways, including by structure, net charge, polarity,
and essentiality.
Amino acid biochemistry 101
Structurally, all amino acids have a central carbon (C), at least one amino group (—NH2), at
least one carboxy (acid) group (—COOH), and a side chain (R group) that makes each amino
acid unique.
Depending on the pH (acidity) of the environment, the amino and carboxy groups can accept
or donate H+, and thus the amino group may be represented as +NH3 and the carboxy group
as COO−. Basically, amino acids can differ in structure in environments with different acidity
levels. The distinctive characteristics of the side chains of the amino acids that make up a
polypeptide bestow on a protein its structure and influence its functional role in the body,
including:
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 whether certain amino acids can be synthesized in the body or must be ingested;
 which metabolic pathways the amino acids can enter.
The 20 ‘natural’ amino acids found in the human genetic code. Don’t worry about the chemistry: you
just need to understand that the structure of an amino acid determines its function.
The tendency of an amino acid to interact with water at physiological pH—that is, its polarity—
represents another means of classifying amino acids. Polarity depends upon the side chain or R
group of the amino acid. Amino acids are classified as polar or nonpolar, although they can have
varying levels of polarity.
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Polar charged amino acids include both the dicarboxylic amino acids (aspartic acid and glutamic
acid) and basic amino acids (lysine, arginine, histidine). Polar charged amino acids interact with
aqueous environments, can form salt bridges, and can interact with electrolytes and minerals
such as potassium, chloride, and phosphate.
The neutral amino acids interact with water to different degrees and can be divided into polar,
nonpolar, and relatively nonpolar categories. The side chains of polar neutral amino acids
contain functional groups—such as the hydroxyl group for serine and threonine, the sulfur
atom for cysteine, and the amide group for asparagine and glutamine—that can interact through
hydrogen bonds with water (the aqueous environment of cells).
Polar amino acids are generally found on the surfaces of proteins. If not found on the surface,
they are oriented inward and function at a protein’s (such as an enzyme’s) binding site.
In contrast, the amino acids contain side chains that do not interact with water and are
categorized as nonpolar or hydrophobic (water fearing). The aromatic amino acids are
considered relatively nonpolar. Tyrosine, for example, because of its hydroxyl group on the
phenyl ring, can to a limited extent form hydrogen bonds with water—hence the term
relatively nonpolar. Because they do not interact with water, the nonpolar (and often the
relatively nonpolar) amino acids are typically found compacted and oriented toward or within
the central region or core portion.
As an illustration of the effect of the polarity of amino acids, the reason why (pure) branched
chain amino acids (BCAAs: you can check their structure to understand why they’re called this
way) are poorly soluble in water is their non-polarity. All 3 BCAAs - leucine, isoleucine and
valine - have a nonpolar side chain.
Take-home message: Proteins are formed by amino acids, which in turn vary in their
chemical structure. The chemical structure of an amino acid determines its function in the
body.
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Essential and non-essential amino acids
In contrast to chemists, nutritionists generally categorize the amino acids in terms of

essentiality. In the context of nutrition, the term essential or indispensable means that the body
cannot create the nutrient (in this case the amino acid) itself and so it must be supplied by the
diet.
The 9 essential amino acids are:
 leucine
 isoleucine
 valine
 lysine
 tryptophan
 threonine
 methionine
 phenylalanine
 histidine
Identifying amino acids strictly as nonessential/dispensable or essential/indispensable is an

inflexible classification, however, that allows no gradations, even in decidedly different or
changing physiological circumstances. Therefore, a third category has been added: conditionally
essential amino acids, also known as acquired indispensable amino acids. For example, a
dispensable amino acid may become indispensable if a certain organ fails to function properly.
Protein biochemistry 101
The 20 natural amino acids are proteinogenic, protein building. Amino acids strung together
form peptidide chains that are in turn incorporated into proteins.
 Peptide: a short amino acid chain.

o Dipeptide: a peptide consisting of 2 amino acids.
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o Tripeptide: a peptide consisting of 3 amino acids.

o Peptides with 2-20 amino acids are called oligopeptides.
o Peptidues with more than 20 amino acids are called polypeptides.
These amino acids are folded into shape by your ribosomes to create complex structures.
Proteins have 4 levels of structure:
1. The primary structure of a protein represents the amino acid sequence of the protein.
2. The secondary structure is the coiling, folding and bending of the protein. This shape is
created by i.a. hydrogen bonds, which are electrical attractions that keep atoms
together and electrostatic/ionic attractions between amino acids (akin to magnets).
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3. The tertiary structure of protein is the overall or total three-dimensional configuration

of the protein. This is basically the resulting total shape of the combined secondary
structure shapes attached together.
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4. Some proteins also have a quatarnary structure as a result of interacting polypeptide

chains that form a so called oligomer.
Now that you understand what proteins and amino acids are, the next step is understanding
how your body acquires amino acids.
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Amino acid sources

When the body needs an amino acid, it derives it by breaking down a protein. Both dietary
(exogenous) and endogenous (internal) proteins provide the body with amino acids for use.
Dietary (exogenous) sources of protein include:

- animal products such as meat, poultry, fish, eggs, and dairy products (with the exception
of butter, sour cream, and cream cheese);
- plant products such as grains, grain products, legumes, and vegetables.
Following ingestion, exogenous proteins serve as sources of the essential amino acids, non-
essential amino acids, and the additional nitrogen needed to synthesize more non-essential
amino acids and nitrogen-containing compounds in the body.
Endogenous proteins presented to the digestive tract represent another source of amino acids
and nitrogen. Endogenous proteins include:
- desquamated mucosal cells, which generate about 50 g of protein per day;
- digestive enzymes and glycoproteins, which generate about 17 g of protein each day.
The digestive enzymes and glycoproteins are derived from digestive secretions of the salivary
glands, stomach, intestine, liver, and pancreas. The intestinal cells contain a variety of proteins
(such as apoproteins, structural proteins, cytosolic enzymes) that are degraded when sloughed
into the gastrointestinal tract. Most of these endogenous proteins, which may total about 70 g
or more per day, are digested and provide amino acids that are available for absorption.
Note, however, that this is not ‘free protein’ that you need to substract from your total protein
need, as ultimately the building blocks of these proteins still have to be consumed in some
form. Ultimately, your body derives all of the building blocks it needs to create proteins from
exogenous sources, notably the consumption of food.
Protein digestion
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No appreciable digestion of protein occurs in the mouth or esophagus, so the first location of
considerable protein digestion is the stomach with the action of hydrochloric acid (HCl), which
is found in gastric juice. The hydrochloric acid content of the gastric juice results in a highly
acidic environement with a gastric pH of about 1 to 2 that enables denaturation (disruption) of
the quaternary, tertiary and secondary structures of protein. Denaturants such as hydrochloric
acid break apart hydrogen and electrostatic bonds to unfold or uncoil the protein (i.e. destroy
the quaternary, tertiary and secondary structure); however, peptide bonds (the primary
structure) are not affected by the hydrochloric acid.
This denaturation is a normal part of protein digestion, so the concern that denaturation of
protein during cooking ‘destroys the protein’ is not a problem at all. Cooking is in many ways
basically a form of pre-digestion.
Another function of hydrochloric acid is to activate the digestive enzyme pepsin. Pepsin
functions as an endopeptidase, which means that it hydrolyzes interior peptide bonds within
proteins or polypeptides at a pH below ∼3.5. (Specifically, pepsin attacks peptide bonds
adjacent to the carboxy end of a relatively wide variety of amino acids (i.e., pepsin has low
specificity).)
Thus, the end products of gastric protein digestion in the stomach include primarily large
polypeptides, along with some oligopeptides (short chains of amino acids bonded to each other)
and free amino acids. These subsequently enter the small intestine.
In the small intestine peptides are broken down further by a number of additional digestive
enzymes, as illustrated below.
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Protein digestion yields two main end products: peptides (principally dipeptides and tripeptides)
and free amino acids. To be used by the body, these end products must next be absorbed.
Protein absorption
Absorption is the process by which the end products of digestion are transported from the
lumen of the gastrointestinal tract, most often the intestine, into the body. To get into the
blood for transport to tissues, amino acids must cross two intestinal membranes, the brush
border (also called apical) membrane and the basolateral (also called serosal) membrane.
Amino acid absorption occurs along the entire small intestine, but most amino acids are
absorbed in parts called the duodenum and upper jejunum. The absorption of amino acids into
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enterocytes requires carriers; however, paracellular absorption — that is, absorption via
passage through the tight junctions of enterocytes (intestinal cells) or by transcellular
endocytosis (transport through the cell by encapsulating the transported molecule into a
vesicle) — can occur in occasional situations in which large volumes of hypotonic fluids
(solution that contains fewer solutes and more water than solutions in the body) containing
some amino acids have been ingested.
The transport systems are responsible for carrying amino acids across the brush border
membrane into the intestinal cell. Most amino acids are thought to be transported across the
enterocyte brush border membrane by so called sodium-dependent transporters.
The affinity of a carrier for an amino acid is influenced both by the hydrocarbon mass of the
amino acid’s side chain and by the net electrical charge of the amino acid. As the hydrocarbon
mass of the side chain increases, affinity increases. Thus, the branched-chain amino acids
typically are absorbed faster than smaller amino acids. Neutral amino acids also tend to be
absorbed at higher rates than basic or acidic amino acids. Essential (indispensable) amino acids
are absorbed faster than nonessential (dispensable) amino acids, with methionine, leucine,
isoleucine, and valine being the most rapidly absorbed.
Ingesting large quantities of one amino acid or a particular group of amino acids that use the
same carrier system may create, depending upon the amount ingested, competition among the
amino acids for absorption. The result may be that the amino acid present in highest
concentration is absorbed but also may impair the absorption of the other, less concentrated
amino acids carried by that same system. Thus, amino acid supplements may result in impaired
or imbalanced amino acid absorption.
Additionally, the absorption of peptides is generally more rapid than absorption of an equivalent
mixture of free amino acids.
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Moreover, nitrogen assimilation following ingestion of protein-containing foods is superior to

that following ingestion of free amino acids. In other words, free amino acids have no
absorptive advantage, in contrast to what supplement companies may claim.
Although peptides, like amino acids, compete with one another for transporters, peptide
transport appears to occur more rapidly than amino acid transport and is thought to represent
the primary means by which most amino acids enter the intestinal cell. In other words, the
majority of amino acids are absorbed as peptides.
Peptides, once within the enterocytes (intestinal cells), are generally hydrolyzed by cytosolic
peptidases to generate free intracellular amino acids. Intact peptides, however, can be found
occasionally in circulation, and this is thought to result from entry of peptides into the body via
paracellular (also called intercellular) routes. With illnesses, especially those affecting the
intestines (such as inflammatory bowel diseases or celiac disease), the gastrointestinal tract can
become more permeable, thus increasing the likelihood of peptides appearing intact in the
blood. Peptides found in the blood are thought to be hydrolyzed by peptidases or proteases in
the plasma, at the cell membrane (especially in the liver, kidneys, and muscle), or, following
transport into tissues, intracellularly in the cytosol or in various organelles.
The ability to administer peptides directly into the blood as in parenteral nutrition is of
nutritional significance since some amino acids (e.g., tyrosine, cysteine, and glutamine) are
insoluble or unstable in their free form. Administration of the peptide form, which can be used
by tissues, thus allows nutrients to be provided in situations in which traditional free amino acid
parenteral mixtures are ineffective.
For amino acids to enter the blood and be used by other body tissues, the amino acids must be
transported across the basolateral (also called serosal) membrane of the enterocyte and into
interstitial fluid, where they enter the blood through capillaries of the villi for transport into the
portal vein leading to the liver and other tissues. The carriers found in the enterocyte’s
basolateral membrane are generally the same as those found in the membranes of nonepithelial
cells of the body.
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After amino acids are transported out of the enterocyte, they enter portal blood and are
transported to the liver. Uptake of the amino acids into liver cells (hepatocytes), as well as cells
of the kidney and other organs, occurs by some carrier systems similar to those found in the
intestinal cell membranes.
Take-home message: Dietary protein is your body’s principle source of amino acids. Protein
is generally digested in the stomach, subsequently absorbed in the intestine and then
transported in the blood to the liver.
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Amino acid metabolism

The liver is the primary site of uptake of most amino acids following ingestion of a protein-
containing meal. The liver is thought to monitor the absorbed amino acids and to adjust the
rate of their metabolism according to the needs of the body by selective catabolism (breaking
down) and anabolism (building up). We’ll first discuss why and how the body breaks down
amino acids.
Amino acid catabolism
The liver is the main site for the catabolism of the indispensable amino acids, with the exception
of the branched-chain amino acids, which tend to be utilized to a greater extent by muscle and
other organs such as the heart. Liver cells have a high capacity for the uptake and catabolism of
amino acids. Catabolism of amino acids occurs to varying degrees in different tissues both
during fasting periods and immediately after eating (the postprandial period). After a meal, the
liver takes up about 50% to 65% of amino acids from portal blood and the liver derives up to
50% of its energy (ATP) from amino acid oxidation. The energy generated may in turn be used
for gluconeogenesis or urea synthesis, among other needs, depending upon the body’s state of
nutrition.
Amino acids are broken down in cells, starting with the transamination or deamination of amino
acids and then the disposal of ammonia, often so that the carbon skeleton of amino acids can be
used and the waste products, like urea, can be discarded.
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Often, the first step of amino acid catabolism is the transfer or removal of an amino acid’s amino group
via transamination.
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Deamination reactions involve only the removal of an amino group from an amino acid via an enzyme,
like dehydrogenase, to produce an α-keto acid (carbon skeleton) and ammonia or an ammonium ion
(the two can be converted into each other).
Subsequently, the disposal of ammonia and ammonium ions takes place in the urea cycle. Urea
is formed by deriving one nitrogen from ammonia (from deamination reaction), a second
nitrogen from aspartate (from transamination reaction) and carbon from CO2/HCO3-.
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The urea cycle.
Once formed, urea typically travels in the blood to the kidneys for excretion in the urine;
however, up to about 25% of urea may be secreted from the blood into the intestinal lumen,
where it may be degraded by bacteria to yield ammonia. Activities of urea cycle enzymes
fluctuate with diet and hormone concentrations. For example, with low-protein diets or
acidosis, urea synthesis diminishes and urinary urea nitrogen excretion decreases significantly.
Thus, substrate availability results in short-term changes in the rate of ureagenesis. In the
healthy individual with a normal protein intake, blood urea nitrogen (BUN) concentrations
range from about 8 to 20 mg/dL, and urinary urea nitrogen represents about 80% of total
urinary nitrogen.
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Several defects (genetic mutations) can occur in urea cycle enzymes that significantly affect our
health. Urea cycle enzyme defects typically result in high levels of blood ammonia (hyper-
ammonemia) and necessitate a protein-restricted diet.
Discarding urea is just waste disposal. The actual usefulness of amino acid catabolism lies in the
produced carbon skeletons.
Carbon skeletons of amino acids can be further metabolized with the potential for multiple uses
in the cell, depending on the original amino acid from which they were derived. Whereas all
amino acids can be completely oxidized to generate energy, not all amino acids can be used, for
example, for the synthesis of glucose. Furthermore, the fate of the amino acid’s carbon skeleton
depends upon the body’s need at the time. In general, an amino acid’s carbon skeleton can be
used to produce:
● energy;
● glucose;
● ketone bodies;
● cholesterol;
● fatty acids.
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Energy generation
The complete oxidation of amino acids generates energy, CO2/HCO3−, and ammonia/
ammonium ions. Amino acids are not preferentially used for energy in the body, however,
unless the diet provides inadequate other energy sources or there is an excess of amino acids
available.
Gluconeogenesis and ketone body production
The production of glucose from a non-carbohydrate source such as amino acids is called
gluconeogenesis.
Gluconeogenesis occurs primarily in the liver but also in the kidneys and small intestine.
The carbon skeletons of all natural amino acids except leucine and lysine, together called the
glucogenic amino acids, can be used to synthesize glucose. To be considered a glucogenic amino
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acid, catabolism of the amino acid must yield pyruvate or an intermediary of the TCA cycle, like
oxaloacetate or succinyl-CoA.
It is less well known in nutritional circles that there several ketogenic amino acids that can be
used to produce ketone bodies in addition to glucose. In fact, leucine and lysine are ketogenic
but not glucogenic. For an amino acid to be considered ketogenic, the catabolism of the amino
acid must generate acetyl-CoA or acetoacetate, which are used for the formation of ketone
bodies.
The ketogenic amino acids are:

 Isoleucine
 Phenylalanine
 Threonine
 Tryptophan
 Tyrosine
 Leucine
 Lysine
The graphic below shows the general fates of amino acid carbon skeletons with respect to key
intermediates of metabolism. The conversion of amino acids to glucose is accelerated by a high
blood glucagon to insulin ratio and by high blood cortisol concentrations. Concentrations of the
hormone glucagon are generally elevated in the blood when blood glucose concentrations are
relatively low, as may occur in between meals or with a fast causing depletion of liver glycogen
stores. Blood glucagon (and in some cases cortisol along with epinephrine) is also elevated in
the presence of infection or trauma/injury and in certain disease states such as untreated
diabetes mellitus and liver disease.
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Cholesterol production
The amino acids whose oxidation yields acetyl-CoA can be metabolized to produce cholesterol.
Additionally, leucine is the only amino acid whose catabolism directly generates β-hydroxy β-
methylglutaryl-CoA, an intermediate in cholesterol synthesis. Leucine oxidation also produces
β-hydroxy β-methylbutyrate (HMB), which appears to promote de novo cholesterol synthesis in
muscle, enabling cell growth and function.
Fatty acid production
The carbon skeleton of amino acids can also be used to synthesize fatty acids. This occurs
parimarily in times of excess energy and protein intakes coupled with adequate carbohydrate
intake. Leucine, for example, can be used to synthesize fatty acids in adipose tissue.
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Other uses of amino acids
In addition to the above compounds that can be produced by the body by oxidizing amino
acids, the compounds in the image below can also be produced.
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Protein synthesis
When the diet is adequate in providing energy and other nutrients, dietary protein is
preferentially used for anabolism rather than catabolism. As a strength trainee, anabolism is
obviously the fate of amino acids that is desirable.
Anabolism, including protein synthesis, increases in tissues following the ingestion of food. Food
provides the amino acids necessary to build body proteins. The form and nature of the dietary
protein influences amino acid use within the body.
For example, amino acids released following ingestion and digestion of “fast proteins”, which
include whey protein, soy protein, amino acid mixtures and protein hydrolyzates (e.g., partially
hydrolyzed protein), appear to be utilized differently in the body when compared with amino
acids released following ingestion and digestion of “slow proteins” such as casein. Fast protein
ingestion causes plasma amino acids to rise more quickly and to higher concentrations but also
to fall more quickly than ingestion of equal amounts of slow proteins. Ingestion of slow proteins
results in lower and prolonged plasma amino acid concentrations and often better nitrogen
(protein) retention than ingestion of fast proteins. The hyperamino-acidemia (higher than
normal amino acid concentrations in the blood) after fast protein ingestion can promote higher
amino acid deamination rates and urea production than slow protein ingestion. In addition,
amino acids from fast proteins appear to be used more for splanchnic (internal organs) protein
synthesis versus peripheral (membrane proteins) or skeletal muscle protein synthesis. Protein
quality will be discussed in detail later on.
Hormones play a major role in amino acid utilization in the body. During prolonged periods in
which food is not eaten (the post-absorptive state), such as during the overnight hours or a
fast, protein synthesis still occurs but at a much lower rate and protein degradation
predominates. The degradative processes are stimulated by epinephrine and cortisol release
and by the higher glucagon to insulin ratio in the blood. This higher glucagon to insulin ratio
diminishes both insulin’s ability to inhibit protein degradation and the overall rate of protein
synthesis. Yet, while skeletal muscle experiences the most protein degradation and limited
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protein synthesis during post-absorptive periods, the glucagon to insulin ratio favoring glucagon
stimulates the hepatic synthesis of some proteins, such as enzymes for gluconeogenesis and
ureagenesis. High blood cortisol concentrations further promote muscle protein catabolism and
hepatic use of the amino acids for gluconeogenesis and ureagenesis. Blood cortisol
concentrations typically increase with infection and injury/trauma but also with depletion of
hepatic glycogen stores, as occurs during an extended fast.
In contrast to the general catabolic nature of glucagon, epinephrine, and cortisol, other
hormones in the body, such as growth hormone and insulin, are anabolic.
Insulin, for example, which is secreted in response to a rise in blood glucose and to a rise in
some blood amino acid concentrations (as occurs with food consumption), generally increases
protein synthesis and decreases protein degradation. To promote protein synthesis, insulin, for
example, affects (generally stimulates) the transcellular movement of amino acid transporters to
the cell membrane and increases the overall activity of amino acid transporters in the liver,
muscle, and other tissues. Insulin also antagonizes the activation of some enzymes responsible
for amino acid oxidation (degradation). Thus, insulin promotes the uptake of the amino acids
into the tissues and inhibits the enzymes that are responsible for the degradation of the amino
acids. Such actions of insulin facilitate the use of the amino acids for protein synthesis. Insulin,
however, along with the amino acid leucine (which by itself acts as an insulin secretogogue, i.e.,
it stimulates insulin secretion), also plays other roles in protein synthesis through effects on the
initiation of protein translation along the ribosomes.
Protein synthesis involves transcription of a gene into messenger (m) RNA followed by its
translation. Multiple factors influence translation. Translation is affected by the amount and
stability of mRNA, the ribosome number, the activity of the ribosomes, the presence of amino
acids in the appropriate concentrations, and the hormonal environment, which in turn can be
influenced by nutrients. Some amino acids may even regulate the expression of some genes
through interactions with amino acid–responsive elements in the gene’s promoter region.
Amino acids, for example, can promote changes in cell volume and protein synthesis through
intracellular signaling pathways. Leucine and a leucine metabolite, β-hydroxy-β-methylbutyrate
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(HMB), for instance, promote protein synthesis through increased phosphorylation of

mammalian target of rapamycin (mTOR). mTOR mediates the effects of insulin on protein
synthesis through different multiprotein complexes but also exerts effects on protein synthesis
that are independent of insulin.
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Diagram detailing the role of amino acid availability in regulating muscle

protein synthesis with amino acid/protein ingestion. (Source: Witard et al. 2016)
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Take-home message: Protein synthesis and catabolism occur in the body at various locations
across the entire day. Net protein balance determines whether there is net tissue build-up or
breakdown. Which tissues are built on and which are broken down is determined by many
factors, notably hormone levels.
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The function of proteins and nitrogen-containing compounds

The molecular architecture and activity of living cells depends largely on proteins, which make
up over half of the solid content of cells and which show great variability in size, shape, and
physical properties. Proteins’ physiological roles are highly varied, and because of this variation,
proteins are categorized according to their function. You don’t have to memorize all these
functions, just understand them so that you can read texts that mention them.
Structural elements
Several proteins have structural roles in the body. They are effectively part of the bricks,
mortar and steel that is your body’s house. Some of these proteins include:
- contractile proteins;
- fibrous proteins.
The two main contractile proteins, actin and myosin, are found in cardiac, skeletal, and smooth
muscles.
Skeletal muscle is found throughout the body and is under voluntary control. Contraction is
calcium induced and involves not only actin and myosin, but also troponin and tropomyosin.
Smooth muscle is found in many tissues including, for example, blood vessels, the lungs, the
uterus, and the gastrointestinal tract. Smooth muscle is under involuntary control and contracts
in response to calcium-induced phosphorylation of the structural protein myosin.
Fibrous proteins, which tend to be somewhat linear in shape, include collagen, elastin, and
keratin and are found in bone, teeth, skin, tendons, cartilage, blood vessels, hair and nails.
Collagen is a group of proteins with very strong structures, because each type of collagen is
made of three polypeptide (tropo-collagen) chains that are cross-linked for strength.
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Take-home message: Contractile proteins are what enable you to flex your muscles. Fibrous
proteins give strength to your connective tissue.
Catalysts
Enzymes are protein molecules (generally designated by the suffix -ase) that act as catalysts.
They change the rate of reactions occurring in the body. Enzymes are necessary for sustaining
life and are found in the body both intracellularly (within cells) and extracellularly (e.g., in the
blood). Enzymes are constructed so that they combine selectively with other molecules (called
substrates) in the cell. The active site on the enzyme (a small region usually in a crevice of the
enzyme) is where the enzyme and substrate bind and the product is generated. Some enzymes,
however, require a co-factor or co-enzyme to carry out the reaction. Minerals such as zinc,
iron, and copper function as co-factors for some enzymes. B-vitamins serve as co-enzymes for
many enzymes. Most human physiological processes require enzymes to promote chemical
changes that could not otherwise occur. Some examples of different types of enzymes include
dehydrogenases, which remove or transfer hydrogens; kinases, which add phosphate groups;
and isomerases, which transfer atoms within a molecule. Some examples of physiological
processes that depend upon enzyme function include digestion (e.g. pepsin, described earlier),
energy production, blood coagulation, and excitation and contraction of neuromuscular tissue.
Take-home message: Enzymes are proteins that speed up or slow down chemical reactions
and processes in the body.
Messengers
Some proteins are hormones. Hormones act as chemical messengers in the body. They are
synthesized and secreted by endocrine tissue (glands) and transported in the blood to target
tissues or organs, where they bind to protein receptors on membranes. Hormones generally
regulate metabolic processes by, for example, promoting enzyme synthesis or affecting enzyme
activity. Whereas some hormones are derived from cholesterol and classified as steroid
hormones, others are derived from one or more amino acids. The amino acid tyrosine, for
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example, is used along with the mineral iodine to synthesize the thyroid hormones. Tyrosine is
also used to synthesize the catecholamines, including dopamine, norepinephrine, and
epinephrine. The hormone melatonin is derived in the brain from the amino acid tryptophan.
Other hormones are made up of one or more polypeptide chains. Insulin, for example, consists
of two polypeptide chains linked by a disulfide bridge.
Take-home message: Certain proteins are hormones that act as messengers in the body to
activate or deactive other processes.
Buffers
Proteins, because of their constituent amino acids, can serve as buffers in the body that help to
regulate acid-base balance. A buffer is a compound that ameliorates a change in pH, acidity
level, that would otherwise occur in response to the addition of alkali or acid to a solution. The
pH of the blood and other body tissues must be maintained within an appropriate range. Blood
pH ranges from about 7.35 to 7.45, whereas cellular pH levels are often more acidic (lower
pH). For example, the pH of red blood cells is about 7.2 and that of muscle cells is about 6.9.
The H+ concentration within cells is buffered by both the phosphate system and the amino
acids in proteins.
Take-home message: Certain proteins act as buffers that regulare acidity levels.
Fluid Balancers
In addition to acid-base balance, proteins influence fluid balance through their presence in the
blood and in cells. More specifically, proteins help attract and keep water inside a particular
area and contribute to osmotic pressure. Diminished blood/plasma concentrations of proteins,
such as albumin (the most abundant of the proteins in the blood), result in a decrease in plasma
osmotic pressure. When protein concentrations in the blood are less dense than normal, fluid
“leaks” out of the blood and into interstitial spaces and causes swelling (edema). Restoring
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adequate concentrations of protein in the blood (e.g., by infusing albumin intravenously)

promotes diffusion of water from the interstitial space back into the blood.
Take-home message: Certain proteins act as fluid balancers that regulate water levels.
Immuno-protectors and acute phase responders
Immuno-protection is provided to the body in part by a group of proteins called immuno-

proteins, also called immuno-globulins (Ig) or, more commonly, anti-bodies (Ab).
Immunoglobulins function by binding to anti-gens — which typically consist of foreign
substances, such as bacteria or viruses that have entered the body — and inactivating them.
Another related group of proteins, called acute phase or positive acute phase reactant proteins,
is as responders in times of acute (sudden), critical illness. These proteins are made in the liver
as part of the body’s response to infection (sepsis), injury or inflammation. Collectively, these
proteins perform a variety of functions that protect the body, such as stimulating the immune
system, promoting wound healing, and chelating and removing free iron from circulation to
prevent its use by bacteria for growth. C-reactive protein is used clinically to evaluate
inflammation in patients. The concentration of this protein rises dramatically within a few hours
of inflammation such as that caused by an infection.
Take-home message: Certain proteins are anti-bodies in your immune system that prevent
you from getting sick; other proteins have similar functions to keep you healthy when you do
get unwell.
Transporters
Transport proteins are a diverse group of proteins that combine with other substances
(especially vitamins and minerals but also other nutrients) to provide a means of carrying those
substances within the blood, into cells, out of cells, or within cells. Transport proteins in cell
membranes, for example, carry and thus regulate the flow of nutrients into and out of cells.
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A few transport proteins of clinical significance include albumin, transthyretin, and retinol-
binding protein. Albumin, the most abundant of the plasma proteins, transports nutrients such
as tryptophan, fatty acids, and vitamin B6, some minerals including zinc, calcium, and small
amounts of copper and some drugs. The protein is synthesized by the liver and released into
the blood. Changes in osmotic pressure and osmolality affect its rate of synthesis. A healthy
person makes about 9 to 12 g of albumin per day. Albumin is often used to assess an individual’s
protein status, specifically visceral (internal organ) protein status. Because of albumin’s relatively
long half-life (∼14–18 days), however, it is not as good or as sensitive an indicator of visceral
protein status as some of the other plasma proteins. The half-life is the time it takes for 50% of
the amount of substance to be degraded. Typically, plasma concentrations of albumin < 3.5
g/dL, prealbumin (transthyretin) < 18 mg/dL, and retinol-binding protein < 2.1 mg/dL suggest
inadequate visceral protein status. Such people need a diet high in energy (kcal) and protein to
promote improvements (assuming the liver is healthy) in status.
Take-home message: Certain proteins act as transporters, akin to tiny ships in your body
that have nutrients, hormones or drugs as their cargo.
Other roles
Proteins carry out many additional roles in the body, but they are generally not relevant for
nutritionists. For example, in cell membranes, proteins function in cell adhesion, and some
others serve to transmit signals into and out of the cell.
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Nitrogen-containing non-protein compounds

In addition to their use in the synthesis of body proteins, amino acids are used to synthesize
nitrogen-containing compounds that are not proteins but nonetheless play important roles in
the body. Again, you don’t need to memorize the functions of all of these compounds in detail,
just understand what they do so that you can understand texts that mention them.
Glutathione
Glutathione is a tripeptide synthesized from three amino acids — glycine, cysteine, and
glutamate — in most body cells. Glutathione has several functions in the body. It is a major
anti-oxidant with the ability to scavenge free radicals (O2• and OH•), thereby protecting critical
cell components against oxidation.
Carnitine
Carnitine, another nitrogen-containing compound, is made from the amino acid lysine. As such,
it does not need to be supplemented to avoid deficiency, in contrast to what supplement
companies sometimes claim. In addition to being synthesized in the liver and kidneys, carnitine
is found in foods, especially meats such as beef and pork. Carnitine from food or supplements is
absorbed in the proximal small intestine. Approximately 54% to 87% of carnitine intake is
absorbed. Intestinal absorption of carnitine is thought to be saturated with intakes of about 2 g.
Muscle represents the primary carnitine pool, although no carnitine is made there. Carnitine,
found in most body tissues, is needed for the transport of fatty acids, especially long-chain fatty
acids, across the inner mitochondrial membrane for oxidation. Carnitine is also needed for
ketone catabolism for energy. Carnitine deficiency, though rare, results in impaired energy
metabolism.
Creatine
33
Creatine, a key component of the energy compound creatine phosphate, also called
phosphocreatine, can be obtained from foods (primarily meat and fish) or synthesized from
certain amino acids in the body.
Once synthesized, creatine is released into the blood for transport to tissues. About 95% of
body creatine is in muscle, with the remaining 5% in organs such as the kidneys and brain.
In tissues, creatine is found both in free form as creatine and in its phosphorylated form.
Phosphocreatine functions as a “storehouse for high-energy phosphate.” In fact, over half of the
creatine in muscle at rest is in the form of phosphocreatine. Phosphocreatine replenishes ATP
in a muscle that is rapidly contracting.
Remember, muscle contraction requires energy. This energy is obtained from the hydrolysis of
ATP. However, the ATP in muscle can suffice for only a fraction of a second. Phosphocreatine,
stored in the muscle and possessing a higher phosphate group transfer potential than ATP, can
transfer a phosphoryl group to ADP, thereby forming ATP or assisting in ATP regeneration,
providing energy for muscular activity. Creatine kinase, also called creatine phosphokinase
(abbreviated CK or CPK), catalyzes the phosphate transfer in active muscle.
There are two types of CK; CK-MB and CK-MM. An elevation in CK-MB in the blood along
with other indicators is used to diagnose a heart attack. Similarly, damage to skeletal muscle, as
may occur with trauma, results in elevations of CK-MM in the blood.
The availability of phosphocreatine and its use by muscle delay the breakdown of muscle
glycogen stores, which upon further catabolism are also used by the muscle for energy.
Creatine and creatine phosphate do not remain indefinitely in muscle; rather, both slowly but
spontaneously cyclize because of non-reversible, non-enzymatic dehydration. This cyclization of
creatine and phosphocreatine forms creatinine. Once formed, creatinine leaves the muscle,
passes across the glomerulus of the kidneys, and is excreted like other nitrogenous waste
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products (e.g., urea, ammonia, uric acid) in the urine. Creatinine clearance is sometimes used as
a means of estimating kidney function. The urinary excretion of creatinine is used as an
indicator of existing muscle mass.
Carnosine
Carnosine is made in the body from the amino acids histidine and β-alanine. In the body,
carnosine is synthesized and found largely in the cytosol of skeletal and cardiac muscle, but it is
also found in the brain, kidneys, and stomach. Carnosine is found in foods, primarily meats, and
may be digested into histidine and β-alanine in the intestine or possibly absorbed intact by
peptide transporters. While not all of the functions of carnosine have been identified, some
studies have shown that carnosine acts as both a buffer and an anti-oxidant within cells; it may
also reduce calcium needs for muscle contractility.
Choline
Choline is made in the body from methylation of the amino acid serine. Choline is found in free
form in foods in small amounts but is more commonly found in foods as part of the
phospholipid lecithin (phosphatidyl choline). Foods rich in lecithin include eggs, liver and other
organ meats, muscle meats, shrimp, cod, salmon, wheat germ and legumes such as soybeans and
peanuts. Lecithin is also added to many foods as an emulsifier. In the body, choline has several
functions, including the synthesis of the neurotransmitter acetylcholine.
Experimental diets devoid of choline can result in decreases in plasma choline and
phosphatidylcholine concentrations as well as alterations in some liver enzymes. Animals devoid
of dietary choline develop a fatty liver accompanied by some hepatic necrosis. Low intakes of
both choline and betaine have been associated with inflammation. The Food and Nutrition
Board has suggested that an Adequate Intake is 425 mg and 550 mg of choline daily for adult
females and males, respectively. Such intakes are easily obtained through dietary consumption
of animal products and foods containing fats. A Tolerable Upper Intake Level of 3.5 g of choline
daily also has been set.
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Purine and pyrimidine bases
Nitrogenous bases, along with a five-carbon sugar and phosphoric acid, are needed for the
synthesis of two nucleic acids, deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), in the
body. It is amino acids that provide the source for the nitrogen in these bases. The nitrogenous
bases can be divided into two categories: pyrimidines and purines.
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Inter-organ ‘flow’ of amino acids and organ-specific

metabolism
While tissues and organs use amino acids to synthesize proteins and some nitrogen-containing
compounds, the metabolism of the amino acids varies to some extent among the different
organs. In many instances, the products generated from amino acid metabolism in one organ
may be needed by another organ, creating a dependence between organs.
Amino acids in the blood
After ingestion of a protein-containing meal, amino acid concentrations typically rise in blood
plasma for several hours, before they return to basal concentrations. In basal situations or
between meals, plasma amino acid concentrations are relatively stable; however, absolute
concentrations of specific amino acids in the plasma vary from person to person.
Amino acids circulating in the plasma and found within cells arise from digestion and absorption
of dietary (exogenous) protein as well as from the breakdown of existing body (endogenous)
tissues.
The endogenous amino acids intermingle with exogenous amino acids to form a “pool” totaling
about 150 g of protein, ‘the amino acid pool’. The pool includes amino acids in the plasma as
well as amino acids in the cytosol of body cells. Reuse of endogenous amino acids is thought to
represent the primary source of amino acids for protein synthesis. Despite differences in
protein intake and rates of degradation of tissue proteins, the pattern of the amino acids in the
amino acid pool appears to remain relatively constant, although the pattern is quite different
from that found in body proteins. The total amount of the essential amino acids found in the
pool is less than that of the non-essential amino acids. The essential amino acids found in
greatest concentrations are lysine and threonine. Of the non-essential amino acids, those found
in greatest concentrations are alanine, glutamate, aspartate, and glutamine. In fact, up to 80 g of
glutamine can be found in the body’s amino acid pool. Amino acids within the pool, regardless
of source, are taken up by tissues and metabolized in response to various stimuli such as
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hormones and your physiological state. Tissues, for example, extract amino acids for energy
production or for the synthesis of nonessential amino acids, protein, nitrogen-containing
nonprotein compounds, biogenic amines, neurotransmitters, neuropeptides, hormones, glucose,
fatty acids, or ketones, depending on the person’s nutritional status and hormonal environment.
Take-home messages: Your body has an amino acid pool in the blood and the fluids in your
cells that it uses to temporarily store amino acids so that your tissues can take up the amino
acids it needs even if they weren’t consumed shortly beforehand. The amino acid pool allows
your body considerable leeway with regard to the need for nutrient timing.
Intestinal cell amino acid metabolism
Intestinal cells, enterocytes, use amino acids for energy production as well as for the synthesis
of proteins and nitrogen-containing compounds. Some of the uses of amino acids in enterocytes
include:
● structural proteins
● nucleotides
● apoproteins necessary for lipoprotein (chylomicron) formation
● new digestive enzymes
● hormones
● nitrogen-containing compounds
In addition, amino acids may be totally or partially metabolized within intestinal cells. It is
estimated that the intestine, which represents about 3–6% of the body weight, uses 30% to 40%
and splanchnic tissues (your abdominal organs) use up to 50% of some of the essential amino
acids absorbed from the diet. Moreover, the intestines are thought to use up to about 90% of
glutamate absorbed from the diet.
Take-home messages: Your intestines require a sufficient protein intake to function well and
use up much of the protein from your diet.
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Organ-specific glutamine metabolism
Glutamine has several major roles in the body, one of which is in ammonia transport. Whereas
ammonia arising in the liver from amino acid reactions is typically shuttled to the urea cycle, this
is not the fate of ammonia in all other tissues. Glutamine is involved in several important body
functions that are especially critical during illness/injury. For example, glutamine is used
extensively by immune system cells and promotes the synthesis of heat shock/stress proteins,
which help protect body cells.
Glutamine also serves several roles in intestinal cells. It is degraded extensively by intestinal
cells as a primary source of energy. It also has been shown to have trophic (growth) effects,
stimulating gastro-intestinal mucosa cell proliferation. Consequently, glutamine helps to prevent
both atrophy of gut mucosa and bacterial translocation. In addition, glutamine has been shown
to enhance the synthesis of heat shock proteins. It is also needed in large quantities along with
threonine for the synthesis of mucins found in gastrointestinal tract mucus secretions. These
roles of glutamine in the gastrointestinal tract have prompted several companies to enrich
enteral and parenteral (intravenous) nutrition products with glutamine. When glutamine is
provided through tube feedings, over 50% of glutamine is extracted by the splanchnic (visceral)
bed. It is estimated that the human gastrointestinal tract uses up to 10 g of glutamine per day,
and that the cells of the immune system use over 10 g per day. In addition to dietary glutamine,
much of the body’s glutamine that is produced by the skeletal muscles (and to lesser extents by
the lungs, brain, heart, and adipose tissue) is released and taken up, mostly by the intestinal
cells.
It is estimated that the average body produces 40 to 80 g of glutamine per day. Glutamine use
by cells increases dramatically during hypercatabolic conditions such as infection and trauma. In
these conditions, muscle glutamine release increases but cannot meet other cellular demands.
Thus, glutamine stores can become depleted and several cell functions may become impaired.
Take-home message: Glutamine is particularly important for intestinal health and your
immune system.
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The alanine-glucose cycle
The amino acid alanine is important in the inter-tissue (between tissues) transfer of amino
groups generated from amino acid catabolism. These reactions are known as the alanine-
glucose cycle detailed in the diagram below. This cycle allows the body to produce significant
amounts of glucose without requiring dietary carbohydrate.
Kidneys
The role of the kidneys in nitrogen metabolism cannot be overemphasized. The kidneys are
responsible for ridding the body of nitrogenous waste that accumulates in the blood plasma.
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Kidney glomeruli act as filters of blood plasma. All the constituents in plasma, with the
exception of plasma proteins, move into the filtrate. Essential nutrients such as sodium, amino
acids, and glucose are actively reabsorbed as the filtrate moves through the tubules. The cell
membranes are relatively impermeable to urea and uric acid, and are particularly impermeable
to creatinine, very little of which is normally reabsorbed.
About 80% of nitrogen is lost in the urine in the form of urea under normal conditions. In acidic
conditions, urinary urea nitrogen losses decrease and urinary excretion of ammonium ions
increases. In addition to urea and ammonia, usual nitrogenous wastes found in the urine include
creatinine and uric acid, with lesser or trace amounts of creatine (<100 mg/day), protein (<100
mg/day), amino acids (<700 mg/day) and hippuric acid (<100 mg/day). Hippuric acid results from
the conjugation of the amino acid glycine and benzoic acid, which is generated mostly in the
liver from the catabolism of aromatic compounds.
In addition to urinary nitrogen losses, nitrogen may be lost in the feces and sweat and with the
loss of hair and skin cells. These losses are referred to as insensible nitrogen.
Take-home message: Your kidneys are your blood’s waste filtering system. They filter out
waste products like nitrogen (in the form of urea), creatinine and ammonia so that they can be
excreted with your urine.
The brain
The brain has a high capacity for the active transport of amino acids, regardless of acidity level.
The transporters for some of the amino acids are almost fully saturated at normal plasma
concentrations; this is especially true of the transporters for the large neutral amino acids like
BCAAs and the aromatic amino acids: tyrosine, tryptophan and phenylalanine. These can
compete with each other for the common carriers. The effects of this competition become
especially apparent in conditions in which the blood concentrations of any of the BCAAs or
aromatic amino acids become elevated. For example, in liver disease, the concentrations of the
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aromatic amino acids exceed those of the BCAAs and cause increased uptake of the aromatic
amino acids by the brain. The elevations of amino acids in the brain alter brain function, causing
a variety of neurologic problems such as impaired brain development and altered behavior and
mental function, among other manifestations.
While it is clear that conditions like liver disease and inborn errors of amino acid metabolism
can alter the brain’s uptake of selected amino acids and cause neurological and behavioral
changes, such effects have not been demonstrated consistently in healthy individuals who
attempt to alter behavior by altering dietary intakes of nutrients. For example, ingestion of
carbohydrate (without ingestion of protein) has been shown to increase the brain’s uptake of
tryptophan and raise serotonin concentrations but does not always result in the expected
behavioral effects (such as feeling calm and relaxed) of elevated serotonin. In other words,
varying nutrient intakes, including carbohydrate and protein, by eating selected foods is thought
to have little effect on the brain’s serotonergic function in many circumstances.
Skeletal muscle
About 40% of the body’s protein is found in muscle tissue and skeletal muscle mass represents
about 43% of the average person’s body mass.
Protein synthesis and protein degradation are under independent controls but together account
for about 10% to 25% of resting energy expenditure in the general population. Rates of
synthesis can be high, as with protein accretion during growth. Alternately, protein degradation
can predominate, as during illness. Rates of protein turnover also vary among the body tissues,
as is evidenced in the more rapid turnover of visceral protein as compared with skeletal muscle.
Yet, because of its mass, muscle accounts for about 25% to 35% of all protein turnover in the
average body. Degradation of proteins occurs primarily by the action of proteases. The
constant degradation of proteins is of prime importance because it ensures a flux of amino acids
through the cytosol that can be used for cellular growth and/or maintenance.
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Uptake of amino acids by the skeletal muscles occurs following ingestion of a protein containing
meal. During this time, skeletal muscles typically experience net protein balance, i.e. protein
synthesis is greater than protein breakdown. In a post-absorptive state such as between meals
or in a fasting situation, the reverse is often true. Protein degradation predominates and amino
acids may be released into the blood for use by other tissues. While alanine is released in the
greatest concentration, other amino acids (including phenylalanine, methionine, lysine, arginine,
histidine, tyrosine, proline, tryptophan, threonine and glycine) are also released in lesser
quantities. Like other tissues, muscles preferentially catabolize some amino acids more than
others; six amino acids (aspartate, asparagine, glutamate, leucine, isoleucine, and valine) appear
to be catabolized to greater extents in the skeletal muscle than other tissues.
BCAA catabolism
Muscle, as well as the heart, kidneys, diaphragm, adipose tissue, and other organs (except, for
the most part, the liver) possess branched-chain aminotransferases for the transamination of all
three branched-chain amino acids. These tissues can thus break down BCAAs. Following
transamination, the α-keto acids of the branched-chain amino acids either remain within muscle
for further oxidation or may be transported (bound to albumin) in the blood to other tissues
(including the liver) for reamination or further catabolism.
Leucine’s metabolism also generates β-hydroxy β-methylbutyrate (HMB). HMB is important for
the production of β-hydroxy β-methylglutaryl (HMG)-CoA, a precursor for de novo (new)
cholesterol synthesis in muscle tissue.
Leucine is one of the few amino acids that can be completely oxidized in muscles for energy.
Leucine is oxidized in a manner similar to fatty acids and its oxidation results in the production
of acetyl-CoA and acetoacetate. Complete oxidation of leucine generates more ATP molecules
on a molar basis than complete oxidation of glucose. Leucine appears to be preferentially
oxidized during fasting situations. During fasting, leucine concentrations rise in the blood and
muscle, and the capacity of the muscle to break down the leucine increases concurrently.
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Indicators of muscle mass and muscle/protein catabolism
While proteolysis (protein breakdown) of muscle generates amino acids that are released into
the plasma for circulation to other tissues, changes in plasma amino acid concentrations do not
reflect changes in muscle mass. Instead, two previously mentioned compounds, creatinine and
3-methylhistidine, are used as indicators of existing muscle mass and muscle degradation,
respectively.
Urinary creatinine excretion is used to assess muscle mass because creatinine is the
degradation product of creatine, which constitutes a fairly standard proportion of muscle
(approximately 0.3–0.5% of muscle mass by weight). Urinary creatinine excretion reflects about
1.7% of the total creatine pool per day and is expressed per 24 hours, as a coefficient based
upon weight or height; however, because of variation in muscle creatine content, urinary
creatinine is not always an accurate indicator of muscle mass.
Take-home message: Creatinine levels correlate roughly with muscle mass. As such,
strength trained individuals often have elevated creatinine levels. Many doctors erroneously
interpret this as a sign of inflammation, kidney malfunction or muscle trauma.
The urinary excretion of 3-methylhistidine is used an indicator of muscle catabolism

(degradation). The amino acid histidine is found in high concentrations as 3-methylhistidine in
the muscle protein actin. Because 3-methylhistidine cannot be reused for protein synthesis
following protein degradation and is excreted in the urine, its urinary excretion can be
measured and serves as an indicator of muscle breakdown. A drawback to its use, though, is
that actin is not found only in muscle but appears to occur in other body tissues, including the
intestine and platelets, which have high turnover rates. Thus, urinary 3-methylhistidine
excretion also may represent an index of protein breakdown for many nonmuscle tissues in the
body.
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Take-home message: 3-methylhistidine levels in your urine are a rough marker of protein
catabolism in your body. You will encounter this marker in research on muscle damage and
protein losses.
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How much protein do you need?

Now that you understand what proteins are and what they do, we can get into more practical
matters.
Protein. It’s every bodybuilder’s favorite macronutrient and for good reason. Protein is
extremely essential, super satiating and amazingly anabolic. Protein is awesome… but you’re
consuming too much of it.
Like most myths, the belief that you should take in 1 g/lb of body weight has become so deeply
entrenched in the fitness world that its validity is rarely questioned. Strangely, very few people
think it’s a bit too accidental that the optimal amount of protein your body can assimilate in a
day is exactly 1 g/lb. 2.2g/kg doesn’t sound as right, does it? Of course, you’re doing this course
for its scientific merit, so let’s look at the literature on the effects of daily protein intake to find
out if 1 g/lb really is the optimal amount of protein intake for maximum muscle gains.
Studies on the optimal protein intake
All values in the bullet point list below are expressed as grams of protein per pound of body
weight per day. All of these studies controlled for energy intake, either based on individual
requirements or by setting energy intake to be equal in all experimental conditions, so that only
the proportion of protein in the diet varied between groups. If the studies were based on
unreliable methods such as nitrogen balance, a marker of lean body mass changes, they were
only included if they controlled for sweating and dietary adaptation periods. (As you’ve learned,
nitrogen is also lost in sweat, which can amount to significant quantities during exercise.
Nitrogen excretion is also affected by habitual intake with the result that increasing someone’s
protein intake can result in negative nitrogen balance for 2 weeks, hence the need for a dietary
adaptation period when studying a new protein intake for an individual.)
• Tarnopolsky et al. (1992) observed no differences in whole body protein synthesis or

indexes of lean body mass in strength athletes consuming either 0.64 g/lb or 1.1 g/lb over a 2
46
week period. Protein oxidation did increase in the high protein group, indicating a nutrient
overload.
• Walberg et al. (1988) found that 0.73 g/lb was sufficient to maintain positive nitrogen
balance in cutting weightlifters over a 7 day time period.
• Tarnopolsky et al. (1988) found that only 0.37 g/lb was required to maintain positive
nitrogen balance in elite bodybuilders (over 5 years of experience, possible previous use of
androgens) over a 10 day period. 0.45 g/lb was sufficient to maintain lean body mass in
bodybuilders over a 2 week period. The authors suggested that 0.55 g/lb was sufficient for
bodybuilders.
• Lemon et al. (1992) found no differences in muscle mass or strength gains in novice
bodybuilders consuming either 0.61 g/lb or 1.19 g/lb over a 4 week period. Based on nitrogen
balance data, the authors recommended 0.75 g/lb.
• Hoffman et al. (2006) found no differences in body composition, strength or resting
hormonal concentrations in strength athletes consuming either 0.77g/lb or >0.91g/lb over a 3
month period.
In addition to the above studies, here’s a full literature overview of studies that measured
muscle mass, strength, or some measure thereof in various groups that consumed a different
protein intake without any other differences between the groups, like different macronutrient
intakes or nutrient timing.
As you can see, in this type of controlled setting, no benefits of more than 0.73 g/lb/d (1.6
g/kg/d) protein have ever been found and many studies suggest that considerably less protein is
enough.
Study Subjects (n) Strength Hypertrophy Maximally

measure(s) measure(s) beneficial
protein
intake
(g/kg/d)
47
Iglay et al. Elderly Composite 1 Body composition (incl. <= 0.9

(2007) males & RM weight, whole body
females (36) protein mass & fat mass)
and skeletal muscle Akt
content
Tarnopolsky Bodybuilders/ N/M Lean body mass and <= 1.05

et al. (1988) cyclists/ Nitrogen balance
sedentary (18)
Hoffman et Young males Back squat & Body composition (incl. <= 1.74
al. (2006) (18) bench press 1 weight, lean body mass,
RM body fat percentage
Tarnopolsky Bodybuilders Bench Press & Body Composition <= 1.35

et al. (1992) (12) Back Squat 1 (incl. body weight, lean
RM body mass
& body fat percentage),
Nitrogen balance and
muscle CSA
Tarnapolsky Bodybuilders Bench press & Body composition (incl. <= 1.4
(1992) (13) back squat 1 weight, lean body mass
RM & fat percentage) and
Nitrogen balance
48
Antonio et al Strength- Bench press & Body composition (incl. <= 2.3
(2015) trained young back squat 1 weight, fat-free mass, fat (No lower
males & RM mass and body fat protein
females (48) percentage) group)
Hambre et Young males Quadricep Body composition (incl. => 1.4

al. (2012) (24) force & weight, fat-free mass and
hamstring force fat mass)
Snijder et al. Young males & Leg press, leg Body composition (incl. => 1.3
(2015) females (39) extension, weight and lean mass),
shoulder press Type I & II fibre size, leg
& chest press 1 volume and muscle CSA
RM
Campbell et Elderly male & N/M Body composition (incl. <= 0.8
al. (1995) females (12) weight, fat-free mass, fat
mass, muscle mass and
body fat percentage),
mean fibre area,
Nitrogen balance,
leucine kinetics & muscle
CSA
Antonio et Strength- N/M Body composition (incl. <= 2

al. (2014) trained young weight, fat-free mass, fat (No Lower
males (30) mass and body fat Protein
percentage) Group)
49
Pasiakos et al Young males & N/M Body composition (incl. <= 1.62
(2013) females (39) fat free mass, fat mass
and body fat
percentage), muscle
protein synthesis and
Nitrogen balance
Pikosky et al. Young males N/M Body composition (incl. => 0.9
(2008) (22) weight, fat free mass, fat (NBAL)
mass and body fat <= 0.9 (MPS)
percentage), Nitrogen
and whole body protein
turnover
Helms et al. Strength- Isometric thigh Body composition (incl. <= 1.6
(2015) trained young pull force weight, fat-free mass,
males (13) sum of skinfolds and
body fat percentage)
Walberg et Male N/M Body composition (incl. <= 0.8 (MPS)

al. (1988) bodybuilders weight, fat-free mass and => 0.8
(19) body fat percentage) and (NBAL)
Nitrogen balance
Antonio et Strength- Bench press 1 Body composition (incl. <= 2.6

al. (2016) trained young RM & bench weight, fat-free mass, fat (No Lower
males (12) press reps to mass and body fat Protein
Failure @ 60%1 percentage) Group)
RM
50
Kukuljan et Elderly males Bench press, lat Body composition (incl. <= 1.32
al. (2009) (180) pulldown and weight, fat-free mass, fat
leg press 1 RM mass and body fat
percentage) and
muscle CSA
Garthe et al. Elite athletes Bench press, Body composition (incl. <= 1.7
(2012) (39) bench pull & weight, fat-free mass, fat
squat 1 RM mass and body fat
percentage)
Rozenek et Healthy Young Bench press, lat Body composition (incl. <= 1.4
al., (2002) Men (73) pulldown & leg weight, fat-free mass, fat
press 1 RM mass and body fat
percentage)
and circumference
measurements
Mettler et al. Strength- Bench press 1 Body composition (incl. => 1.0
(2010) trained males RM weight, fat-free mass, fat
(20) mass and body fat
percentage)
Consolazio Young healthy N/M Body composition (incl. => 1.4

et al. (1975)* males (8) weight, body fat and dry
protein mass) and
Nitrogen balance
51
* - Study participants participated in calisthenics and aerobic type training, not resistance
training.
N/M - Not Measured
Note: studies not controlling for the timing of protein intake, like when the protein is provided
pre- or post-workout in the higher protein group(s) and not in the lower protein group(s), are
discussed in the course topic on nutrient timing.
Based on this research, many review papers have concluded 0.82 g/lb/d or 1.8 g/kg/d is the
upper limit at which protein intake benefits body composition (Phillips & Van Loon, 2011). This
is total daily protein intake from all sources expressed in relation to total bodyweight.
This recommendation often includes a double 95% confidence level, meaning they took the
highest mean intake at which benefits were still observed and then added two standard
deviations to that level to make absolutely sure all possible benefits from additional protein
intake are utilized. As such, this is already overdoing it and consuming 1 g/lb ‘to be safe’ doesn’t
make any sense. 0.82 g/lb is already very safe.
The picture below summarizes the literature. As you can see, 1.8g/kg (0.82g/lb) is the point at
which additional protein intake ceases to yield any benefits.
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“But I train harder!”
This is the common objection from bro bodybuilders who have been indoctrinated by decades
of absurd protein recommendations from their fellow bros on drugs and the supplement
industry. If you think you need more than 0.82 g/lb because you think you train harder than the
participants of all the above studies, think again. For example, Lemon et al. (1992) studied
bodybuilders training 1.5 hours per day, 6 days per week and still concluded 0.75 g/lb is the
highest intake at which body composition benefits could occur.
However, it is true that your protein intake logically depends on your training intensiveness. If
you’re not stimulating much protein turn-over and muscle growth with your training, you don’t
need as much protein for maximum muscle growth. So if you don’t train very hard, the protein
intake could be decreased. If you optimize someone your clients’ programs as you’ll learn in this
course though, this is not a relevant issue, because your clients should certainly be training hard
enough.
Conversely, if someone is performing a high volume of both aerobic and anaerobic training,
such as athletes, you may want to increase the protein intake. You can use the folllowing table
to assess how much additional protein is needed for athletes with extraordinary high training
volumes.
How protein needs change based on exercise type and training level according to Fielding & Parkington
(2002).
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Note that these recommendations are not additive. You always have to subtract the baseline
protein requirement, which is ~0.9 g/kg/d, from all numbers you add up. So if you add an hour
of moderate intensity jogging to daily strength training, you only have to add 0.1 g/kg/d. (In fact,
you probably don’t have to add any protein in this case due to the inference effect, but we’ll
discuss that later in the course topic on cardio.)
“But I’m more advanced than these geeks!”
Another frequently heard objection to the idea almost no one needs more than 0.82 g/lb/d of
protein is that people need more protein because they are more experienced than the studied
populations. Sorry, no cigar this time either. For example, Tarnopolsky et al. (1988) used elite
bodybuilders and found that less protein was needed than in novice bodybuilders.
In fact, the finding that the more experienced you are, the less protein you need, has been
replicated in several studies (Rennie & Tipton, 2000; Hartman, Moore & Phillips, 2006; Moore
et al., 2007). This makes perfect sense if you logically think about it.
In everyone there is both constant protein synthesis and breakdown. Resistance training causes
both breakdown and synthesis to increase, normally with a favorable balance towards synthesis.
As you progress in your training, the body becomes more efficient at stopping the breakdown
of protein resulting from training. This is one of the reasons you don’t get sore as much
anymore when you’re more advanced. Since less muscle protein now needs to be rebuilt, this
means less protein is subsequently needed for optimal growth.
Secondly, the more advanced you are, the less protein synthesis increases after training. As you
become more muscular and you get closer to your genetic limit, progressively less muscle is
built after training. This is very intuitive. The slower you can build muscle, the less protein is
needed for optimal growth. It wouldn’t make any sense if the body needed more protein to
build less muscle, especially considering that the body becomes more efficient at metabolizing
protein.
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As such, since more advanced trainees experience both less protein breakdown as well as less
protein synthesis, they need less protein than novice level trainees. However, since the extent
of this adaptation has not been studied in a practical training setting, it is advisable that
advanced trainees still consume 0.82 g/lb/d of protein.
What about when cutting? Or you’re really lean?
A final objection that is often heard is that these values may be true during ‘bulking’ or
maintenance periods, but ‘cutting’ in an energy deficit requires more protein to maintain muscle
mass. Several studies have found that this is not the case.
 Walberg et al. (1988) studied cutting weightlifters and they still found 0.73 g/lb was
sufficient to maintain lean body mass.
 A perhaps even more telling study is by Pikosky et al. in 2008. The researchers took a
group of endurance trained subjects and had them consume either 0.41 or 0.82 g/lb of
protein per day. They also added a thousand calories worth of training on top of their
regular exercise. So these guys were literally running on a 1000 calorie deficit while
drastically increasing their training volume. Talk about a catabolic state… Of course the
nitrogen balance in the low protein group plummeted. However, the protein intake of
0.82 g/lb in the other group completely protected the subjects from muscle loss.
Nitrogen balance, whole-body protein turnover and protein synthesis remained
unchanged.
 Jose Antonio et al. recently published a study in strength trained subjects in a deficit.
There was no effect of 3.4 vs. 2.3 g/kg protein on changes in lean body mass or strength.
(See the section on whether protein can make you fat below for more details on the research
by Antonio et al.)
 Prior research from Jose Antonio in strength trained individuals had found the same
thing: increasing protein from 1.9 to 4.4 g/kg did not affect body composition (including
fat loss) or strength. Now, technically these subjects weren’t supposed to be in a deficit,
but body fat percentage went down in both groups. This was non-significant, but since
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it’s yet another study finding no benefits of very high protein intakes, it’s worth taking
into account.
 Pasiakos et al. studied physically active individuals (3-4 x per week) in a 40% deficit.
Many subjects were military personnel. There was no difference in fat or muscle losses
between the group consuming 1.6 and the group consumign 2.4 g/kg protein. A major
strength of this study is that they also looked at muscle protein synthesis, anabolic
signaling, gene expression, nitrogen balance ánd resting metabolic rate. None of the
measures were different between the 2 protein intakes. Note: Eric pointed out the higher
protein group did have higher postprandial protein synthesis levels, but given the non-
significantly greater lean body mass loss in the moderate protein group, the relevance of this is
highly questionable. A limitation was that the strength training was pretty light: full-body
training 3x per week with 3 sets of 15 reps per muscle group. For beginners that’s not
terrible though. They also did a decent amount of cardio, which would further increase
protein needs. These subjects were not strength trained before the study, but that only
makes the results more convincing, because we know that strength training experience
decreases protein needs.
 Verdijk et al. (2009) found no difference in fat loss, muscle growth, nitrogen balance or
strength development between a group consuming 1.1 g/kg protein and a group adding
20 g casein to the diet. The subjects were elderly men who performed strength training
3x per week. They only trained their lower body though, which probably explains the
very low protein intake. In case you think their age is a confounder, it is, but most
research actually suggests the elderly have a higher protein requirement, if anything, due
to anabolic resistance, i.e. less efficient protein metabolism (discussed in further detail in
the course topic on strength training for the elderly).
 Campbell et al. found no difference in fat loss, strength development or muscle growth
in subjects consuming 1 – 1.2 g/kg protein compared to more than 1.2 g/kg. The
subjects were overweight men and women that performed full-body strength training
twice a week along with 1 weekly cardio session of 60 minutes. A notable strength of
this study was that it had 117 subjects and it lasted 9 months.
As you can see in the figure below, there was even a trend for better fat loss and more
muscle growth in the group consuming 1 – 1.2 g/kg protein than the group consuming
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more than 1.2 g/kg protein.
 Maltais et al. recently found no difference in lean body mass gains in strength training
elderly subjects consuming 1.1, 1.3 or 2.1 g/kg protein. They did 3 full-body strength
training sessions per week for 4 months. The 2.1 g/kg protein group did lose fat though
while the 1.3 g/kg group didn’t. This was probably in part because this group was
consuming soy-based protein while the 2.1 g/kg group was consuming dairy. The 1.1 g/kg
group actually lost only 200 grams less fat than the 2.1 g/kg group (0.9 kg vs. 1.1 kg fat),
but their fat loss did not reach statistical significance.
 Campbell & Meckling (2015) found that a protein intake of 0.9 g/kg (1.4 g/kg FFM)
resulted in significantly more fat loss and better muscle retention than a protein intake of
1.2 g/kg (1.9 g/kg FFM) with no difference in strength development. The subjects were
overweight women and they performed 3 full-body strength training sessions per week.
(See the lecture below on protein requirements for how an excessive protein intake can be
harmful.)
Moreover, we have good research in untrained individuals that shows increasing your protein
intake in a deficit does not protect your muscles [2], even when doubling your protein intake at
just 440 calories a day [3].One particularly nice study is the Hill et al. study that looked at 3
different protein intakes at weight maintenance and during weight loss. Protein content did not
affect body composition and this effect was not different between weight maintenance and
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weight loss phases. How much muscle you lose is instead determined by things like the rate of
weight loss, your training and your genetics.
We also know that increasing your protein intake has little effect on muscle atrophy during
disuse, like during bedrest. If extra protein can’t ward off that kind of catabolism, why would it
help during a cut?
Also, the supposed difference in nitrogen sparing effects of carbs and fat are negligible
(McCargar et al. 1989; Millward, 1989). Neither actually spares much protein though. Only
protein spares protein. The protein sparing idea seems to have come from a wrong
interpretation of the nitrogen balance literature showing more lean mass is lost in more severe
caloric deficits. A simple explanation for that finding is that the more total mass you lose, the
more lean mass you lose. No surprises there.
In fact, protein quality often increases in energy deficit, because more of the diet’s protein is
generally composed of high quality protein sources, like chicken breast, rather than, for
example, grains.
As such, there is simply no empirically substantiated reason to think we need more than 0.82
g/lb of protein per day when cutting. If anything, you could reason the body should be able to
use more protein during bulking periods, because more muscle is being built and a lot of other
nutrients are ingested that may enable more protein to be used.
Still, many people point to Eric Helms’s publication on protein requirements in negative energy
balance as the proof you need more protein when cutting, especially when you’re very lean,
than when bulking, so let’s review this paper in detail. Eric published a literature review and an
experimental study as part of his Master’s dissertation to research if we need more protein
when in a deficit. We start with the literature review, because that was the introduction for the
actual study in his dissertation.
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Note: If you’re not interested in the scientific details and don’t know who Eric Helms is, you can skip to
the conclusion of this section using the bookmarks on the left.
Here’s the abstract. (Note that Eric and Menno are friends, so Eric will be referred to by his
first name here.)
A systematic review of dietary protein during caloric restriction in resistance

trained lean athletes: a case for higher intakes.
by Eric Helms et al.
Introduction: Caloric restriction occurs when athletes attempt to reduce body fat or make
weight. There is evidence that protein needs increase when athletes restrict calories or have
low body fat. The aims of this review were to evaluate the effects of dietary protein on body
composition in energy-restricted resistance-trained athletes and to provide protein
recommendations for these athletes.
Methods: Database searches were performed from earliest record to July 2013 using the
terms protein, and intake, or diet, and weight, or train, or restrict, or energy, or strength, and
athlete. Studies (N = 6) needed to use adult (> 18 yrs), energy-restricted, resistance-trained (>
6 months) humans of lower body fat (males 23% and females 35%) performing resistance
training. Protein intake, fat free mass (FFM) and body fat had to be reported.
Results: Body fat percentage decreased (0.5-6.6%) in all study groups (N = 13) and FFM
decreased (0.3-2.7kg) in nine of 13. Six groups gained, did not lose, or lost nonsignificant
amounts of FFM. Five out of these six groups were among the highest in body fat, lowest in
caloric restriction, or underwent novel resistance training stimuli. However, the one group that
was not high in body fat that underwent substantial caloric restriction, without novel training
stimuli, consumed the highest protein intake out of all the groups in this review (2.5-2.6g/kg).
Conclusion: Protein needs for energy-restricted resistance-trained athletes are likely 2.3-
3.1g/kg of FFM scaled upwards with severity of caloric restriction and leanness.
The theory that you need more protein in a deficit

The introduction of Eric’s dissertation makes the case that theoretically, we need more protein
when energy intake is below maintenance levels. Eric et al. state:
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“When in negative energy balance, the anabolic response to protein is enhanced (Saudek
& Felig, 1976), which can be erroneously interpreted to mean that less protein is needed
during weight loss.”
Why would this be an erroneous interpretation? In an energy deficit, the body becomes more
conservative with protein. The more efficient the body is with protein, the less it logically
needs. This is similar to the finding that the more advanced you are as a strength trainee, the
less protein you need, because your body’s protein metabolism becomes more efficient. It’s
counter-intuitive, but it’s perfectly logical.
Of course, Eric et al. do offer an explanation:
“When supply is limited, efficiency is increased, indicating the body’s increased need for
protein in states of negative energy balance (Fielding & Parkington, 2002).”
As pointed out above, this statement is not logical. But there’s a reference! Yet if we actually
look at the research paper by Fielding & Parkington, it does not support increased protein
requirements in a deficit at all. In fact, based on their literature review, Fielding & Parkington
recommend less protein than this course does:
“The current recommended intakes of protein for strength is 1.6 to 1.7 g/kg.”
Fielding & Parkington actually go so far as to conclude that as athletes get more advanced, these
protein needs decrease to 1.2 g/kg/d even if the athlete is performing ‘habitual heavy resistance
training for 75 minutes a day’.
So purely based on the body’s physiology, we would actually expect protein needs to decrease
in a deficit, not increase.
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The evidence that you need more protein in a deficit

Eric et al. offer indirect evidence for increased protein needs during weight loss. They cite
research showing that certain endurance athletes and Olympic weightlifters during a training
camp were found to be in negative nitrogen balance, a measure of muscle loss, even while
consuming 2 g/kg protein. Aside from the methodological limitations of these old studies and
nitrogen balance, this again is a logically fallacious argument. Just because someone is in negative
nitrogen balance, does not mean their protein intake is insufficient. You need a second study
group consuming a different protein intake to assess the effect of protein intake on nitrogen
balance. Here’s an example if it doesn’t logically click why negative nitrogen balance does not
imply protein intake is insufficient.
John is an investment banker. He sleeps 2 hours a day, works 18 hours a day, is on the verge of
burn-out and trains 3 times a week doing P90x and CrossFit but stays far away from heavy
weights because the local PT said they are unsafe. He consumes 3 g/kg protein and cuts on a
50% deficit. He has 4% body fat. His diet consists of nothing but protein shakes and white rice
because he thinks that’s what all the pros do. He has low testosterone, but his doctor said he’s
too young for replacement therapy. John has negative nitrogen balance. Now, what do you
think is the cause of his negative nitrogen balance? If you answered ‘his protein intake is too
low’, please re-read this entire document. Twice.
As such, no prior research had demonstrated that protein needs are higher in a deficit. Let’s
move on to Eric et al.’s new evidence: their literature review.
The aim was to assess the weight loss success of the study groups as a function of their protein
intake. In other words, Eric et al. tried to find out if studies with higher protein intakes achieved
better body composition results than studies with lower protein intakes.
Before we get to the actual results, we can already identify a problem with the data. Only 6
studies were selected in the literature review. These studies differed in the rate of weight loss,
the body fat percentage of the subjects, the training the subjects did, how muscular the subjects
were and how much protein they consumed. So we have 5 relevant independent variables and
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just 6 studies. For a literature review to be somewhat conclusive, it is necessary to have at the
very least 3 studies for each combination of independent variables (as 2 data points can’t make
a trend). So even in the best case scenario, this review would be useful to generate an
hypothesis. (And that’s exactly what this was, since it was Eric’s disseration introduction for his
actual study, which is discussed below.)
In spite of these limitations, Eric et al. concluded, based on a narrative review, that this evidence
supported a trend towards greater protein requirements in a deficit. They didn’t perform any
analysis on the data, however, so Menno did one himself. He calculated the ratio of lean body
mass loss (‘muscle’) to total bodyweight loss. Researchers use these kind of p-ratios to assess
the effectiveness of the diet. After all, that’s what we want from a diet: high levels of fat loss
with minimal loss of muscle mass. This gives us a measure of diet effectiveness.
When looking at the relation between 2 variables, you plot it. That’s what has been done
below: the ratio of loss of fat-free mass to total bodyweight is on the vertical axis and protein
intake in grams per kilogram of total bodyweight a day is on the horizontal axis.
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Do you actually notice any relation between protein intake and weight loss success? Of course,
visual inspection is not very scientific, so Menno performed a regression analysis in SPSS. He got
a beta coefficient of -1.7, p = 0.36. That means there was no statistically significant relation
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between these 2 variables. Higher protein intakes were not associated with better muscle
retention during the studied diets.
Technical note: Menno averaged Umeda’s 2 groups as a single data point in the graph (not in the
analysis) due to the study’s data heterogeneity. Since the higher protein group actually had a worse p-
ratio here (11.5, literally off the chart), this only strengthens the lack of a relation.
In fact, the top 5 most successful study groups were by Garthe et al. and Mero et al. Their
protein intakes in order of best body composition results were 1.6, 1.6, 1.4, 1.4 and 1.5 g/kg
protein.
The argument from Eric et al. rests mostly on 2 studies. The first is Mettler et al.’s high protein
group with a protein intake of 2.3 g/kg. However, going by the data, their lean body mass to
total bodyweight loss ratio was 0.2. That was actually just the average in the dataset, so you
can’t say they were doing spectacularly well because of their high protein intake. They were just
doing average. Which fits all the other data: the extra protein wasn’t harmful, but it wasn’t
beneficial either.
The second study is Maestu et al. with a p-ratio of 0.1 and a protein intake of ~2.6 g/kg. So this
group performed slightly better than average and had a far higher than average protein intake
(the average protein intake was 1.6 g/kg). Was it the protein intake that caused them to be
relatively successful in preserving muscle mass during the diet? If you look at Maestu et al.’s
study, there’s another factor that just may be relevant: these guys were the only bodybuilder
group in the dataset. Not only that, they were national and international level competitors with
an average of 8 years of training experience. Training 600 minutes per week, increasing even to
~1000 minutes per week throughout the study. So we have international level bodybuilders
training over 16 hours a week and the reason their muscle retention was slightly more
successful than average in athletes and regular Joes must have been… their protein intake? You
can name a dozen other factors that makes someone an international bodybuilding champion
(genetics, a solid training program, an appropriate contest prep strategy etc.). Most importantly,
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since there was no relevant control group, this study can’t demonstrate a positive effect of a
high protein intake.
As such, the data presented by Eric et al. do not provide any evidence that you need more
protein when cutting than when bulking.
In the conclusion, Eric et al. derive their final recommended protein intake from Phillips and
Van Loon (2011). Yet those authors state “protein intakes in the range of 1.3–1.8 g /kg/day […]
will maximize muscle protein synthesis” with the lower end being for advanced trainees. The
abstract does list 1.8 – 2 g/kg may be advantageous when cutting to promote fat loss and the
1.8 – 2.7 g/d figure is given in the text, but those figures are not substantiated by any research
or formal analysis. This is what you call a ‘journalistic reference’, where newspapers are ok with
citing a figure if another newspaper did as well, even though the original source is unknown.
Basically, Eric came up with the hypothesis that you need more protein when cutting than when
bulking. However, the research review itself was just that: it generated an hypothesis. There is
no actual evidence in it that shows the hypothesis is true. So Eric then performed an
experimental study to test his hypothesis. (Eric is after all a true scientist and that can be said
with nothing but respect.)
Here’s the abstract.
High-protein, low-fat, short-term diet results in less stress and fatigue than
moderate-protein, moderate-fat diet during weight loss in male weightlifters: a
pilot study
by Eric Helms et al.
PURPOSE: Athletes risk performance and muscle loss when dieting. Strategies to prevent
losses are unclear. This study examined the effects of two diets on anthropometrics, strength,
and stress in athletes.
METHODS: This double-blind crossover pilot study began with 14 resistance-trained males
(20-43 yr) and incurred one dropout. Participants followed carbohydrate-matched, high-protein
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low-fat (HPLF) or moderate-protein moderate-fat (MPMF) diets of 60% habitual calories for 2
weeks. Protein intakes were 2.8g/kg and 1.6g/kg and mean fat intakes were 15.4% and 36.5% of
calories, respectively. Isometric midthigh pull (IMTP) and anthropometrics were measured at
baseline and completion. The Daily Analysis of Life Demands of Athletes (DALDA) and Profile
of Mood States (POMS) were completed daily. Outcomes were presented statistically as
probability of clinical benefit, triviality, or harm with effect sizes (ES) and qualitative
assessments.
RESULTS: Differences of effect between diets on IMTP and anthropometrics were likely or
almost certainly trivial, respectively. Worse than normal scores on DALDA part A, part B and
the part A “diet” item were likely more harmful (ES 0.32, 0.4 and 0.65, respectively) during
MPMF than HPLF. The POMS fatigue score was likely more harmful (ES 0.37) and the POMS
total mood disturbance score (TMDS) was possibly more harmful (ES 0.29) during MPMF than
HPLF.
CONCLUSIONS: For the 2 weeks observed, strength and anthropometric differences were
minimal while stress, fatigue, and diet-dissatisfaction were higher during MPMF. A HPLF diet
during short-term weight loss may be more effective at mitigating mood disturbance, fatigue,
diet dissatisfaction, and stress than a MPMF diet.
Noticed the effect on body composition? Even in a 40% deficit, 2.8 g/kg protein had no
beneficial effect on fat loss, muscle retention or strength compared to 1.6 g/kg. Magnitude-
based inference showed with 99% certainty that the greater protein intake had no effect on
bodyweight or muscle mass, measured in 3 different ways. For fat loss the certainty was 97%
and there was actually a 2% chance that the higher protein was harmful compared to a 1%
chance that it was beneficial.
What about the beneficial effect of protein on mood? Even if there’s no effect on your body
composition, doesn’t this research show extra protein still helps you psychologically?
Unfortunately, it doesn’t, or at least this study doesn’t show it. This study was confounded in a
major way by using maltodextrin powder as the protein placebo. That meant the medium
protein group was consuming 95 grams of pure sugar. When you’re in a 40% deficit and over
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20% of your diet is sugar, it’s not surprising you feel poorly. They were likely starving and the
major difference in hunger alone could easily explain the difference in mood.
Conclusion
No research shows that protein requirement increase in negative energy balance, not even in
very lean individuals. Theoretically, we may even need less, since the body’s protein metabolism
becomes more efficient in a deficit.
Should you set protein relative to bodyweight or lean body mass?
Theoretically, you would expect protein requirements to be influenced primarily by lean body
mass and less so by fat mass. However, the vast majority of research we have uses bodyweight,
so within the studied ranges of body fat percentage, this is suitable. The reason for this is likely
that while muscle mass requires more protein than fat mass, organ mass is still the most
protein consuming tissue in your body.
Plus, any greater predictive power from using lean body mass rather than total body mass is
easily lost as a result of the inaccuracy of trying to determine someone’s body fat percentage.
Unless, of course, you have access to a reliable body composition technique, like a recent DXA
scan of the person.
(Note: Measuring body fat percentage will be discussed further in the course topic on measuring
progress.)
In Menno’s re-analysis of Helms’s data, he also found that the optimal protein intake is not
better estimated by basing protein needs on lean body mass. The p-value increased from 0.36
to 0.43 and explanatory variance (R^2) went down from 8% to 6%. In other words, calculating
protein relative to lean body weight instead of total body weight did not strengthen the relation
between protein intake and weight loss success: there was still no significant trend towards
higher protein intakes being beneficial. Technically, the relation was actually slightly worse
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The little research we have that directly studied the effect of lean body mass on protein
utilization also found that lean body mass does not influence the dose-response of protein
intake for protein synthesis and anabolic signaling.
However, in obese individuals, 1.8 g/kg/d may be excessive. If you have a reasonably reliable
estimate of someone’s body fat percentage (discussed in the course topic on measuring progress),
you can use the lower bound of Eric Helms’s recommendation and prescribe 2.3 g/kg LBM/d if
this results in a lower protein recommendation than 1.8 g/kg/d.
Gender
Women don’t need as much protein as men, as we’ll get into in the course topic on gender
differences, but suffice to say for now that in the absence of data, 1.8 g/kg/d of protein is still
advisable for women.
PEDs
Performance enhancing drugs (PED, particularly androgenic-anabolic steroids (AAS), may

increase protein needs.
In itself AAS actually decrease your protein needs. AAS increase nitrogen retention: they make
your body’s protein metabolism more efficient.
However, this lower rate of protein breakdown is easily offset by far greater rates of protein
synthesis, assuming proper drug use, so you probably want to increase the protein intake in line
with the increased rate of muscle growth.
As a rule of thumb, you can estimate by which factor someone’s muscle growth will exceed
that of a natural lifter. Then take the logarithm of that – a common measure statisticians use to
characterize an unknown dose-response curve with diminishing returns – and use that plus one
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as a factor to multiple the protein intake. So for an individual who's expected to grow 5 times
as fast as a natural lifter, you prescribe a protein intake of 1.8 g/kg/d * (1 + log(5)) = 3 g/kg/d.
Note: PEDs will be discussed in detail in their own course topic.
When the bros may be right
If you look at all the factors that influence protein requirements and you combine all of those
just so that protein requirements are maximal, it’s conceivable that 1.8 g/kg/d may not suffice to
support maximal muscle growth. Specifically, for hard training male novices that start an intense
strength training program for the first time or that have a lot of muscle memory (more on that in
the course topic on understanding muscle growth), 1.8 g/kg/d protein may not be enough. With the
optimized program design you’ll learn in this course you’ll be able to achieve far greater rates of
muscle growth than what we see in the literature. In this scenario, the bro rule of thumb of 2.2
g/kg/d may be advisable and you could go as high as 2.7 g/kg for individuals with extraordinary
genetic potential (more on that in the course topic on customized program design).
The optimal protein intake is primarily a factor of how high protein synthesis can be pushed, so
in these cases exceptional protein intakes are required to keep up with the exceptional rates of
muscle growth that these individuals can achieve.
The one-gram-per-pound myth’s origin
Why is it that so many people say you need to consume 1 g/lb? Aside from the facts that there
don’t need to be any good reasons for people to believe something that is not true, that myths
tend to perpetuate themselves via conformism and tradition, and that the fitness industry is
flooded with myths, here are some plausible grounds for the ‘confusion’.
• People copy the dietary practices of pro bodybuilders on androgens. Steroids enable you to
assimilate far more protein than you’d normally could.
• The more is better heuristic. There are so many studies showing protein is good for you,
it’s hard not to think more of it is even better.
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• Supplement companies have an obvious financial incentive to make you want to believe you
need more protein than you really do. Supplement companies fund many of the major fitness
expos, they sponsor many pro bodybuilders and they have a stake in many of the popular
fitness magazines. They even sponsor a lot of the scientific research. Really, there’s not that
much else to sell in fitness, so if you follow the money, you often end up at a supplement
company.
• People can’t be bothered with decimals and just round up to the nearest convenient integer,
which so happens to be an easy to remember 1.
Often, more is better, but at some point it’s just too much.
Many people also don’t realize that the 1 g/lb myth only exists in bro bodybuilding culture. The
scientific community has long refuted the idea that strength trainees require more protein than
untrained individuals in the first place(!)
On a final note, there’s nothing inherently wrong or unhealthy about consuming more protein
than your body can use to build muscle. The excess will simply be used as energy. Even highly
excessive protein intakes consumed for months on end generally do not affect your health
negatively and that includes your kidney health.
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Take-home messages
• There is normally no advantage to consuming more than 0.82 g/lb (1.8 g/kg) per day of
protein. This already includes a mark-up, since most research finds no more benefits after 0.64
g/lb.
• Optimal protein intake decreases with training age, because your body becomes more
efficient at preventing protein breakdown resulting from training and less protein is needed for
the increasingly smaller amount of muscle that is built after each training session.
In-text section references
 Effect of protein intake on strength, body composition and endocrine changes in strength/power athletes.
Hoffman JR, Ratamess NA, Kang J, Falvo MJ, Faigenbaum AD. J Int Soc Sports Nutr. 2006 Dec 13;3:12-8.
 Tarnopolsky, M. A., Atkinson, S. A., MacDougall, J. D., Chesley, A., Phillips, S., & Schwarcz, H. P. (1992).
Evaluation of protein requirements for trained strength athletes. Journal of Applied Physiology, 73(5), 1986-
1995.
 Macronutrient content of a hypoenergy diet affects nitrogen retention and muscle function in weight
lifters. Walberg JL, Leidy MK, Sturgill DJ, Hinkle DE, Ritchey SJ, Sebolt DR. Int J Sports Med. 1988
Aug;9(4):261-6.
 Protein requirements and muscle mass/strength changes during intensive training in novice bodybuilders.
Lemon PW, Tarnopolsky MA, MacDougall JD, Atkinson SA. J Appl Physiol. 1992 Aug;73(2):767-75.
 Influence of protein intake and training status on nitrogen balance and lean body mass. Tarnopolsky MA,
MacDougall JD, Atkinson SA. J Appl Physiol. 1988 Jan;64(1):187-93.
 Dietary protein for athletes: From requirements to optimum adaptation. Phillips SM, Van Loon LJ. J Sports
Sci. 2011;29 Suppl 1:S29-38.
 Protein and amino acid metabolism during and after exercise and the effects of nutrition. Rennie MJ,
Tipton KD. Annu Rev Nutr. 2000;20:457-83.
 Hartman, J. W., Moore, D. R., & Phillips, S. M. (2006). Resistance training reduces whole-body protein
turnover and improves net protein retention in untrained young males. Applied Physiology, Nutrition and
Metabolism, 31, 557–564.
 Moore, D. R., Del Bel, N. C., Nizi, K. I., Hartman, J. W., Tang, J. E., Armstrong, D. et al. (2007). Resistance
training reduces fasted- and fed-state leucine turnover and increases dietary nitrogen retention in
previously untrained young men. Journal of Nutrition, 137, 985–991.
 Effects of exercise on dietary protein requirements. Lemon PW. Int J Sport Nutr. 1998 Dec;8(4):426-47.
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 Effects of high-calorie supplements on body composition and muscular strength following resistance
training. Rozenek R, Ward P, Long S, Garhammer J. J Sports Med Phys Fitness. 2002 Sep;42(3):340-7.
 Increased protein maintains nitrogen balance during exercise-induced energy deficit. Pikosky MA, Smith
TJ, Grediagin A, Castaneda-Sceppa C, Byerley L, Glickman EL, Young AJ. Med Sci Sports Exerc. 2008
Mar;40(3):505-12.
 Dietary carbohydrate-to-fat ratio: influence on whole-body nitrogen retention, substrate utilization, and
hormone response in healthy male subjects. McCargar LJ, Clandinin MT, Belcastro AN, Walker K. Am J
Clin Nutr. 1989 Jun;49(6):1169-78.
 Macronutrient Intakes as Determinants of Dietary Protein and Amino Acid Adequacy. Millward, DJ. J.
Nutr. June 1, 2004 vol. 134 no. 6 1588S-1596S.
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Protein intake & meal frequency
lecture
Protein intake & meal frequency
Note: meal frequency will be discussed in further detail in the course topic on nutrient timing.
Can your body absorb more than 20 g of protein per meal?
An additional reason that “your body can’t absorb more than 20 g protein per meal” is a myth
is that while protein synthesis has a very limited ceiling, protein breakdown can still be
decreased with higher protein intakes in each meal. Less protein breakdown has the same
benefit for total protein balance as more protein building.
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The leucine threshold & BCAAs

Total dietary protein intake is not all that matters. Ultimately, your body mainly cares about
total amino acid intake and there’s one amino acid in particular that’s important to understand,
because it affects nutrient timing and protein quality: leucine.
Leucine is the king of amino acids when it comes to muscle protein synthesis. This essential,
branched chain amino acid (BCAA) stimulates a protein - known as mTOR - that is highly
important for muscle growth. Leucine basically gives the signal to start muscle protein synthesis.
For the optimization of muscle protein synthesis, two things are thus important:
1. Consuming enough protein to provide building blocks (amino acids) to build muscle.
2. Optimizing the signal for muscle protein synthesis by leucine consumption.
Interestingly, the amount of leucine required to maximize the signal for muscle protein
synthesis is actually likely to exceed the amount that can be incorporated into muscle. So the
excess leucine will be oxidized and eliminated after it has done its signaling job. This ‘leucine
wasting’ is fine, as the goal is to maximize the signal for muscle protein synthesis and not only
to provide enough amino acids as building blocks.
So how much leucine do I need?
The individual leucine requirement depends on multiple factors, such as your daily activity level,
health, age and body composition. However, there is generally no need to overcomplicate this
or even calculate your leucine intake. You can generally assume that if you follow the
recommended protein intakes from this course and you include at least one source of complete
animal protein with each meal, like meat, dairy, fish or poultry, you will automatically consume
enough leucine and other BCAAs across the day.
It just takes about 10 grams of essential amino acids to make additional leucine supplementation
redundant. Interestingly, additional leucine can still modulate the anabolic signaling of the meal,
but the total amount of actual muscle protein that is built. Even post-workout in elderly men, a
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scenario where theoretically you would see maximal benefits of leucine supplementation,
adding leucine to just 0.16 g/kg whey protein (that’s just 13 g for a man weighing 176 lb / 80 kg)
failed to increase the amount of muscle protein that is built or decrease the amount of muscle
protein that is broken down.
Indeed, adding several grams of leucine to your meals for several weeks does not affect muscle
growth, muscle function or fat loss in the the elderly [2, 3] or competitive canooists and leucine
generally does not prevent muscle loss even in cases of severe muscle wasting.
In a study comparing several weight loss diet groups of elite wrestlers, BCAA supplementation
on top of a sufficient protein intake had no effect on muscle retention, subcutaneous fat loss,
aerobic performance, anaerobic performance or strength.
Another study by Spillane et al. (2012) found that "When combined with heavy resistance
training for 8 weeks, supplementation with 9 g/day of BCAA 30 min before and after exercise
had no preferential effects on body composition and muscle performance."
Basically the only positive ‘research’ on BCAA supplementation comes from the notorious
team of Scivation, who – surprise! – sell BCAA supplements.
Since BCAAs are generally so expensive that it’s still cheaper to get the BCAAs from protein
powder or even whole foods, I almost never recommend BCAA supplementation to my clients.
Vegeterians are a notable exception, as we’ll discuss below.
The leucine threshold
This is not the end of the story, however. Even if your total daily of leucine is sufficient, you
may still not be optimizing your muscle growth because of the leucine threshold. The leucine
threshold is a concept that has changed in definition over the years. Its original and most
relevant definition for us is that there is a threshold, i.e. a minimum amount, of leucine required
to start the protein synthesis process and cell growth. Below this threshold, the signal for
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muscle protein synthes is not strong enough and nothing or little happens. So if you consume a
meal or snack that does not have enough leucine, that protein may effectively be wasted.
For example, Mamerow et al. (2014) compared 2 protein distributions for total daily protein
balance. The EVEN group consumed an even protein distribution at each meal: ∼30 g at
breakfast, 30 g at lunch and 30 g at dinner.
The SKEW group consumed ∼10 g at breakfast, 15 g at lunch and 65 g at dinner. Total daily
protein was 1.2 g/kg/d (~90 g), which is generally seen as more than sufficient for non-strength
training individuals.
In spite of this, the EVEN group had a ~25% higher daily average protein synthesis rate (see
figure below). The large 65 g protein meal was unable to compensate for the 2 smaller meals
not stimulating protein synthesis enough. Since other research shows benefits of adding leucine
to these kind of very small protein servings, it is likely that the leucine threshold is responsible
for the greater protein synthesis in the EVEN group.
The leucine threshold is still not fully validated in research. It may be that human muscle protein
synthesis responds to leucine intake in a simple dose-dependant manner (see figure below)
without a true threshold. The leucine ‘threshold’ is sometimes arbitrarily defined as enough
leucine to spike protein synthesis by 50% above baseline, so it may be that the increase in
76
protein synthesis below that intake in other research just didn’t reach statistical significance
even though it did occur (see the course topic on DIY research for what stistical significance means).
Arnal et al. (2000) found that it doesn’t matter if you distribute your protein equally across the
day in 4 meals or have a 7% / 79% / 14% distribution across breakfast, lunch and dinner,
respectively. However, total protein intake was 1.67 g/kg/d and the subjects didn’t perform
strength training, which means the protein intake was likely excessive enough to negate all
benefits of finetuning the protein distribution across the day.
Figure modified from Breen & Phillips (2011)
To be safe, it is advisable to keep your protein intake in each meal above the leucine threshold.
Based on the research of Stuart Phillips, who is arguably the world’s leading researcher on
protein intake, that means you should consume a minimum of 0.3 g/kg protein in each meal and
each meal should have a complete amino acid profile. For protein sources high in leucine like
egg protein and dairy, you can probably get away with just 0.24 g/kg.
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In general, plant protein sources are lower in leucine (6-8% of total protein) than animal protein
sources (8-11%). As such, vegeterians will need to either supplement BCAAs or consume a
total daily protein intake of 2.4 g/kg in addition to making sure each meal contains a complete
amino acid profile and hits the leucine threshold.
The following table lists the exact amount of food you need to consume to hit the leucine
threshold. As you can see, in practice this is often not problematic.
78
% leucine per 100 g Amount needed to stimulate 50% MPS

Food source protein (0.5 g leucine)
Animal protein sources

Beef 8% 35 g
Chicken 7.5 % 35 g
Pork 8% 40 g
Fish 8% 40 g
Egg 9% 50 g
Dairy
Milk 8.5 % 200 ml
Yogurt 10 % 90 g
Cheese 11 % 20 g
Plant protein sources

Tofu 8% 70 g
Legumes 7-8 % 40 g
Nuts 6.5-7 % 35 g
Protein powders (ratio)

Milk protein 8.5 % 8g
Whey-casein-mix 9.5 % 6g
(1:1)
Whey-casein-mix 9% 6g
(1:4)
Egg white protein 8.5 % 7g
Pea-rice-mix (7:3) 7% 9g
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Protein quality
BCAA content is not the only determinant of protein quality. Several other factors determine
how useful certain proteins are to stimulate anabolism or prevent catabolism and ultimately
make your muscles bigger. Let’s review what determines protein quality.
Protein quality describes characteristics of a protein in relation to its ability to achieve defined
metabolic actions, like muscle growth. Traditionally, this has been discussed solely in the
context of a protein’s ability to provide specific patterns of amino acids to satisfy the demands
for synthesis of protein as measured by animal growth or, in humans, nitrogen balance.
However, protein is not only important for muscle growth. Protein also plays crucial roles for
signalling, in regulation of body composition and bone health, gastro-intestinal function and
bacterial flora, glucose homeostasis and satiety.
Measuring protein quality
One of the most well established measures of protein quality is the protein
digestibility-corrected amino acid score (PDCAAS), which is an improvement over several
previous measures of protein quality.
PDCAAS is based on the combination of an age-related amino acid reference pattern that is
representative of human requirements plus estimates of the digestibility of the protein. The
amount of potentially limiting amino acids in the test protein is compared with their respective
content in the appropriate reference pattern, identifying the single most limiting amino acid that
determines the amino acid score. See, your body doesn’t have a ‘total dietary protein'
requirement. It ultimately only cares how much it gets of each amino acid. For example, even if
you consume all the valine in the world and your total protein intake is easily adequate,
insufficient leucine consumption still limits muscle growth.
The current consensus is that meeting the minimum requirements for lysine, methionine, and
tryptophan, the most limiting amino acids in poor quality proteins, determines the amino acid
score and when nitrogen retention plateaus. At the plateau of nitrogen balance, any further
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increase in plasma amino acids would stimulate increased oxidation and elimination of the
excess amino acids, implying that protein quality above requirements does not matter.
The PDCAAS also takes digestibility of protein into account. If amino acids aren’t digested, your
body obviously can’t use them. As an example of poor digestibility, up to ~50% of raw egg’s
protein is not absorbed or digested in the small intestine [2], so you need about twice as much
protein from raw eggs as from cooked eggs.
The PDCAAS is a nice concept in theory, but it has several major limitations.
 It ignores the regulatory functions of amino acids before their oxidation. Leucine’s
influence on initiation of muscle protein synthesis is a prime example, as you’ve learned
in the section on the leucine threshold: leucine strongly influences whether other amino
acids are oxidized or used for protein synthesis.
 PDCAAS scores that are higher than 100% are truncated to 100% because the available
protein in food is first limited by digestibility. As the digestibility cannot exceed 100%, so
cannot PDCAAS. This leads to problems when calculating PDCAAS for mixed meals. In
mixed meals, it’s possible to have complementary aminio acid sources that together
create a high quality protein. Moreover, it may be possible for high amounts of essential
amino acids to compensate for a lack of non-essential amino acids.
 PDCAAS values are not adaptable for individuals consuming higher habitual protein
intakes. As protein intake increases, for example toward the upper half of the current
acceptable macronutrient density range, both the metabolic demands for amino acids
and the consequent fate of the dietary amino acids will become increasingly difficult to
predict in terms of generating a single reference amino acid pattern against which to
judge protein quality, especially across the entire life span and in all physiologic
conditions. Emerging experimental evidence suggests that there is an age related change
in the regulatory influence of essential amino acids on muscle protein synthesis that will
reduce the effectiveness of dietary protein to maintain muscle mass (anabolic resistance:
discussed in detail in the course topic on strength training for the elderly).
More importantly for us, the PDCAAS amino acid reference pattern may not apply very
well to lean, muscular strength trainees on a high protein diet.
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 The food matrix in which a protein is consumed can have significant impact on the
bioavailability of amino acid for metabolic needs. Digestive losses and structural changes
of amino acids are caused by numerous anti-nutritional factors in foods (discussed in
further detail in the topic on health science). Digestibility factors developed from rat bio-
assays may not appropriately correct for the range of anti-nutrient effects in the food
matrix, both naturally occurring and formed through processing methods. Although
heat, oxidation, and other treatments are carried out for consumer protection and
benefit, they can lead to formation of i.a. Maillard compounds, oxidized sulfur amino
acid, D-aminoacids, or cross-linked peptide chains, which limit amino acid bioavailability.
Basically, the PDCAAS does not take into account interaction effects between different amino
acids, interaction of the amino acids with other substances in the food in which they are
consumed and it is not created with lean, muscular strength trainees on a high protein diet in
mind.
As such, you can be skeptical of all the tables you see giving specific protein quality scores to
certain foods with, e.g. beef protein being a 90 and egg protein being a 95. These are highly
theoretical numbers. To see which protein sources are really best at stimulating muscle growth,
we need to turn to the empirical research.
Protein quality studies
The table below summarizes the literature on protein quality in relation to muscle growth. The
following sections go into the conclusions from these data.
Study Methodology Compared proteins Findings

Ingestion of whey - 18 young 10 g EAA in rapidly (i.e., Mixed MPS at rest
hydrolysate, casein, or resistance- whey hydrolysate and and after exercise
soy protein isolate: trained men soy) and slowly (i.e., was greater in whey >
effects on mixed - mixed MPS at micellar casein) digested soy > casein
muscle protein rest and after proteins
synthesis at rest and exercise
following resistance - time examined:
180 min
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exercise in young
men.
Greater stimulation of - 14 older men 20 g isolated micellar Myofibrillar MPS at

myofibrillar protein who were light- casein or pure whey rest and after
synthesis with to-moderately protein isolate exercise was greater
ingestion of whey active in the whey
protein isolate v. - myofibrillar supplementation
micellar casein at rest MPS at rest and group than casein
and after resistance after exercise group
exercise in elderly - time examined:
men 240 min
Whey protein - 48 untrained 20 g whey, casein, and Whey protein
stimulates older men casein hydrolysate stimulates
postprandial muscle - Postprandial postprandial muscle
protein accretion mixed muscle protein accretion
more effectively than protein fractional more effectively than
do casein and casein synthetic rates do casein and casein
hydrolysate in older (FSRs) hydrolysate
men - time examined:
360 min
Whey and casein - 17 untrained 0.3 g/kg LBM of casein Whey and casein
labeled with L-[1- young men or whey or placebo intake immediately
13C]leucine and - myofibrillar after resistance
muscle protein MPS after exercise results in an
synthesis: effect of exercise overall equal MPS
resistance exercise - time examined: response. There was
and protein ingestion 6 hours a trend toward a
higher but temporally
shorter MPS response
with whey compared
with a
more moderate but
prolonged MPS
response with casein.
Ingestion of casein and - 25 untrained 20 g casein or whey Acute ingestion of

whey proteins result young males and protein or placebo both whey and casein
in muscle anabolism females after exercise
after resistance - NBAL resulted in similar
exercise - 300 min increases in muscle
protein net balance,
resulting in net
muscle protein
synthesis despite
different patterns of
83
blood amino acid

responses (greater
oxidation of leucine
when whey
protein was ingested).
The Effects of Pre- - 16 female 24 g whey or casein Both groups
and Post-Exercise basketball player protein immediately experienced
Whey vs. Casein starting a pre- and post-exercise significant
Protein Consumption resistance (4 times a week for improvements in
on Body Composition training program eight weeks) body composition,
and Performance - body strength, and
Measures in Collegiate composition, anaerobic
Female Athletes. strength and performance. There
performance was no significant
- 8 weeks difference in any
markers between
whey and casein
Whey protein - 63 non- Whey supplement (22 Daily
supplementation resistance- g carbs + 22 g protein) supplementation with
during resistance trained men and or whey was more
training augments lean women soy supplement (24.5 g effective than soy
body mass - LBM gains carbs + 20 g protein) protein or isocaloric
- 9 months or carbohydrate carbohydrate control
resistance supplement (45 g) treatment conditions
training in promoting gains in
lean body mass
The effects of 8 weeks - 24 resistance 48 g of rice or whey Both whey and rice
of whey or rice trained males protein isolate protein isolate
protein - body (isocaloric and administration post
supplementation on composition, isonitrogenous) post- resistance exercise
body composition and strength and workout improved indices of
exercise performance. recover body composition and
- 8 weeks exercise
performance;
however, there were
no differences
between the two
groups.
Consumption of fluid - 8 resistance isonitrogenous, Milk-based proteins
skim milk promotes trained men isoenergetic, and promote muscle
greater muscle - NBAL and macronutrient-matched protein accretion to a
protein accretion after fractional soy or milk beverages greater extent than
resistance exercise synthetic rate of containing 18 g protein do soy-based proteins
than does muscle protein when consumed after
consumption of an - time examined: resistance exercise
isonitrogenous and 180 min
84
isoenergetic soy-
protein beverage
Consumption of fat- - 56 resistance fat-free milk; fat-freeNo strength

free fluid milk after trained men soy protein that was differences between
resistance exercise - Muscle fiber isoenergetic, groups
promotes greater lean size, maximal isonitrogenous, and Type II muscle fiber
mass accretion than strength, and macronutrient ratio area increased in all
does consumption of body matched to Milk (17.5 ggroups, but more in
soy or carbohydrate composition protein, 25.7 g the Milk group
in young, novice, male - 12 weeks carbohydrates, 0.4 g Type I muscle fiber
weightlifters fat); or maltodextrin area increased after
that was isoenergetic training only in the
with Milk and Soy Milk and Soy groups
LBM increased in all
groups, with a greater
increase in the Milk
group
Differences in - 12 30 g protein from beef Both milk and beef
postprandial protein recreationally or skim milk after a ingestion augment the
handling after beef active young men single bout of resistance postexercise
compared with milk - myofibrillar exercise myofibrillar protein
ingestion during MPS synthetic response in
postexercise recovery: - time examined: young men. Milk
a randomized 5 hours ingestion increased
controlled trial. myofibrillar protein
synthesis rates to a
greater extent during
the 0- to 2-h
postexercise phase.
However, the
increase in
myofibrillar protein
synthesis rates did
not differ between
milk and beef
ingestion during the
entire 0- to 5-h
postexercise phase.
Protein blend - 19 young, ∼18 g whey protein (1.9 Resting muscle
ingestion following recreationally g leucine, 8.9 g EAAs) protein synthesis did
resistance exercise active male and or ∼19 g of protein not differ between
promotes human female blend (1.8 g leucine, 8.7 groups. Postexercise
muscle protein - fractional g EAAs) composed of FSR increased
synthesis synthetic rate 50% protein from equivalently in both
(FSR) following sodium caseinate, 25% groups during the
85
resistance whey isolate and 25% early period,

exercise soy protein isolate however, FSR
- time examined: remained elevated
180 min only in the protein
blend group during
the late period.
Soy-dairy protein - 16 young, 0.305 g/kg whey or Myofibrillar MPS was
blend and whey recreationally 0.337 g/kg protein blend not different between
protein ingestion after active adults composed of 50% the groups at rest and
resistance exercise - myofibrillar protein from sodium increased above
increases amino acid MPS at rest and caseinate, 25% whey resting values post-
transport and after acute isolate and 25% soy exercise with no
transporter resistance protein isolate difference between
expression in human exercise Leucine and EAA the groups.
skeletal muscle - time examined: content was matched
240 min for both conditions.
Pea proteins oral - 161 moderately - 25 g whey concentrate No strength

supplementation physically active or 24 g pea isolate differences between
promotes muscle men containing ~ 14-15 g groups.
thickness gains during - muscle protein or placebo No difference
resistance training: a thickness and (maltodextrin) between increases in
double-blind, strength thickness between
randomized, Placebo- - 12 weeks both protein groups.
controlled clinical trial resistance Pea protein group
vs. Whey protein training protocol experienced
significantly greater
gains in muscle
thickness than the
placebo group.
Ingestion of Wheat - 60 elderly men 35 g wheat protein, 35 g Wheat vs. wheat
Protein Increases In - myofibrillar wheat protein hydrolysate: no
Vivo Muscle Protein MPS hydrolysate, 35 g significant difference
Synthesis Rates in - 240 min micellar casein, 35 g in MPS
Healthy Older Men in whey protein, or 60 g 35 g whey and wheat
a Randomized Trial wheat protein hydrolysate did not
hydrolysate significantly stimulate
MPS
35 g casein stimulated
MPS to a significantly
higher extent than 35
g wheat
MPS significantly
increased after
ingestion of 60 g
wheat protein
86
Effects of Hydrolyzed - 56 resistance- 30 g/serving carbohydra, Similar increases in

Whey versus Other trained men e placebo or strength and weight
Whey Protein - strength and 30 g/serving protein between all groups,
Supplements on the body from either however, greater fat
Physiological composition (a) 80% whey protein loss in the group that
Response to 8 Weeks - 8 weeks concentrate (WPC), consumed extensively
of Resistance Exercise resistance (b) high-lactoferrin- hydrolyzed WPC
in College-Aged Males training containing WPC, or
(c) extensively
hydrolyzed WPC.
All subjects consumed 2
servings per day; pre-
and post-workout or
between meals on
nontraining days
The effects of protein - 36 resistance- 48 g per day No strength
and amino acid trained males carbohydrate placebo, differences between
supplementation on - body 40 g/d whey + 8 g/d the group
performance and composition, casein, or Greatest increase in
training adaptations strength, 40 g/d of whey protein fat-free mass in the
during ten weeks of muscular + 3 g/d BCAA + 5 g/d whey + casein group
resistance training endurance, and L-glutamine (WBG)
anaerobic
capacity
- 10 weeks
resistance
training
Protein - 68 22 g protein blend (25% No difference in
Supplementation Has recreationally soy, strength gains
Minimal Effects on active young men 25% whey, and 50% between the groups
Muscle Adaptations - strength, thigh sodium caseinate), whey There was a trend
during Resistance muscle thickness, protein isolate, or an towards higher LBM
Exercise Training in myofiber cross- isocaloric maltodextrin increase in the
Young Men: A sectional area, (carbohydrate) placebo protein blend group,
Double-Blind and lean body however, changes
Randomized Clinical mass between whey and
Trial - 12 weeks placebo group did not
differ
The Effect of - 26 elderly, protein-rich supplement There was a
Resistance Training sedentary men 12 g of protein, 7 g of significant increase in
and Different Sources - strength and essential amino acids fat-free mass and
of Postexercise body from milk, soy, or rice strength in all groups,
Protein composition milk (nonprotein with no differences
Supplementation on -12 weeks control). between groups.
Muscle Mass and
Physical Capacity in
87
Sarcopenic Elderly
Men
Post Exercise - 45 untrained Milk (21 g Consumption of all

Ingestion of Plant vs women carbohydrates, 21 g protein supplements
Animal Protein - strength protein) resulted in significant
Enhances Exercise - 10 weeks Whey (10 g C, 34 g P) increases in arm and
Performance Hemp (18 g C, 21 g P) leg muscle strength,
Outcomes Similarly Placebo (49 g C) with an insignificant
effect on body fat
percentage.
Changes in muscle
hypertrophy were
also seen regardless
of drink consumed.
Slow and fast dietary - 16 young men 30 g whey (WP) or 40- Whole body protein
proteins differently - protein 43 g casein (CAS) breakdown was
modulate postprandial breakdown, inhibited by 34% after
protein accretion protein synthesis CAS ingestion but not
and net leucine after WP ingestion.
balance Postprandial protein
- time examined: synthesis was
420 min stimulated by 68%
with the WP meal
and to a lesser extent
(+31%) with the CAS
meal.
Net leucine balance
over the 7 h after the
meal was more
positive with CAS
than with WP.
Hydrolyzed dietary - 21 healthy Intact or hydrolysed Gut cells preferably

casein as compared adults casein containing 320 use the amino acids
with the intact protein - nitrogen mmol nitrogen from hydrolysed
reduces postprandial kinetics casein, thus, less
peripheral, but not - 8 hours amino acids are taken
whole-body, uptake of up into the blood
nitrogen in humans stream to be available
for MPS.
Effects of an - 19 elderly, Either habitual Consumption of a
omnivorous diet overweight or omnivorous diets, which meat-containing diet
compared with a moderately provided approximately contributed to
lactoovovegetarian obese men 50% of total dietary greater gains in fat-
88
diet on resistance- - strength and protein from meat free mass and skeletal
training-induced lean body mass sources (beef, poultry, muscle mass. There
changes in body - 12 weeks with pork, and fish) or was no difference in
composition and resistance ovolactovegetarian diet strength gains
skeletal muscle in training protocol between the groups.
older men.
Effect of protein - 21 elderly men After consuming a Increases in muscle

source on resistive- - strength and lactoovovegetarian strength and size
training-induced lean body mass (LOV) diet for 2 wk, were not influenced
changes in body - 4 weeks with subjects were randomly by the predominant
composition and resistance assigned to either source of protein
muscle size in older training protocol consume a beef- consumed by older
men containing (BC) diet or men with adequate
to continue the LOV total protein intake
diet. The BC diet
included 0.6 g
protein/kg/d from beef
and the LOV diet
included 0.6 g
protein/kg/d from
textured vegetable
protein (soy) sources.
Compared with casein - 23 men and ~ 23 g micellar caseins, Despite MSPI’s high
or total milk protein, women milk soluble protein PDCAAS, ensures a
digestion of milk - postprandial isolate (MSPI), and total rate of amino acid
soluble proteins is too utilization of milk protein delivery that is too
rapid to sustain the dietary nitrogen rapid to sustain the
anabolic postprandial - 8 hours anabolic requirement
amino acid during the
requirement postprandial period.
Milk proteins had the
best nutritional
quality, which
suggested a
synergistic effect
between soluble
proteins and caseins
Coingestion of whey - 18 healthy 1) [¹⁵N]WP, [¹³C]Cas, Early after meal
protein and casein in a participants and lactose; ingestion, amino acid
mixed meal: - rate of 2) [¹³C]WP, [¹⁵N]Cas, absorption and
demonstration of a appearance and lactose; retention across the
more sustained across the 3) lactose alone. leg were similar for
anabolic effect of splanchnic bed WP and Cas, but as
casein and the rate of WP – whey protein rates for WP waned,
89
disappearance Cas - casein absorption and

across the leg assimilation into
- time measured: skeletal muscle were
420 min better retained for
Cas
Protein feeding - 41 obese men 22.6% of energy as No significant
pattern, casein and women protein in the form of differences in hunger
feeding, or milk- - body milk-soluble protein perception between
soluble protein feeding composition and (MSP, = whey) or groups. In the last
did not change the hunger caseins, week, due to a higher
evolution of body - 6-wk restricted inhibition of protein
composition during a energy diet degradation and
short-term weight loss despite a lower
program. stimulation of protein
synthesis,
postprandial balance
between whole body
protein synthesis and
degradation was
better with caseins
than with MSP. It
seems likely that the
positive effect of
caseins on protein
balance occurred only
at the end of the
experiment.
Effect of a Hypocaloric - 38 overweight Three conditions: The casein group
Diet, Increased men 25 % of consumed experienced
Protein Intake and - strength and calories from whey or significantly greater
Resistance Training on body casein or no protein gains in strength and
Lean Mass Gains and composition supplementation and muscle mass and
Fat Mass Loss in - 12 week weight not exercise significantly greater
Overweight Police loss (80% deficit) (hypocaloric diet only) decrease in fat mass
Officers + resistance than the whey and
program diet-only group.
Whey protein - 15 elderly 15 g of whey protein or Phenylalanine balance
ingestion in elderly participants 6.72 g of EAA or 7.57 g improved in the whey
persons results in - leg of nonessential amino group, but not in the
greater muscle phenylalanine acids (NEAA) EAA group or NEAA
protein accrual than balance (index of group
ingestion of its muscle protein
constituent essential accrual)
amino acid content - 3.5 hours
90
Milk ingestion - 24 untrained 237 g of fat-free milk, Milk ingestions

stimulates net muscle participants 237 g of whole milk, or increases NBAL. A
protein synthesis - NBAL 393 g of fat-free milk significantly higher
following resistance - 240 min isocaloric with the WM uptake for threonine
exercise was observed in the
whole milk suggesting
that whole milk may
have increased
utilization of available
amino acids for
protein synthesis.
Postexercise net - 6 not trained Subjects consumed in Net balance was
protein synthesis in subjects random order 1 liter of similar for MAA and
human muscle from - net protein 1) a mixed amino acid EAA. However, more
orally administered balance after (40 g) solution (MAA = amino acids were
amino acids. exercise EAA + NEAA), taken up into the leg
- time examined: 2) an essential amino from the MAA than
225 min acid (40 g) solution EAA.
(EAA), and
3) a placebo solution
Leucine - 40 men Ingestion of A low-protein (6.25
supplementation of a - MPS after - 25 g whey protein (3.0 g) mixed
low-protein mixed exercise g leucine), macronutrient
macronutrient - time examined: - 6.25 g whey protein beverage can be as
beverage enhances 4.5 h (0.75g leucine), effective as a high-
myofibrillar protein - 6.25 g whey protein protein dose (25 g) at
synthesis in young supplemented with stimulating increased
men: a double-blind, leucine to 3.0 g total MPS rates when
randomized trial leucine, supplemented with a
- 6.25 g whey protein high (5.0 g total
supplemented with leucine) amount of
leucine to 5.0 g total leucine. However,
leucine, or enriching a low
- 6.25 g whey protein protein mixture with
supplemented with BCAA leads to
leucine, isoleucine, and inferior results
valine to 5.0 g total compared with
leucine enriching with leucine
alone, as valine and
isoleucine compete
for the same intestinal
transporters and can
impede leucine
absorption.
Ingestion of a protein - 10 elderly men 35 g bolus intact casein Ingestion of a protein
hydrolysate is or hydrolyzed casein hydrolysate, as
91
accompanied by an - muscle protein opposed to its intact

accelerated in vivo fractional protein, accelerates
digestion and synthetic rate protein digestion and
absorption rate when - 6 hours absorption from the
compared with its gut, augments
intact protein postprandial amino
acid availability, and
tends to increase the
incorporation rate of
dietary amino acids
into skeletal muscle
protein.
Ingestion of Casein in - 32 healthy 25 g casein dissolved in Casein ingestion in a
a Milk Matrix older men bovine milk serum or milk matrix delays
Modulates Dietary - MPS water protein digestion and
Protein Digestion and - time examined: absorption but does
Absorption Kinetics 300 min not modulate
but Does Not postprandial muscle
Modulate Postprandial protein synthesis
Muscle Protein when compared to
Synthesis in Older the ingestion of
Men micellar casein only in
healthy older men
Minced beef is more - 10 older men 135 g beef steak or Minced beef is more
rapidly digested and - whole-body minced beef rapidly digested and
absorbed than beef protein balance absorbed than beef
steak, resulting in and skeletal steak, which results in
greater postprandial muscle protein increased amino acid
protein retention in synthesis rates availability and greater
older men. - time examined: postprandial protein
6 hours retention. However,
this does not result in
greater postprandial
muscle protein
synthesis rates.
Fast vs. slow protein
Speed of absorption is often marketed by supplement companies as a major advantage of a

protein source. You consume it and BOOM, all that protein is right there in your muscles
instantly. In reality, rapid aborption is not purely an advantage.
92
The 2 prototypes of slow and fast protein are the 2 major protein fractions of cow milk: casein
and whey protein. Other than differing significantly in amino acid composition, whey protein is
soluble in milk, so it remains in liquid form and passes quickly through your stomach, hence
having a fast digestion and being named a ‘fast protein’. Caseins, on the other hand, are not milk
soluble and will clot in your stomach, delaying the amino acid delivery to your intestines. Hence
casein is a ‘slow protein’.
While it’s true that faster proteins result in higher spikes of muscle protein synthesis (MPS),
they don’t stimulate MPS for as long as slower proteins do. So while there are significant
differences in the concentration of amino acids in the blood over time (see illustration below),
the total amount of net protein build-up is ultimately generally similar for whey and casein
proteins. Most studies that suggested whey is superior to casein simply did not measure MPS
for a sufficient duration, which biased the results in favor of faster proteins.
Mean (±SD) changes from baseline in serum total amino acid (AA) concentrations in the subjects after
the ingestion of ~23 g protein from total milk protein (TMP), micellar casein (MC) or milk soluble
protein isolate (MSPI). Source
93
Secondly, as you learned in the course topic on protein digestion and absorption, fast
absorption can result in greater protein oxidation rates, particularly leucine oxidation. Whey
protein in isolation actually seems to be digested too rapidly in humans to sustain postprandial
amino acid requirements, resulting in fewer of the amino acids actually being incorporated into
muscle tissue compared to casein or whole milk protein.
Rapid digestion can also decrease the uptake of amino acids by your muscles. Hydrolyzed
casein, marketed for its rapid and easy absorption, is in fact preferentially used by your gut cells,
leaving less amino acids available for your muscles.
Note: A study in elderly individuals found the opposite, but this may have been related to the study
period being 6 rather than 8 hours, thus biasing the results in favor of the faster protein. Also, elderly
individuals may have more to gain from fast proteins in low quantities because of their anabolic
resistance, which will be discussed in the course topic on strength training for the elderly.
Thirdly, while whey protein is superior to casein production in stimulating peak MPS, casein
protein is more effective at reducing protein breakdown. In fact, whey protein doesn’t prevent
catabolism much at all. As a result, when looking at total muscle protein balance, not just MPS,
over a time period of 7 hours, casein results in greater net muscle protein balance. In long term
studies, casein protein has also been shown to result in higher whole body protein balance
levels, greater strength development, more fat loss and more muscle growth than whey
protein.
In conclusion, the idea that faster proteins, like hydrolyzed proteins and whey, are better for
bodybuilding than slower proteins, like casein, is nothing but supplement company marketing.
The truth is often exactly the opposite for young individuals.
The food matrix
The digestion and absorption of amino acids is affected not just by the structure of the protein
but also by its interaction with the food matrix in which the protein is consumed.
94
There are various mechanisms by which other components in dietary protein than just the
BCAAs stimulate protein balance. As Hulmi et al. (2010) noted based on Kimball (2007): "via
the PEPT-1 cotransporters' high capacity, low specificity rate of transport, and an apparent
increased transport affinity for L-valine bound peptides [...] the bound form of an essential
amino acid (EAA) may be more efficiently utilized than when delivered in its free-form."
Due to the synergy between amino acids, pure EAA and BCAA mixtures don’t stimulate
protein balance as well as complete amino acid profiles.
The same applies on levels higher up in the food matrix.
 We already saw that micellar casein and whole milk proteins are better at stimulating
cumulative muscle protein balance over time than whey.
 Blends of protein tend to promote greater protein balance than pure whey, even when
the blend contains soy, which in isolation performs very poorly (discussed below).
 One level further up, we see that whole milk tends to stimulate greater protein
synthesis than skimmed milk, indicating that other substances than just the protein in
milk promote MPS.
As such, the evidence and evolutionary theory suggest that good old actual milk consumption
rather than processed protein powders is the most reliable and effective way to stimulate both
protein synthesis as well as reduce protein breakdown. Nature provided you with a package
deal of whey and casein in highly absorable form: take advantage of it.
In contrast to milk, many plants have a food matrix that negatively affects protein quality. Plant
foods contain many anti-nutrients as part of their evolutionary defence against being eaten.
These substances in the food matrix of plants make their protein difficult to use for the human
body. Wheat and soy in particular are poorly capable of stimulating muscle protein balance,
even when they provide high amounts of essential amino acids. In general, vegetarian diets are
inferior to omnivorous diets from a bodybuilding perspective and require more meticulous
95
management of protein quality to compensate for this. Rice, hemp and pea protein seem to
provide relatively usable amino acids. Taking their amino acid composition into account, this
means it’s advisable for vegetarians to choose pea protein powder or a blend of pea and rice
protein powders if they’re going to supplement protein powder.
(Note: anti-nutrients will be discussed in greater detail in the topic on health science.)
Conclusion
lecture
Protein quality
The following table provides a guideline of protein quality categories. In short, whole foods
generally reign supreme as anabolic fuel for bodybuilders. If you do decide to supplement
protein for whatever reason, a protein blend is best, e.g. whole milk protein, a mix of whey
with 50-80% micellar casein, a pea-rice protein blend (for vegetarians).
High quality Moderate quality Low quality

Whole dairy Pea protein Soy protein
Meats Rice protein Other vegetarian protein sources
Fish Hemp protein
Boiled/baked eggs Whey protein
Poultry Hydrolyzed protein
Milk protein
Casein protein
96
Can protein make you fat?

In 2015, Antonio et al. conducted a study showing that a group consuming 4.4 g/kg/d of protein
– yes, that’s a ton of protein – did not gain fat compared to a control group that was consuming
1.8 g/kg/d of protein even though the higher protein group was consuming more calories. The
study design was such that both groups were instructed to maintain their baseline eating habits.
The high protein group was simply instructed to consume more protein.
The authors stated: “This is the first interventional study to demonstrate that consuming a
hypercaloric high protein diet does not result in an increase in body fat.” And boom, the
internet concluded that a surplus of protein isn’t stored as fat.
Antonio et al. followed this up with another study in 2015 comparing protein intakes of 2.3 vs.
3.4 g/kg/day. This time the higher protein group lost more fat.
And in 2016 Antonio et al. followed up with another similar study on 2.6 vs 3.3 g/kg/d.
Sponsorship bias?
Before we go into detail on these studies, a disclaimer is warranted. The first study was
sponsored by MusclePharm, who are notorious for their sponsored research practices, and
Adept Nutrition (Europa Sports Products brand). The second and third studies did not disclaim a
competing interest, but the protein powder for the second study was provided by Essentia
Metabolic Proteins.
All studies were published in JISSN, a journal that is notorious for publishing many questionable
studies sponsored by the supplement industry. Unlike many more reputable journals, JISSN
charges a hefty sum of about $2000 to publish articles. This means JISSN has a strong financial
incentive to publish whatever’s requested and to publish papers that will attract a lot of
popularity.
97
Peer review?
It is also notable that the first study passed peer review – a process that often takes month – in
a single day. The second and third studies’ peer review processes took just a week.
Personally though, while I think peer-review was lacking for these studies, after having met Jose
Antonio, I do not believe that any data frauding took place during these studies.
Replication failure
However, a subsequent study published by a different lab in a different journal failed to replicate
the findings of Antonio’s studies.
Spillane & Willoughby (2016) also had a group of trained men overeat for 8 weeks, this time
either on protein and carbohydrates with a little fat (HPC) or purely on carbohydrates (HC).
So the groups ate the same amount of calories, but the main difference was that the HPC group
ate more protein and the HC group ate more carbs. The protein intakes relative to baseline
bodyweight were 2.5 g/kg/d for HPC and only 1.1 g/kg/d for HC. (See table below for their
exact macros if you’re interested.)
98
The result: both groups gained the same amount of fat during the bulk.
By the way, there was also no difference in the training volume load achieved by the groups,
how much lean body mass they gained, how much muscle protein content they gained, how
much upper or lower body strength they gained or the changes in their muscle DNA
concentrations and their anabolic hormone levels.
However, there was a trend for increased lower body strength in the 2.5 g/kg/d protein
group. This group also gained more body water, which may be indicative of muscle gain,
though that’s a stretch given that they did not gain more muscle protein content. So this
study shows that there are probably benefits of more protein than just 1.1 g/kg/d in trained
men during a serious bulk, but even at this protein intake, the benefits of more protein are
questionable.
So why did the studies from Antonios’s lab not find fat gain, while Spillane & Willoughby did?
Rather than accept these findings at face-value, a wise person would ask: what’s the mechanism
whereby a higher protein diet would result in less fat storage or more fat loss?
The mechanisms of weight loss by high protein diets
First, you have to realize one very important point about how your body reacts to your diet:
your body doesn’t know or care about which part is ‘the surplus’. When food is consumed, it
all enters your digestive tract and it’s all mixed together. It’s not like your stomach has some
sensory valve that opens after consuming a certain amount of nutrients that corresponds with
energy maintenance and then shuttles the remainder to a separate compartment. So the idea
that “a surplus of protein isn’t converted to fat” is utter nosensense. You consume a certain
amount of calories and a certain amount of protein. This proportion of protein can impact fat
loss, as we discuss below, but primary relevant variables are protein intake and energy intake.
99
Not only is it nonsense that your body treats ‘a surplus of protein’ differently than ‘the protein
that came before the surplus’ (see how silly that is?), it is also nonsense that protein cannot be
converted to fat. As you’ll read about in detail in this week’s recommended reading, the carbon
skeleton of dietary amino acids can easily be used by the body as a source of energy.
Glucogenic amino acids, the vast majority, can be converted to glucose, which can in turn be
converted to fat. The 4 ketogenic amino acids can be converted to ketone bodies.
Some confusion occurs because it is actually true that the protein from your diet is rarely
actually converted to fat. What usually happens, is that the protein is oxidized and used as fuel.
This frees up fat, so that more of the fat from your diet is stored as fat. Regardless of if protein
itself is directly converted to fat or if it simply contributes as a fuel source, the end result is the
same: protein intake from the diet can result in fat storage. (We will go into more detail on this
in the course topic on understanding human metabolism.)
Instead of some magical property by which protein defies the laws of thermondynamics,
research has identified 3 mechanisms whereby higher protein diets can promote fat loss or
prevent fat gain.
1. Anabolism. More protein intake can result in more muscle growth. Muscle growth is a
highly energy intensive process. Greater energy expenditure can cause fat loss or
prevent fat gain.
2. The thermic effect of food (TEF). Protein has a higher thermic effect than carbohydrates
and fats, which means the body burns more calories when metabolizing protein than
when metabolizing fats or carbs. Again, greater energy expenditure can cause fat loss or
prevent fat gain.
3. Satiety. Pure protein is generally more satiating than pure carbs or fats. Greater satiety
means people will eat less if they don’t track their macros and lower their energy intake.
Let’s go through the plausibility of these explanations for Antonio’s studies one by one.
100
1. Anabolism. We can rule this out, because in no study did the higher protein group gain more
lean body mass than the lower protein group (no trend either) or had any performance
advantage.
Specifically, there was no difference in the achieved training load (sets x reps x weights) in the
first study, showing that the higher protein group did not gain more strength, unless they
somehow lost a lot of work capacity.
In the second study there was also no difference in the development of any strength or power
measure: maximal strength (1-RM squat and bench press) vertical jump and pull-ups.
In the third study there was no difference in the development of bench press 1 RM strength,
bench press strength-endurance or total training load.
2. How about TEF? This explanation is often heard: protein has a higher thermic effect than
carbs or fat. Indeed, Sutton et al. (2016) showed that high protein meals have a higher thermic
effect than low protein meals.
However, the TEF of protein is unlikely to be relevant in practice for strength trainees, because
TEF does not increase in a dose-response fashion to protein intake. In this particular study, for
example, TEF was higher with 14 g protein than with 9 or 4 grams but there was no difference
between 4 or 9 grams. These results are shown in the graph below.
101
Protein mainly has a higher TEF than carbs or fats when the macronutrients are consumed in
isolation. In the context of lean individuals consuming mixed whole food meals with sufficient
protein to induce an anabolic response (see the section on the leucine threshold), the
difference disappears and the TEF of a mixed meal is generally above 20%, which is just as large
as, if not larger than that of pure protein. (We will go into more detail on this in the course
topic on calories.)
So while it is possible that the TEF of protein played a role in these studies to ward off fat gain
due to the possible reliance on pure protein powder as meals to consume their extreme
protein intakes, but if so, there is no considerable benefit we can get from this in practice,
unless you really enjoy slamming down protein shakes in between meals.
Moreover, any potential effect of TEF is far too small to be the sole explanation of these study
findings.
102
3. That leaves satiety. The subjects in the higher protein groups must have eaten less. However,
this poses a paradox. In both Antonio’s studies, the higher protein groups consumed
significantly more calories than the moderate protein groups.
Self-report
At least, that’s what they said… In no study was there any diet control. In the first study, the
subjects were “instructed to maintain the same dietary and training habits over the course of
the study. On the other hand, the subjects in the high protein diet group were instructed to
consume 4.4 grams of protein equal to 4.4 g/kg/d.” In the second study, it was the same
instruction but now protein was ≥ 3 g protein/kg/d. In the third study, there isn’t even any
mention of instructions to the subjects. Most subjects entered their own macros into
MyFitnessPal and a few went with old school with pen and paper. MyFitnessPal has a
notoriously unreliable database, so even if they selected the foods they actually ate, this
increases the margin of error significantly. (In the course topic on setting calories, we go into a
better way to track your macros.)
Everyone intuitively knows that self-reported data on what people ate is not perfectly accurate.
However, serious strength trainees who have gotten used to tracking their macros meticulously
for years often forget just how unreliable self-reported food intake is. You have to realize that
people that track their macros meticulously are the fitness elite. When you conduct a study,
you don’t have the luxury of recruiting only these kinds of people. In fact, many researchers
have gone so far as to say that self-reported diet data has no place in scientific research:
“uncritical faith in the validity and value of memory-based dietary assessment methods (M-BMs)
has wasted substantial resources and constitutes the greatest impediment to scientific progress
in obesity and nutrition research… M-BMs are fundamentally and fatally flawed owing to well-
established scientific facts and analytic truths… Given the overwhelming evidence in support of
our position, we conclude that M-BM data cannot be used to inform national dietary guidelines
and that the continued funding of M-BMs constitutes an unscientific and major misuse of
research resources.” Ouch.
103
Now, I personally do not agree with this and I would not outright dismiss diet data when it’s
self-reported. However, these studies had a few major red flags. In the first study, the drop-out
rate was a whopping 25%: 10 out of 40 subjects did not complete the study. In the second
study, it was 34.2% with 25 out of 73 subjects not completing the study. In the third study, no
compliance data is provided.
Those are very high numbers for this type of study. For behavioral weight loss studies, Wadden
& Bell (1990) found an average drop-out rate of 13.8% based on 13 studies. Wilson (1980)
found a strikingly similar average of 13.5% based on 17 studies. And that’s generally when you
try to get fat, sedentary men and women to lose weight. These studies just tried to get strength
trained men to eat more protein for 8 weeks. (Actually, the second study doesn’t list how long
it lasted anywhere, just ‘several weeks’. Based on the training program, it appeared to be 6
weeks.) Hell, these guys volunteered for these studies.
So why did they drop out? Just try to consume 3.4 – 4.4 g/kg/d of protein yourself and you’ll
see. For a 176 lb (80 kg) man, that’s 272 – 352 grams of protein. The studies don’t list a lot of
attrition data, as it’s called: “Of the 10, three stated an inability to consume the protein needed
for the study and one subject complained of gastrointestinal distress. Six did not provide a
reason.” I talked to Jose Antonio in person and he said many subjects complained of digestive
issues and had trouble consuming this much protein.
When you volunteer for a study and you often know the people involved in the research from
your college, you don’t plan to drop out. You don’t want to let people down. What people
generally do is first try to fudge their compliance. Say you want to go to bed, but you still have
150 grams of protein left, but you’re already bloated and full. Do you forcefeed it or do you log
the 150 g of protein and just go to bed ‘just this once’? This is a well known problem called
social desirability bias. People tell you what they think you want to hear. So before someone
completely drops out of the study, you can bet they haven’t adhered to the program very well
for quite a while.
104
Jose Antonio is very open about the difficulty in getting people to do what you want in
research. In both studies, they even used unequal randomization to put more people in the high
protein groups. “The purpose of unequal randomization was to take into account the loss of
subjects from potential lack of compliance due to the high protein diet…”
So we have several studies from the same lab that were questionable in terms of peer review,
sponsorship bias and the reliability of the data, while one independent study showed that the
high protein group gained just as much fat as the low protein group. We need a reliable arbiter.
Metabolic ward data
Fortunately we have the perfect arbiter, metabolic ward research from Bray et al. [1, 2] with
gold-standard measurement techniques with groups consumign 5 vs. 15 vs. 25% of energy as
protein while massively overeating by 40%. A metabolic ward is basically a science lab where
oxygen and carbon dioxide consumption and production by your body are measured. So energy
expenditure can be accurately measured. All meals in a metabolic ward are prepared for the
subjects. So energy intake can also be tightly controlled. Basically, this research is as good as it
gets.
105
The study design of Bray et al.’s second study for those interested in the details. The first study was
similar in design.
In both of Bray’s studies, there was no difference in the amount of fat gain between the groups.
Even in a 40% energy surplus, there was no difference in fat gain between the woefully
insufficient 5% protein intake (~0.7 g/kg/d) and the massively superfluous 25% protein intake
(~3 g/kg/d). Fat gain was significantly predicted by energy intake, but there was no significant
relation with protein intake.
Moreover, in both studies the low protein group did not gain as much lean body mass as the
medium and high protein groups, but there was no difference between the medium and high
groups. The medium protein group even gained non-significantly less fat and more muscle than
the high protein group in the second study. The exact same pattern held true for energy
106
expenditure in both studies: low protein did not cut it, but there was no benefit to increasing
protein above the medium intake.
You can see the body composition changes from both studies in the following figures.
Body composition change over time in Bray et al.’s first study.
107
The results of Bray et al.’s second study.
Interestingly, as a side note, the second study also showed that both nitrogen and protein
balance overestimated muscle growth due to changes in the metabolism of residual mass, i.e.
the skin, intestines, kidneys and liver. This suggests that the acute protein balance research may
overestimate protein requirements.
In support of the findings of Bray et al’s careful metabolic ward data, a meta-analysis and
systematic review shows that high protein diets do not result in more fat loss than low protein
diets when both diets have the same amount of calories.
Conclusion
Gold-standard research shows that when you are already consuming enough protein in each
meal and across the day (~1.8 g/kg/d), there is no metablic, thermic or body composition
benefit to increasing your protein intake further. How much fat you gain or lose is primarily
dictated by energy balance, not by protein intake, since protein can be easily used as fuel by the
108
body or even stored as fat. The explanation for the questionable Antonio studies is most likely
simply that the subjects overreported their energy and protein intakes because they literally
couldn’t stomach the extreme protein intakes they were required to consume.
109
Practical appliations for protein intake

 Consume a minimum of ~1.8 g/kg of protein every day, possibly more for hard training
novice men, mixed athletes, drug users and vegeterians; possibly less for women,
advanced trainees, people that don’t follow an optimized program design and obese
individuals. It doesn’t matter whether you’re in an energy surplus or deficit.
 Protein supplementation is generally not advisable in comparison to whole foods when
it comes to stimulating muscle protein balance. Animal protein sources are preferable to
plant protein sources.
 Make sure each meal contains a complete amino acid profile and hits the leucine
threshold.
 Protein needs to be factored into your total daily energy intake just like carbohydrates
and fat.
 Meal frequency per se does not generally have any considerable effect on total protein
balance or energy expenditure, but the protein distribution across the day does.
110

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Bayesianbodybuilding.

Amino acid biochemistry 101

Essential and non-essential amino acids

In contrast to chemists, nutritionists generally categorize the amino acids in terms of

The 9 essential amino acids are:

Identifying amino acids strictly as nonessential/dispensable or essential/indispensable is an

Protein biochemistry 101

 Peptide: a short amino acid chain.

o Tripeptide: a peptide consisting of 3 amino acids.

3. The tertiary structure of protein is the overall or total three-dimensional configuration

4. Some proteins also have a quatarnary structure as a result of interacting polypeptide

Amino acid sources

Dietary (exogenous) sources of protein include:

Moreover, nitrogen assimilation following ingestion of protein-containing foods is superior to

Amino acid metabolism

Amino acid catabolism

The urea cycle.

Gluconeogenesis and ketone body production

The ketogenic amino acids are:

Fatty acid production

Other uses of amino acids

(HMB), for instance, promote protein synthesis through increased phosphorylation of

Diagram detailing the role of amino acid availability in regulating muscle

The function of proteins and nitrogen-containing compounds

adequate concentrations of protein in the blood (e.g., by infusing albumin intravenously)

Immuno-protectors and acute phase responders

Immuno-protection is provided to the body in part by a group of proteins called immuno-

Nitrogen-containing non-protein compounds

Purine and pyrimidine bases

Inter-organ ‘flow’ of amino acids and organ-specific

Amino acids in the blood

Intestinal cell amino acid metabolism

Organ-specific glutamine metabolism

The alanine-glucose cycle

Indicators of muscle mass and muscle/protein catabolism

The urinary excretion of 3-methylhistidine is used an indicator of muscle catabolism

How much protein do you need?

Studies on the optimal protein intake

• Tarnopolsky et al. (1992) observed no differences in whole body protein synthesis or

Study Subjects (n) Strength Hypertrophy Maximally

Iglay et al. Elderly Composite 1 Body composition (incl. <= 0.9

Tarnopolsky Bodybuilders/ N/M Lean body mass and <= 1.05

Tarnopolsky Bodybuilders Bench Press & Body Composition <= 1.35

Hambre et Young males Quadricep Body composition (incl. => 1.4

Antonio et Strength- N/M Body composition (incl. <= 2

Walberg et Male N/M Body composition (incl. <= 0.8 (MPS)

Antonio et Strength- Bench press 1 Body composition (incl. <= 2.6

Consolazio Young healthy N/M Body composition (incl. => 1.4

“But I train harder!”

“But I’m more advanced than these geeks!”

What about when cutting? Or you’re really lean?

more than 1.2 g/kg protein.

A systematic review of dietary protein during caloric restriction in resistance

The theory that you need more protein in a deficit

Of course, Eric et al. do offer an explanation:

The evidence that you need more protein in a deficit

Here’s the abstract.

Should you set protein relative to bodyweight or lean body mass?

Performance enhancing drugs (PED, particularly androgenic-anabolic steroids (AAS), may