Activity Proper

1. Why is the brain the most important consumer of glucose?

- Because our brain is considered as energy-hungry organ which demands high
amounts of energy, the foods we eat greatly affect how will our brain function, with
everything from learning and memory to emotions and the glucose is main source of
energy for humans. The central nervous system depends greatly on glucose from
surrounding extracellular fluid because the nerve cells cannot concentrate or store
carbohydrates it is dangerous to maintain a steady supply.
2. Explain the process of carbohydrate digestion.
- It begins when you digest carbohydrates the minute the food hits your mouth. The
saliva secreted from your salivary glands moistens food as it’s chewed. This saliva
releases an enzyme called amylase, which begins the breakdown process of the
sugars in the carbohydrates you’re eating.
- From there, you swallow the food now that it’s chewed into smaller pieces. The
carbohydrates travel through your esophagus to your stomach. At this stage, the
food is referred to as chyme. Your stomach makes acid to kill bacteria in the chyme
before it makes its next step in the digestion journey.
- The chyme then goes from the stomach into the first part of the small intestine, called
the duodenum. This causes the pancreas to release pancreatic amylase. This
enzyme breaks down the chyme into dextrin and maltose.
- From there, the wall of the small intestine begins to make lactase, sucrase, and
maltase. These enzymes break down the sugars even further into monosaccharide
or single sugars. These sugars are the ones that are finally absorbed into the small
intestine. Once they’re absorbed, they’re processed even more by the liver and
stored as glycogen. Other glucose is moved through the body by the bloodstream.
The hormone insulin is released from the pancreas and allows the glucose to be
used as energy.
- Anything that’s left over after these digestive processes goes to the colon. It’s then
broken down by intestinal bacteria. Fiber is contained in many carbohydrates and
cannot be digested by the body. It reaches the colon and is then eliminated with your
stools.
3. Monosaccharide other than glucose cannot be directly used by cells. What
process do these monosaccharides undergo in the body? Explain.
- Because glucose is the most important type of monosaccharide, and the only one
who can be involved in cellular respiration to produce ATP and NADH. Meanwhile,
galactose and fructose must be converted to glucose before used. It is the goal of
the cell to convert glucose to carbon dioxide and alter during the process to obtain
high energy molecule ATP from inorganic phosphate ad ADP.
4. Explain comprehensively the following processes
a. Glycolysis
- is the metabolic pathway that converts the main source of energy, the glucose into
pyruvate or lactate for energy production. The free energy released in this pcorss is
used to form the high energy compounds, ATP or adenosine triphosphate. And
NADH which is a reduced nicotinamide adenine dinucleotide. This process is a
sequence of 1ten reactions involving ten intermediate compounds.
b. Gluconeogenesis
- is the process that uses other non-carbohydrates rather than glucose like amino
acids to form the glucose-6-phosphate. Aspartic acid is also a metabolite in the urea
cycle and participates in gluconeogenesis, the generation of glucose from non-sugar
carbon substrates. The liver is the major organ for removing lactate by converting
lactate back to glucose by a process called gluconeogenesis.
c. Glycogenolysis
- is the process in which the glycogen is break down to be a glucose that utilized as
energy or be converted to glucose-6-phosphate for entry to glycolytic pathway.
During a brief fast, glucose is supplied to the ECF from the liver through
glycogenolysis. Galactosemia occurs because of the inhibition of glycogenolysis and
is accompanied by diarrhea and vomiting.
d. Glycogenesis
- is the term given for the converting glucose to glycogen to be the most important
storage polysaccharide in liver and muscle.
e. Lipogenesis
- Is the synthesis of fatty acids from non-lipid precursors like glucose and other
substrates. HNF1A mutation increases lipogenesis by promotion of fatty acid
synthesis and downregulation of liver-type fatty acid–binding protein 1 (L-FABP),
leading to diffuse intralesional Steatosis. Glucocorticoids also stimulate adipocyte
 differentiation and promote lipogenesis through the activation of enzymes such as
lipoprotein lipase and increased messenger ribonucleic acid (mRNA) expression for
leptin.
f. Lipolysis
- Is the process occurs in adipose tissue by which fat is broken down or being
decomposed through enzyme of water or through hydrolysis. If lipolysis of
triglycerides is unregulated, it will form ketone bodies then the body will use it as the
energy source through the Tricarboxylic Acid Cycle.
5. What are the hormones that regulate carbohydrate metabolism? Explain their
respective functions.
- The two major hormones that regulate the CHO metabolism are the insulin and
glucagon, in which insulin let the entry of glucose in the cell. It is synthesized by B
cells of islet. Insulin is known as the hypoglycemic hormone because it is the only
one that can decrease plasma glucose whenever glucose is in peak, it will increase
the movement of glucose inside the cell. While glucagon is release whenever
glucose is low and it is known as the hyperglycemic hormone. It will be produce
during stress and fasting. It also promotes glycogenolysis and gluconeogenesis.
- Another two hormones produced by adrenal gland are the epinephrine and
glucocorticoids. Epinephrine increase plasma glucose by inhibiting insulin secretion
and release in times of stress while glucocorticoids increase glucose by lowering
intestinal entry into the cell.
- Meanwhile, growth hormone and ACTH are produced by the anterior pituitary gland.
Growth hormone increase plasma glucose by decreasing entry of glucose in cells
and increasing glycolysis and decrease levels of cortisol stimulates anterior pituitary
to release ATCH that will increase plasma glucose by converting liver glycogen to
glucose and promote gluconeogenesis.
- Other hormones like thyroxine which is stimulated by the TSH that increase plasma
glucose by increasing glycogenolysis, gluconeogenesis, and intestinal absorption of
glucose.
6. What is the standard specimen used in the clinical laboratory determination of
glucose?
- In choosing the standard specimen for glucose determination, the phlebotomist
should use venous plasma glucose.
7. For fasting blood sugar (FBS) testing, one needs to fast (of course). Why is so?
And how do you instruct the patient to fast?
- Because of the reason that fasting helps ensure the blood test records an accurate
measure of fasting blood sugar levels.
- The results help a doctor to diagnose or rule out diabetes.
- The health professional must be able to talk to the patient in verbal form or use
layman’s term in giving instructions to the patient. The patient must be told to not to
eat or drink anything other than water for 8 to 10 hours before a fasting
blood glucose test.
8. Why do we need to separate cells from serum or plasma within 30 to 60 minutes?
- We need to separate cells from serum or plasma within 30-60 minutes to ensure and
prevent the lysis of cells that may lead to releasing cellular components not usually
found in serum samples. While serum or plasma samples that are allowed to sit less
than 30 min are likely to retain cellular elements and other contaminants impacting
future analysis. In short, this will prevent substantial loss of glucose by cellular
fraction particularly, if white blood cell count is high.
9. What is the ideal tube for glucose testing? Explain the function of its additives.
- The most ideal tube for glucose testing is the grey-top tube which has an additive of
2mg potassium oxalate or sodium fluoride used to inhibit glycolytic enzyme enolase
by binding into magnesium and acts as an anticoagulant resulting in whole blood or
plasma. It interferes with Na, K, and most BUN determinations
10. How does CSF glucose compare with that of plasma?
- The glucose level in CSF should be proportional to the blood glucose level and
corresponds to 60-70% of the concentration in blood. Therefore, normal CSF
glucose levels lie between 2.5 and 4.4 mmol/L (45–80 mg/dL).
11. If one needs to measure CSF glucose, what are the pre-analytical considerations?
- The health professional should remember that blood glucose must be collected 1-2
hours before spinal tap and if delayed, but should deliver the samples immediately, it
must be freeze at -20 degrees Celsius or ref at -4 degrees Celsius.
12. What are the chemical methods of glucose determination? Explain each (include
the reagent and positive result).
a. Folin Wu
- When glucose or other reducing agents are treated with an alkaline copper solution
they reduce the copper with the result insoluble cuprous oxide is formed. The
 cuprous oxide form is allowed to react with phosphomolybdate to form positive result
Phosmolybdenum blue measured at 520 nm.
b. Nelson Somogyi
- The reducing sugars when heated with alkaline copper tartrate reduce the copper
from the cupric to cuprous state and thus cuprous oxide is formed. It is commonly
used to measure the ‘true’ glucose. When cuprous oxide is treated with
arsenomolybdic acid or arsenomolybdate and the positive result will be
arsenomolybdenum blue.
c. Neocuproine
- One method for determining copper is the “Neocuproine Method”. In this method,
copper in a +1 oxidation state reacts with the reagent neocuproine to form an
organge-red or yellow to orange result. The complex is extracted into a chloroform-
methanol mixture, giving a yellow solution with a molar absorptivity of 8000 M -1 cm-1
at 457 nm. Beer’s law is obeyed up to a concentration of 0.2 mg Cu/25 mL of
extraction solvent. Full color development occurs when the sample’s pH is between 3
and 9. 
13. If the result of oxidation-reduction methods is positive or high, does it mean that
there is a presence or there is high level of glucose? Explain.
- Yes, because when oxidation-reduction methods are high, the glucose tends to
increase too because of their linear relationship and because glucose is can be
oxidized and thus considered to be reducing sugar, it has free anomeric carbon at
carbon 1 position.
- Analytic methods that are based on oxidation–reduction reactions may be influenced
positively or negatively by ingested substances such as ascorbic acid (vitamin C).
This interference is observed in chemical testing of serum on automated analyzers
and it can also occur in urine testing for glucose which has positive interference for
reducing substance method.
14. Why is Hagedorn Jensen Method also known as inverse colorimetry?
- Because it is the only oxidation reduction method that uses alkaline ferric reduction
method or ferricyanide method unlike other test that uses alkaline copper reduction
methods.
15. What are the enzymatic methods of glucose determination? Explain each (include
the reagent and positive result)
a. Glucose Oxidase
 - Is a kind of flavoenzyme and an enzymatic method that catalyzes the oxidation of
glucose to gluconic acid and hydrogen peroxide. It is highly specific to measure B-D-
glucose by using mutarotase which converts α-D-glucose to β-D-glucose. The
reagents used are glucose oxidase, peroxidase and chromogen. This chromogens
are positive with blue color in O-dianisidine and in aminoantipyrine with the positive
color of rose-pink.
b. Hexokinase Method
- is the most specific glucose method because it is commonly used as reference
method. It is a two-step reaction of glucose concentration that is proportional to the
rate of production of reduced NADPH which is followed spectrophotometrically. The
absorbance of this method is measured at 340 nm. One thing to remember about
hexokinase method is that NAD is required if G6PD is obtained from Leuconostoc
mesenteroides and NADP is required if G6PD is obtained from yeast. The product
for this reaction is 6-phosphogluconate or 6-phosphogluconolactone.
c. Glucose Dehydrogenase Method
- An enzymatic method that catalyze oxidation of glucose to gluconolactone with
concomitant reduction of NAD+ to NADH. In addition, the variations of
spectrophotometric use the reagents mutarotase, GD, diaphorase and end with the
product MTTH (Blue) + NAD. While through electric current uses the reagents
pyrroloquinoline quinone, GD, Ferricyanide and will result to Ferricyanide + 2e.
- This method reduced the glucose to produce chromophore that is measured
spectrophotometrically or an electric current.

16. In glucose oxidase method, why is there a need for a pretreatment of serum with
mutarotase?
- Glucose oxidase converts B-D-glucose to gluconic acid. Mutarotase may be added
to the reaction to facilitate the conversion of a-D-glucose to b-D-glucose. Oxygen is
consumed and hydrogen peroxide (H2O2) is produced. The reaction can be
monitored polarographically either by measuring the rate of disappearance of oxygen
using an oxygen electrode or by consuming H2O2 in a side reaction.
17. What are the sources of error in glucose oxidase methods and why.
- The sources of error in glucose oxidase methods can be strong oxidizing agents like
bleach that will lead to falsely increase values. By using oxygen consumption
electrode which directly measure oxygen by polarographic technique, it will prevents
 interference. Another error is the consumption of reducing agents like ascorbic acid,
uric acid, bilirubin, hemoglobin that will lead to false negative and the addition of K
ferrocyanide that will decrease interference with Bilirubin.
18. What are the remedies if the sample to be tested for glucose using glucose
oxidase method (polarographic or colorimetric) is icteric?
-
19. What is the reference method for glucose determination? Why is it considered as
the reference method?
- Hexokinase method because the results for the glucose samples taken were mostly
the same to theoretical values and does not suffer from any interference, therefore
the appearance of ascorbic or uric acid in the samples did not affect the results
unlike the glucose oxidase in which uric or ascorbic acid inhibits the enzyme that
may cause low catalytic activity, but in hexokinase method, they do not disturb and
inhibit their catalytic activity.
20. Is the reference method routinely performed in the clinical lab? Why or why not?
- The answer is no. It is because even though it is highly accurate and precise, the
reference method is too demanding and time-consuming for routine use in a clinical
laboratory.
21. What are the remedies if the sample to be tested for glucose using hexokinase
method is
a. Hemolyzed
- Hemolyzed specimens can be problematic in that contents released from the RBC
may interfere with the stoichiometric relationship between glucose andNADPH
accumulation. So with the remedy, the specimen containing more than 0.5g
haemoglobin/dl are unsatisfactory because phosphate esters and enzymes released
from red blood cells interfere with the assay. Also the red blood cell count and
haematocrit may be decreased so might as well repeat the process.
b. Icteric
- Hexokinase method for icteric specimens can be sort out by using a sample blank.
This blank is prepared by adding 10ul of sample to isotonic saline or buffer instead of
reagent. The absorbance of this mixture, read against water at 340 nm, is
subtracted.
22. What is the most accurate method of glucose measurement?
 - Isotope Dilution Mass Spectrometry or Gas Chromatography is the most accurate and
reliable method. Since it is an analytical technique, the results tend to fulfils all
requirements to be used in increasing order reference measurement procedure.
23. Enzymatic methods are most employed in the clinical labs and POCT instruments
in measuring glucose. What is the rationale behind this?
- The rationale behind this is the fact that enzymatic methods provide specificity an
can be package to furnish point of care determinations. The different types of test
under enzymatic methods produce an electric current that is proportional to the initial
glucose concentration, or a product that measures spectrophotometrically is
proportional to the initial glucose concentration. The assays can be initial rate-of-
change assays, where the velocity of the reaction is dependent on the initial glucose
or end-point assays.
24. What are the types of glucose testing? Differentiate them in terms of process,
uses, and reference interval.
a. Home Glucose Monitoring
- Glucose oxidase and reagents to measure the generation of hydrogen peroxide can
be bonded to filter paper and the system used to measure glucose concentrations in
a drop of capillary blood. This has resulted in the most important change in diabetes
management since the introduction of insulin
b. Random Blood Glucose (RBS)
- Taken anytime of the day without fasting and used for emergency cases examples
when the patient is having hyperglycemic ketonic coma, insulin shock.
c. Fasting Blood Sugar (FBS)
- Taken after at least 8 to 10 hours of overnight fasting and usually done during the
morning to prevent the effects of diurnal variation. With the categories of normal
FBS: <100 mg/dL; Impaired FBS/Prediabetes: 100 to 125 mg/dL; Provisional DM
diagnosis: ≥126 mg/dL
d. 2-hours Post-Prandial Glucose
- FBS initially, the patient is given CHO load (usually 75g) and plasma glucose is
measured after 2 hours. Categories: Normal: <140 mg/dL; Impaired glucose
tolerance/Prediabetes: 140-199 mg/dL; Provisional DM diagnosis: ≥200 mg/dL
e. Oral Glucose Tolerance Test (OGTT)
- Series of glucose testing after 8 to 10 hours of testing.
f. Glycosylated Hemoglobin
 - A test that reflects long-term blood glucose control in diabetics is the concentration of
hemoglobin A1c. When hemolysates of red cells are chromatographed, three or
more small peaks named hemoglobin A1a, A1b, and A1c are eluted before the main
hemoglobin A peak.
25. In OGTT(Single dose)
25.1 Why is there a need for patients to go with normal carbohydrate diet for 3
days prior the test?
- The patients that will undergo test for OGTT must have a 3-day diet with at least
150g carbohydrate per day to reduce the false-positive diagnoses of gestational
diabetes.
25.2 Why should the OGTT be discontinued if the FBS is >140 mg/dL?
- If the blood glucose in OGTT extends in more than 140mg/dL after drinking the
glucose mixture, a person is said to have a pre-diabetes or the physician would say
that the patient has an impaired glucose tolerance, indicating the relatively high risk
for the development of diabetes in these patients.
25.3 Why should the test be discontinued is the patient vomits?
- Perhaps the patient has a needle phobia or has been fasting too long. It can also be
because of the patient’s condition. It happens sometimes in Glucose Tolerance
Testing. The phlebotomists must have a waste container available. Offer patient
tissues or washcloth for possible clean up and may need to discontinue test and
reschedule for another day. Regardless of an uneventful outcome, it demands your
immediate termination of the procedure before things get worse. Be thankful you
have time to remove the needle before he passes out or vomits on your shoes.
26. Why is HBA1c considered as the test for long term glucose control?
- Because hemoglobin are proteins that glucose sticks over time, and this proteins are
inside the red blood cells which has a normal life span of 120 days, glucose
molecules often attached to hemoglobin, the more sugar in your blood, the more it
forming glycated hemoglobin. In individuals with poorly controlled diabetes,
increases on the quantities of these glycated hemoglobins are noted, therefore,
HBA1C does not measures the amount of glucose in a drop of blood, instead it
measures the percentage of glucose stuck to the hemoglobin in the blood or the
percentage of glycation of the hemoglobin.
27. Why is EDTA anti-coagulated WB the preferred specimen in HBA1c testing?
 - Because EDTA is testing for hemoglobin and that means it should be prevented from
clotting. We cannot use serum or red top because this will lead into coagulated
blood, even though HBA1C is part of the clinical chemistry section, it is the only test
that can have a lavender top and whole blood specimen.
28. What is the most commonly employed HBA1c testing method in the clinical labs?
In POCT instruments?
- The affinity chromatography method, which is preferred by most technicians in the
laboratory, uses a boronate resin group to attach the glycosylated hemoglobin, which
is then selectively eluted from the resin bed using a buffer. This method is not
temperature dependent and not affected by fetal hemoglobin, hemoglobin with
sickle-cell trait, or hemoglobin C.
- The HbA1C point-of-care assay is based on a latex immunoagglutination inhibition
method. In this method, the concentration of HbA1C and the concentration of total
hemoglobin are measured; the ratio is reported as percentage of HbA1C. Because
glycated hemoglobin F is not measured using this method, very high levels of
hemoglobin F of more than 10%, it will cause the HbA1C level to be lower than
expected because a greater proportion of the glycated hemoglobin will exist in the
form of glycated hemoglobin F. Also, HPLC and electrophoresis methods can be
used to separate the various forms of hemoglobin into A1A,A1B and A1C.
29. What is the reference interval for HBA1c
- Reference intervals for HbA1c is expressed as median and 2.5th to 97.5th percentile
for each trimester were: T1: 5.1 (4.5–5.6%), T2: 5.0 (4.4–5.5%), and T3: 5.1 (4.5–
5.6%).
30. Differentiate Type 1 and Type 2 diabetes in terms of pathogenesis
a. Type 1 diabetes
- is the sudden onset of disease that affects any age but mostly young people with the
body habitat of being thin or normal. It is commonly known as Juvenile-onset or
Insulin-Dependent diabetes. Ketoacidosis is common and usually, autoantibodies are
present with beta cells being destroy due to autoimmunity and results in lost insulin
production, while endogenous insulin are low or absent with less prevalent. It can
have a gene-environment interaction and a chromosome 6-human leukocyte antigen.
b. Type 2 diabetes
- is the gradual onset of disease that mostly affects adults and people with this
disease are often obese. This is non-insulin dependent diabetes and is correlates on
 insufficiency of insulin with poorly utilized by tissues. Ketoacidosis is not commonly
detected and autoantibodies are absent with endogenous insulin being normal or
decreased/increased. It is more prevalent than Type 1 but considered to be
preventable.
31. Why is c-peptide levels detectable in type 2 DM but not in type 1?
- C-peptide is used to measure insulin secretion in both types of diabetes. When there
is an absence or low c-peptide levels, this suggests that there is a phenotype of DM1
while type 2 DM will have a normal or high level of C-peptide, so it is therefore
detectable in type 2 DM than type 1 DM. This test may be ordered to monitor the
status of beta cell function and insulin production over time and to determine when
insulin injection may be required.
32. How do we confirm the diagnosis of DM using lab methods?
- The preferred test in diagnosing diabetes mellitus is the measurement of the fasting
plasma glucose level.
33. Why is kidney disease one of the complications of DM?
- High blood glucose, also called blood sugar, can damage the blood vessels in your
kidneys. When the blood vessels are damaged, they don’t work as well. Many people
with diabetes also develop high blood pressure, which can also damage your
kidneys.