Process biochemistry

Wine production is a very old finding and it is defined as an alcoholic beverage obtained from grape juice using yeast as a fermenting organism [1]. With the progress of biotechnology many changes were brought in traditional wine making procedures [2]. In the mid of 19th century application of immobilized enzymes was developed and this technique was introduced and applied in the field of analysis and industrial production of biomolecules [3]. In recent years, cell immobilization techniques have become increasingly important and are being successfully applied in industrial processes such as the production of alcohols (ethanol, butanol and isopropanol), organic acids (including malic, citric, lactic and gluconic acids), enzymes (cellulase, amylase, lipase and others) and biotransformation of steroids for hormone production, wastewater treatment, and food applications (beer and wine) [4]. Cell immobilization in alcoholic fermentation is a rapidly expanding research area because of its beneficial technical and economic advantages compared to the conventional free cell system [3,5]. The host for yeast immobilization for wine making is always biologically inert [6]. In alcoholic fermentations, many investigators proposed wide variety of supports for immobilization of yeast cells like alginate and other inorganic materials [7-9]. Various supports have been proposed for potential use in main or secondary fermentations of wine, including inorganic supports such as, mineral kissiris [10] and Ca-alumina [11] since they are abundant and are of low cost there is a possibility of unlimited reuse. But none of them fulfilled the food-grade properties. Foodgrade natural supports such as, delignified cellulose materials [12], gluten pellets [13] and also fruit pieces like apple and quince [14,15] were successfully used as immobilization supports for wine production. However, for industrial wine production, it is important to identify a suitable support for cell immobilization that is of food grade, readily available, cost-effective and which may contribute to an overall improvement in the sensory characteristics of the final wine product.

The effects of Ni, Cu and Hg on thiol and exopolysaccharide synthesis in Nostoc spongiaeforme has been investigated. LD 50 s for Ni, Cu and Hg based on the specific growth rate, were 8, 4 and 0.2 mM, respectively. The presence of Ni was most favourable to thiol production (6d) at 1, 2 and 5 mM as the average value reached 43.37 mmol/g dry wt over the control cells (22.5 mmol/g dry wt). Cu in the same concentration range, favoured the process (average 33.64 mmol/g dry wt). The thiol level in cells dosed with Hg (0.2 mM, LD 50) was similar to that in the control sets. Nevertheless, lower concentrations of Hg (0.15 mM) produced a rise in thiol to 31.12 mmol on 6d. A time course study on thiol biosynthesis at the respective LD 50 concentrations of Ni, Cu or Hg indicated that with the less toxic cations more time was required to induce thiol biosynthesis with ultimate hyperproduction while the more toxic cations reduced the lag as well as the final thiol level. The effectiveness of cations, therefore, followed the order Ni \Cu \Hg while that of toxicity was Hg \ Cu\ Ni. Exopolymer systhesis was also metal-specific as the maxima corresponded to 80, 26 and 48 mg glucose equivalent/mg dry wt for Cu (2 mM), Ni (5 mM) and Hg (0.05 mM), respectively relative to the control (25 mg glucose equivalent). The sequence Cu \Hg \Ni, therefore, presents a contrast with those for thiol production. These data form the first report on the metal-specific nature of thiol and exopolymer production in a cyanobacterium.

The activated sludge process is the most widely used biological wastewater treatment method. The measurement of some physicochemical parameters in aeration tanks does not allow clarification operation failures to be avoided. This study focuses on the determination of a rheological parameter of filamentous activated sludge at standard concentrations (2-7 g L À1) in order to assess sludge settleability. On filamentous sludges, rheological measurements in shear flow display a hysteresis area (or reduced hysteresis area (rHa) when the volume of sludge considered is 1 mL), which corresponds to the presence of filamentous bacteria in the sludges. In numerous cases of filamentous activated sludges, significant relationships with the zone settlement velocity (ZSV) parameters of settleability point out that hysteresis area could be used to predict the ability of filamentous activated sludges to settle. With the protocol used, a filamentous activated sludge with a rHa >20 Pa s À1 mL À1 , always showed a very low or null ability to settle whatever the value of activated sludge total suspended solid (TSS) content over a standard range (2-7 g L À1)

Fluorene degradation using immobilized Pleurotus ostreatus on various supports including polystyrene foam, Amberlite XAD-7, Amberlite XAD-2000, pumice particles and sand has been reported. These supports have yielded positive results with regard to performing metabolic functions, but the drawback of synthetic resins is that they are not eco-friendly. If natural supports could be used for cell immobilization for the degradation of fluorene, they would be both eco-friendly and perform metabolic functions. Therefore, This study attempts to design an immobilized P. ostreatus using pumice as a support specifically for the application of fluorene biodegradation in batch shaking systems. Laccase and MnP enzyme activities were monitored in these systems. The maximum fluorene degradation level was 89.16%. Batch degradation of fluorene in a packed bed reactor was also studied. The maximum fluorene degradation level was found to be 82.46%. Additionally, two different, synthetically concocted wastewater samples containing fluorene were studied in a recycled packed bed reactor in repeated mode. At the end of biodegradation process, the metabolites of the fluorene biodegradation produced by immobilized P. ostreatus were analyzed via GC-MS and FT-IR. It was concluded that the biotransformation of fluorene by immobilized P. ostreatus was achieved.

Dysregulation of human epidermal growth factor receptor pathways by over-expression or constitutive activation can promote lung tumor processing including angiogenesis and metastasis and is associated with poor prognosis in non-small cell lung cancers. Ganoderma also known as Lingzhi has been one of the most popular chemoprevention mushrooms in East Asia for centuries. Among many bioactive components identified from Ganoderma, an immunomodulatory protein is the major ingredient for the treatment of lung cancer. Recombinant fungal immunomodulatory protein, GMI, was cloned from Ganoderma microsporum and purified. However, knowledge on the pharmacological and molecular mechanisms in suppressing EGF-mediated tumor invasion and metastasis is poorly understood. The goal of study is investigate in suppressing tumor invasion and metastasis activity of GMI. GMI exhibited an inhibitory effect on EGF-induced migration and invasion. GMI treatment with EGF presented the most potent anti-migration and anti-invasion properties on Boyden chamber assay. GMI inhibited EGF-induced phosphorylation and activation of EGFR and AKT pathway kinases in a dose-dependent manner. Additionally, the EGF-induced activation of Cdc42 GTPase was inhibited by GMI, while GMI had little effect on the EGF-induced activation of Rac1 GTPase. GMI also inhibited the EGF-induced microfilament depolymerization. These findings are the first to reveal the novel functions of GMI in tumor anti-metastasis and the molecular basis for its anticancer action.

The enzymatic acylation of isoquercitrin with fatty acid esters of various carbon chain lengths was carried out in 2-methyl-2-butanol using Novozym 435 1 . The conversion yield and the initial rate decreased from 66% to 38% and from 17.7 to 10.1 mmol/h respectively, as the carbon chain of the acyl donor increased from C4 to C18. Isoquercitrin acylated derivatives showed higher xanthine oxidase inhibition activities than isoquercitrin. This property increased with the lipophilicity of the derivative esters. The scavenging activity of isoquercitrin esters against ABTS and DPPH radicals decreased with the acyl chain length. Conversely, for esters from C6 to C18, a linear growing relationship can be established between the chain length and the superoxide radical scavenging activity. Furthermore, an improved antiproliferative effect on Caco2 cancer cells was induced by addition of isoquercitrin esters comparing with isoquercitrin. ß

... The use of starter cultures in the fermentation of boza, a traditional Turkish beverage. Murat Zorba a , Omre Hancioglu a , Mahmut Genc a , Mehmet Karapinar Corresponding Author Contact Information , a , E-mail The Corresponding Author and Gulden Ova a. ...

The scale-up of a 10 L air pulsed bioreactor for the continuous treatment of textile wastewater by pellets of the white rot fungus Trametes versicolor has been carried out, based on the geometric similitude with lab-scale bioreactors (0.5 and 1.5 L). Decolourisation experiments of 150 mg L À1 Grey Lanaset G dye solution carried out in the pilot-scale bioreactor showed that in both discontinuous and continuous treatment with an HRT of 48 h, the decolourisation levels were higher than 90%. Some operational changes were carried out in the continuous decolourisation treatment of the dye solution in order to adapt the process to industrial conditions such as, non-sterilization of the dye solution, use of tap water instead of distilled water plus macronutrients and micronutrients and the use of industrial quality co-substrate instead of reagent grade. The pilot system was working continuously during 3 months and over 70 days without sterilization of the dye feeding solution, achieving good decolourisation levels (78% average during the treatment). Continuous treatment of real industrial textile wastewater under non-sterile conditions was carried out during 15 days in the pilot-scale bioreactor, with colour reduction levels between 40 and 60%. These dye concentrations are regarded as environmentally acceptable to be discharged into a municipal wastewater treatment plant if necessary according to the local regulation.

The genes, cDNA alpES1 and alpES1, encoding Aspergillus clavatus ES1 alkaline protease were amplified from complementary DNA (cDNA) and genomic DNA, respectively, cloned in pCR ® II-TOPO plasmid and then sequenced. Sequence analysis of the cDNA alpES1 gene revealed an open reading frame (ORF) of 1212 bp encoding a pre-pro-protein of 403 amino acid residues consisting of a 21-aa signal peptide, a 100-aa pro-peptide and a 282-aa mature protein with a calculated molecular weight of 28.5 kDa. Compared to the cDNA alpES1 gene, the alpES1 gene contained three introns, which had 53, 57 and 54 bp, respectively. The cDNA alpES1 gene was then sub-cloned in pET-30b(+) and expressed in Escherichia coli BL21 (DE3). The purified recombinant protease had a molecular weight of about 32 kDa estimated by SDS-PAGE. Kinetic parameters, K m and k cat values of the recombinant AlpES1 for casein, were 0.23 mM and 12.38 min −1 , respectively. The catalytic efficiency (k cat /K m ) was 53.82 min −1 mM −1 .

A method to obtain high-expression levels of recombinant horseradish peroxidase isozyme C (HRP C) in Spodoptera frugiperda Sf-9 cell culture and a strategy for its purification are described. HRP C was secreted into the culture medium where it accumulated to 25.6 mg/l. Addition of hemin to the insect cell culture increased the level of active enzyme expression up to 41.3 mg/l. A selective staining procedure using 3,3 -diaminobenzidine allowed visualisation of HRP C in the infected insect cells and provided an alternative staining strategy for titration of recombinant baculovirus carrying the HRP gene. Immobilised metal ion affinity chromatography using a Ni-NTA matrix with elution in the gradient-step mode yielded a 68% HRP C recovery with a RZ of 2.8. When the displacement elution mode was utilised, the yield was essentially the same and the product was electrophoretically pure, having a RZ of 3.2.

Various agro-industrial residues in combination with peptone, NH 4 Cl and/or soy bran were screened as substrates for extracellular bglucosidase (BGL) production by Monascus purpureus NRRL1992 on submerged fermentations (SmF). Higher BGL production was achieved when the agro-industrial residues were combined with peptone, and the utilization of NH 4 Cl (inorganic nitrogen source) had not supported high enzyme production. The combination between grape waste and peptone was the best for enzyme production, and was selected as the growth substrate for further investigations. The evaluation of the effects of the medium components on enzyme production showed that the influence of peptone was more important than grape waste. The production of extracellular BGL by M. purpureus was inducible and controlled by carbon (glucose) catabolite repression. #

The combined effects of macronutrients of media on a-amylase production by Bacillus sp. were studied using response surface methodology. A 2 2 full-factorial central composite design was chosen to explain four medium constituents, viz. starch, glycerol, yeast extract (YE) and peptone, and to analysis of the results. This procedure limited the number of actual experiments performed while allowing for possible interactions between four components. The P-value of the coefficient for quadratic effect of glycerin concentration was <0.0001, suggesting that this was the principal experimental variable having the greatest effect on the production of a-amylase. It was found that YE had no effect on a-amylase production. The optimal combinations of media constituents for maximum a-amylase production were determined as 17.58 g/l starch, 12.37% (v/v) glycerin, 8.77 g/l peptone and 0.00 g/l YE. #

The potential of noninvasive determination of glucose, lactic acid, and nisin in Lactococcus lactis subsp. lactis biofilm fermentation was investigated through fourier transform mid-infrared (FTIR) spectroscopy. Samples obtained from a biofilm bioreactor were analyzed with traditional methods and FTIR spectroscopy. The FTIR spectra were interpreted by using suitable spectra wavenumber regions through multivariate statistical techniques such as partial least square (PLS) and principal component regression (PCR). The standard error of calibration for the PLS-1 st derivative calibration models for glucose, lactic acid, and nisin were 3.87 g/l, 2.62 g/l, and 189.6 IU/ml, respectively. Prediction errors were low for glucose and lactic acid, whereas nisin could be reliably quantified when its concentration is higher than 800 IU/ml. Results indicated that FTIR spectroscopy could be used for rapid detection of glucose and lactic acid concentrations, and nisin activity in nisin fermentation.

The production of laccase by Trametes versicolor in airlift bioreactors was studied. In order to enhance laccase production several inducers were tested, among which 2,5-xylidine led, by far, to the highest activities (around 1500 U/l). The bioreactor operated in two successive batches for 40 days with no operational problems and pellets of regular size were maintained throughout the fermentation. The bioreactor configuration employed is appropriate for laccase production by free cells of T. versicolor. The extracellular liquid obtained was studied in terms of pH and temperature activity and stability. The ligninolytic fluid produced was fully active at pH 3 and 60 °C. Moreover, it was stable at pH 5 and retained about 70% activity at 30 °C for 12 days. In addition, it was able to decolourise the sulphonephthalein dye Phenol Red. This decolourisation was most efficient at pH 5.

a b s t r a c t ␣-l-Rhamnosidase [E. C. 3.2.1.40] cleaves terminal ␣-l-rhamnose specifically from a large number of natural products. The enzyme has wide occurrence in nature and is reported from animal tissues, plants, yeasts, fungi and bacteria. It is a biotechnologically important enzyme due to its applications in debittering and clearance of citrus fruit juices, enhancement of wine aromas and derhamnosylation of many natural products containing terminal ␣-l-rhamnose to compounds of pharmaceutical interests. Though ␣-l-rhamnosidases have been investigated actively during recent years, there is no recent review on ␣-l-rhamnosidases. An attempt has been made to fill up this gap in this review. It consists of a brief introduction of ␣-l-rhamnosidase which is followed by a critical description of the methods used for assaying the enzyme activity. Purifications, characterizations and properties of ␣-l-rhamnosidases from different sources have been discussed and the available structural and molecular biological studies on the enzyme have been given. Biotechnological applications of this enzyme in different processes have been briefly described. The review concludes with the identification of areas which needs further extensive studies.

Conidiation of an industrial strain of Penicillium (P.) camemberti, a ripening fungus, was examined on solid media. In order to evaluate the influence of nutritional factors on conidiation, we developed an inoculation and transfer procedure that allowed to obtain an homogenous mycelial biomass. Absence of conidiation was observed when ammonium sulphate and sodium nitrate were used as nitrogen sources. In contrast, conidiation increased significantly in the presence of ammonium phosphate or potassium nitrate with 4.8 Â 10 6 and 12 Â 10 6 spores ml À1 , respectively. By using those optimal conditions, the influence of nutrient starvation or calcium supply on conidiation was studied. Under the conditions tested, nitrogen starvation and calcium supply were better inducers than carbon starvation. A high carbon-to-nitrogen (C/N) ratio provided the highest level of with 4.2 Â 10 7 spores ml À1 and a sporulation index of 8.2 after 16 days of cultivation. #

The study proposes to get a better knowledge of the physicochemical properties of yeast extract (YE) molecules involved in the improvement of CHO cell growth and to reduce YE complexity without losing positive effects. Various chromatographic processes were implemented for fractionation of a nanofiltrated YE (nYE). The nYE was first fractionated by one-step preparative chromatography, either with anion exchange (AEC), hydrophobic interaction (HIC) or size exclusion (SEC) methods. After analysis of its main components, each fraction was added in a control chemically defined medium to assess its impact on CHO cell growth. Results mainly underlined that AEC was the most selective separation process to purify nYE in one step without decreasing cell growth promoting effect. A three-step chromatographic process including successive AEC, HIC, and SEC was then developed to refine the physicochemical properties of nYE compounds. Among fractions that triggered similar cell growth promoting effect than nYE, one also improved IgG specific production. It mainly included cationic and hydrophilic peptides with a great proportion of lysine and arginine, low quantities of polysaccharides and no nucleic acids. Thus, this study allowed us to deepen the YE contribution to animal cell culture as well as to evaluate fractionation strategies to simplify such a complex mixture.

The effect of magnetic fields (MF) on glutathione (GSH) production by Saccharomyces cerevisiae ATCC 7754 was studied. For this purpose, a factorial design of experiments was used to determine the influence of the time of exposure (8-16 h) and MF induction (25.0-34.3 mT), in GSH and biomass production. Additionally, control experiments (CE), without the application of MF, were performed. The results indicated the existence of favourable alterations in GSH and biomass concentrations due to the application of MF. In all experiments, the amount of biomass produced was higher than in CE and, with regard to GSH yield, in all the experiments at 24 and 48 h it was higher and in three experiments at 72 h of culture. The highest specific GSH yield (20.9 mg GSH /g biomass ), GSH yield (340.0 mg/L) and biomass (16.26 g/L) were obtained using a MF induction of 25.0 mT for 16 h. These results were 16.1%, 39.0% and 19.6% higher than in the CE, respectively. Through statistical analysis it was found that the MF induction was a significant factor in GSH yield, and also it was observed that, within the range of the experimental conditions used, the lower MF induction, the higher the GSH yield.

The performance of mesophilic anaerobic digesters of four large Italian wastewater treatment plants without primary sedimentation were studied. Only the waste activated sludge is stabilised by means of the mesophilic (35-37 8C) anaerobic digestion process. The anaerobic digesters generally worked with a hydraulic retention time in a range of 20-40 days and an organic loading rate of some 1 kg VS/m 3 reactor day. The solids content of the sludge fed to the digesters was in the range 2.6-3.9% and the gas produced per kilogram of volatile solids added was in the range 0.07-0.18 m 3 /kg VS fed . The specific gas production per kilogram of volatile solids destroyed was in the range 0.5-0.9 m 3 / kg VS destroyed and the reduction of the volatile solids concentration was in the range 13-27% (average 18%). These figures are particularly significant when designing anaerobic digesters for the treatment of waste activated sludge as single substrate. Moreover, it was observed that the higher the applied solid retention time in the activated sludge process for wastewater treatment, the lower the gas production. In particular, the specific gas production decreased from 0.18 to 0.07 m 3 /kg VS fed when increasing the solid retention time in the wastewater treatment line from 8 to 35 days. Finally, a mathematical model for the prediction of biogas production on the solid retention time applied in the wastewater treatment process was developed. #

A kinetic study was made of the anaerobic purification or biomethanation of wine distillery wastewaters ('vinasses '), using bioreactors containing various suspended clayey supports (sepiolite, bentonite and saponite), on to which the microorganisms eflecting the puriJication were immobilised. Assuming that the overall anaerobic fermentation process conforms to.first-order kinetics, experimental data pairs, namely the methane volume yielded (G) and the time (t), jitted the fo/Iowing equation: G = G, x (I-exp (K, x t)). The specific rate constant, K,, was determined in each of the situations studied. The support used has a marked influence on the kinetic constant of the process,. the saponite support yielding sign$cantIy the highest values. On the other hand, the specific rate constant decreased over the chemical oxygen demand (COD) range studied (I-MI g/litre) when the volume of wastewater added or substrate concentration was increased; this showed an inhibition phenomenon. Also, the mean rate biogas production and the methanogenic activity decreased irrespective of the support used. The yield coeficient, Y,, was 0.32 litres CH., STP/g COD.

Proteases have applications in food, detergent and pharmaceutical industries. A novel protease has been purified from the latex of Calotropis procera and characterized. As another cysteine protease, procerain, is reported from the same source, the newly purified enzyme was named as procerain B. The enzyme shows distinct properties compared to procerain, in terms of cleavage recognition site, immunological properties and other physical properties like molecular weight, isoelectric point, etc. The newly purified enzyme shows a broad optimum pH (6.5-8.5) as well as broad optimum temperature (40-60 8C). Additionally, the enzyme retains its activity where most of other proteases are not active. Moreover, the enzyme appeared to be very efficient in hydrolysis of blood stain and may have potential application in detergent industries. Simple and economic purification of procerain B, together with easy availability of latex, makes the large-scale production of procerain B possible, thus enables to explore various industrial as well as biotechnological applications. ß

We have investigated the changes of microbial community structures and the concomitant performance in two biological wastewater treatment systems (conventional and inverted A 2 /O processes) over a whole cycle of sludge bulking. A low level of filament abundance was observed during non-bulking period, with types 0092 and 0041 as the dominant filamentous bacteria. With the increase of the sludge volume index values from 76 (73) to 275 (300) mg/L, the filament abundance estimated by microscopic examination increased from 1 (few) to 5 (abundant), with Microthrix parvicella becoming the dominant filament bacteria. Sludge bulking resulted in a significant shift in bacterial compositions from Proteobacteria to Actinobacteria dominance, characterized by the significant presence of filamentous M. parvicella (from not detected to higher than 60% of clones) and decrease of the dominant Betaproteobacterial population (from higher than 40% to less than 1%). Important relevant bacterial populations including polyphosphateaccumulating organism (PAO, Candidatus Accumulibacter phosphatis), ammonia-oxidizing bacteria (AOB, Nitrosomonas), nitrite-oxidizing bacteria (NOB, Nitrospira) and denitrifying bacteria (Thauera) were absent under the serious bulking condition. Accumulation of nitrite and ammonia was observed during serious bulking, while the phosphorus removal performance was not decreased because M. parvicella could behave as a PAO.

The ability of two plant materials, Cactus latifaria and the seeds of Prosopis juliflora, to act as natural coagulants was tested using a synthetic water formulated to resemble the drinking water supplied to the city of Maracaibo. Turbidity was added as kaolin. The coagulation ability of the two materials was assessed by the use of standard jar test measurements. Both materials produced comparable turbidity removals and were able to produce a final water whose turbidity was close to the required standard of 5NTU with both high (100-200 NTU) and low (30 -40 NTU) initial turbidities. This was comparable with the performance achieved by previous workers using Moringa oleifera extracts. The optimum coagulant dose was found to be lower than that for aluminium sulphate. Treatment of the raw vegetable solids with solvents produced coagulants which, in some cases, were different from the raw materials and suggested that non-polar material might be involved with the coagulation process.

The feasibility of sulphate removal from sulphate-rich wastewater using an anaerobic fixed-bed reactor was investigated. The bioreactor was installed at a chemical industry producing organic peroxides, which generate wastewater with sulphate concentrations ranging from 12,000 to 35,000 mg SO 4 2 − l − 1 . A pilot-scale anaerobic fixed-bed reactor with a 94.2-l volume was tested to treat part of the wastewater. The reactor was filled with 1-cm 3 polyurethane foam cubes and operated, initially, in discontinuous regime. Five batch tests were performed with diluted industrial wastewater. The sulphate reduction efficiency and the chemical oxygen demand (COD) removal efficiency were evaluated as a function of the COD to [SO 4 2 − ] ratio in each batch test. The effect of the addition of supplementary ethanol on the sulphate-reducing bacteria growth was also evaluated. The reactor was then fed in a semi-continuous regime with raw industrial wastewater with high sulphate concentration. The addition of ethanol stimulated the sulphate-reducing bacteria, which predominated over the methane-producing organisms even at a high COD to [SO 4 2 − ] ratio. A maximum sulphate removal efficiency of 97% was reached during discontinuous and semi-continuous operations.

Invertase (EC.3.2.1.26) belongs to glycoside hydrolases that catalyzes sucrose (table sugar) into two monosaccharides i.e. glucose and fructose. Invertases produced by a large number of microorganisms having molecular mass in the range of 47-430 kDa, K m value from 0.063 to 470 mM, K cat from 5.68 to 7.4 × 10 6 s −1 and half-life of 2.42-78.8 min (at 65 • C) worked optimally between temperature of 30-75 • C and pH of 2.9-6.2. They are extensively used in the preparation of jams, marshmallows, infant formulas, candies containing liquefied sugar centers, chocolate covered cherries, digestive aid tablets, artificial honey, beverages and bakery items. Invertases are also important for the prevention of diseases (ulcers, intestinal ailments, reduce colds etc.) in humans, in anti-aging process and physical rejuvenation. Their ability to convert glucose into hydrogen per oxide categorize them as powerful antimicrobial and antioxidant agents, hence, assist in defense reaction against bacterial infections and gut fermentation by oxidation. Microbial invertases have been extensively studied during last decades. However, their wider applications ranging from industry to human health demands a comprehensive review on production, purification, properties and their benefits.

Several non-pathogenic yeast species are described to synthesize sophorolipids: glycolipid molecules with surface tension lowering capacities. These so-called biosurfactants offer an environmental friendly alternative for the petrochemical derived surfactants used in various sectors and attract quite some industrial attention due to the high yields of over 400 g/L which can be obtained by cultivations with Candida bombicola. On top of their surface lowering properties, sophorolipids display several interesting biological activities, allowing applications in the cosmetics, pharmaceutical and medical field. In this review, various aspects of the sophorolipid production process are outlined as well as recent findings on the biosynthetic pathway. Furthermore, the synthesis of modified sophorolipids derived either microbial or by post-fermentative modification is discussed. Finally, recent and novel applications of sophorolipids are summarized.

Whole cells of bakers’ yeast (Saccharomyces cerevisiae) exhibited very low catalase activity (16 U g−1). Various treatments altered membrane permeability and increased cellular catalase activity. Treatment with cetyltrimethylammonium bromide (CTAB) yielded maximum catalase activity (51000 U g−1) followed by digitonin (27072 U g−1). After permeabilization of the membrane, the catalase slowly leaks out of the cells. The half-life of intracellular catalase in CTAB-permeabilized cells was 3.4 days at 4°C and 1.2 days at 26°C. Catalase in permeabilized cells was more stable against self-inactivation during catalysis than the enzyme in cell-free extract and purified catalase. Catalase of CTAB-permeabilized cells was effective in removing the residual H2O2 from H2O2-treated fresh milk/heat pasteurized milk.

A process for producing non-monosaccharide and high-purity galactooligosaccharides (GOS) was proposed. GOS were produced from lactose by Penicillium expansum F3 β-galactosidase immobilized in calcium alginate. The effects of pH, temperature and lactose concentration on GOS production were investigated. A purity of 28.7% (w/w) GOS was obtained from 380 g/L lactose solution at pH 5.4 and 50 °C. The immobilized enzyme was used for repeated GOS synthesis and showed good operational stability. Digestible sugars in the GOS could be depleted after fermentation with Saccharomyces cerevisiae L1 or Kluyveromyces lactis L3 entrapped in the calcium alginate. Purity greater than 37% with yields greater than 27% of non-monosaccharide GOS were maintained by S. cerevisiae L1 for 19 batches and purity greater than 97% with a yield greater than 20% of high-purity GOS was produced using K. lactis L3 for two batches.

The antioxidant and antimicrobial activities of various solvent extracts from Mirabilis jalapa tubers (MJT) were investigated using various in vitro assays. The total phenolic and flavonoid contents varied from 21.45 to 364.6 mg gallic acid equivalent (GAE)/g dried extract and 5.2 to 71.6 mg quercetin/g dried extract, respectively. Water extract of MJT was the most potent antioxidant in all assays used, followed by methanol extract. The five solvent extracts were screened for antibacterial and antifungal activities. Water extract was the most effective with minimum inhibitory concentration <200 g/ml against Staphylococcus aureus, Micrococcus luteus, Pseudomonas aeruginosa, Klebsiella pneumoniae, Bacillus cereus and Enterococcus faecalis. Only water extract showed antifungal activity against Aspergillus niger, Fusarium solani, Fusarium oxysporium and Fusarium granularium. GC/MS analysis of MJT dichloromethane and methanol extracts showed that oleic acid and ␤-sitosterol were, respectively, the major compounds. LC/MS analysis of the aqueous extract showed a high content of flavanol and flavonol compounds. Phenolic acids such as ferulic and caffeic acid were also detected.

The amino acid composition of CHO cell culture media for manufacturing an antibody fusion protein (B1) was optimized using data obtained through metabolic flux analysis. The metabolic flux analysis was performed using the metabolic rates derived from two semi-steady states of continuous culture. The amino acid composition was the same for media used in both the fed-batch and continuous cultures, which made the modification of amino acid composition suggested by the metabolic flux analysis applicable to the fed-batch culture. The flux analysis indicated that methionine, tryptophan, asparagine and serine were limiting, while alanine, arginine, glutamine and glycine were in excess at the semi-steady states. Based on this result, the limiting amino acids were increased while those in excess were reduced to obtain balanced amino acid composition in both the batch and feed media. The fed-batch cell cultures using the modified media generated less lactate and ammonia, and more significantly, increased peak cell densities by 55% and B1 protein titer by 27% without any impact on final sialic acid content, a quality attribute of the B1 protein.

The stability of penicillin G was analyzed as a function of temperature, pH, and the combined effects of pH and temperature were investigated by proposing a second-order polynomial model for penicillin G decomposition reaction rate constant. The modeling results obtained for the first-order decomposition reaction of penicillin G are in good agreement with the experimental data obtained at the pH range of 1.8-10.0 and temperature range of 0 -52°C. The stability analysis shows that penicillin G is more stable at the pH range of 5.0-8.0 than out of it and lower temperatures for subsequent isolation. The maximum stability of penicillin G is at about pH 6.0 and its instability its much higher at acidic than basic pH values. The stability of penicillin G decreases by increase in temperature for all pH values. The stability results as well as the proposed second-order model for penicillin G decomposition reaction rate constant can be used for investigating penicillin G losses during its isolation process.

In this study, the optimum operating conditions of a lab-scale completely stirred acidogenic reactor treating a chemical synthesis-based pharmaceutical wastewater were determined. Individual effects of hydraulic retention time (HRT), organic loading rate (OLR) and pH on the degree of acidification were investigated at a mesophilic temperature (35 AE 1 8C), for HRT in the range 8-24 h, OLR up to 14 kg COD/m 3 d and pH in the range of 5.0-6.3. A maximum acidification level of 44% and COD removal efficiency of 13%, were obtained at an OLR of 13 kg COD/m 3 d at pH of 5.5 AE 0.1 with an HRT of 12 h. Under these conditions, acetic, propionic and n-butyric acids were found to be predominant volatile fatty acids in the acidogenic reactor. #

A novel toxic protein from Glycine max seeds, named Gm-TX, was purified by combination of differential precipitation with ammonium sulphate, DEAE-cellulose, anhydrotrypsin-sepharose 4B and Superdex 200 HR fast-protein liquid chromatography. Gm-TX is composed of one polypeptide chain of 28 kDa (SDS-PAGE), with pI 5.1-5.2, contains <0.5% neutral sugar, has N-terminal sequence comprised of KTISSEDSPFFNCREK, maximum absorption spectrum at 280 nm and molar extinction coefficient of 16.9. It exhibited ribonuclease activity (1821.42 AE 3.34 UA/h/mgP) toward yeast RNA and promoted rabbit erythrocyte agglutination (120.4 HU/ mgP) mediated by anti-Gm-TX IgG, which was inhibited by mannose. It was devoid of trypsin inhibitory, urease and chitinase activities. Gm-TX was lethal to mice by intraperitoneal route with LD 50 of 4.5 mg/kg. Moreover, it showed dose-dependent insecticidal activity on the major economically important pests Callosobruchus maculatus and Dysdercus peruvianus, producing ca. 50% and 56% mortality when incorporated into artificial diets at a level of 1.16% and 0.25%, respectively. These data indicate that Gm-TX is a candidate protein that could be expressed in genetically transformed plants for improved resistance to insect pests.

A sequencing batch reactor (SBR) has certain advantages over conventional activated sludge processes (ASP) for the treatment of complex wastewater. The performance of a sequencing batch reactor in treating complex chemical effluents was investigated with a suspended biomass configuration and operating under aerobic conditions. A total sequence of 24 h (15 min: filling phase; 23 h: reaction phase (aeration with recirculation); 30 min: setting; and 15 min: withdrawal) was employed and studied with various organic loading rates (1.0 kg COD/m3/day: 1.7 kg COD/m3/day and 3.5 kg COD/m3/day). The SBR performance was assessed by means of carbon removal and operational parameters such as pH, oxidation–reduction potential (ORP), sludge volume (SV), sludge volume index (SVI), suspended solids (SS) and volatile suspended solids (VSS), which were monitored during reactor operation. The SBR showed relatively more efficient performance over conventional suspended growth systems. About 8% sulphate was removed under anoxic conditions at all the studied organic loading rates. Enhanced performance with SBR over conventional ASP may be due to enforced short term unsteady state conditions coupled with periodic exposure of the microorganisms (physiological state) to defined process conditions which facilitate the required metabolic conditions for treating complex chemical effluents.