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Diatoms are currently considered valuable feedstocks for different biotechnological applications. To deepen the knowledge on the production of these microalgae, the diel pattern of batch growth, photosystem II performance, and accumulation of target metabolites of two commercially relevant diatoms, Phaeodactylum tricornutum and Skeletonema costatum, were followed outdoors in 100-L flat panel photobioreactors. S. costatum presented a higher light-to-biomass conversion resulting in higher growth than P. tricornutum. Both fluorescence data and principal component analysis pointed to temperature as a limiting factor for the growth of P. tricornutum. Higher protein and carbohydrate contents were found in P. tricornutum, whereas S. costatum fatty acids were charac-terized by a higher unsaturation degree. Higher productivities were found at 1 p.m. for protein, lipid, and ash in the case of S. costatum. Overall, S. costatum showed great potential for outdoor cultivation, revealing a broader temperature tolerance and increased biomass productivity than P. tricornutum. info:eu-repo/semantics/publishedVersion

Aim of this study The major aim of this work was to consistently optimize the production of biomass of the dinoflagellate Karlodinium veneficum and evaluate its extracts biotechnological potential application towards food, nutraceutical or/and pharmaceutical industries. Methods and results A successful approach of biomass production of K. veneficum CCMP 2936 was optimized along with the chemical characterization of its metabolite profile. Several temperatures (12, 16, 20, 25, 30°C), L1 nutrient concentrations (0.5×, 2×, 2.5×, 3×) and NaCl concentrations (20, 25, 30, 40 g L−1) were tested. The growth rate was maximum at 16°C, 2× nutrient concentration and 40 g L−1 of NaCl; hence, these conditions were chosen for bulk production of biomass. Methanolic extracts were prepared, and pigments, lipids and phenolic compounds were assessed; complemented by antioxidant and anti-inflammatory capacities, and cytotoxicity. Fucoxanthin and derivatives accounted for 0.06% of dry weight, and up to 60% (w/w) of all quantified metabolites were lipids. Said extracts displayed high antioxidant capacity, as towards assessed via the NO•− and ABTS•+ assays (IC50 = 109.09 ± 6.73 and 266.46 ± 2.25 µgE ml−1, respectively), unlike observed via the O2•− assay (IC25 reaching 56.06 ± 5.56 µgE ml−1). No signs of cytotoxicity were observed. Conclusions Karlodinium veneficum biomass production was consistently optimized in terms of temperature, L1 nutrient concentrations and NaCl concentration. In addition, this strain appears promising for eventual biotechnological exploitation. Significance and Impact of the Study This work provides fundamental insights about the growth and potential of value-added compounds of dinoflagellate K. veneficum. Dinoflagellates, as K. veneficum are poorly studied regarding its biomass production and added-value compounds for potential biotechnological exploitation. These organisms are difficult to maintain and grow in the laboratory. Thus, any fundamental contribution is relevant to share with the scientific community.

Fermented olives are an important vehicle for delivering probiotics, contributing to magnify their known health benefits, especially in view of their efficacy to prevent or ameliorate specific chronic diseases by influencing the bioavailability and activity of particular bioactive compounds. This chapter attempts to provide an overview of available information on olive fermentation phenomena, and to make a claim for health-promoting nutrients and probiotics from fermented olives, aiming at a better understanding of its potential toward improvement of health.

Consumption of functional probiotic foods has increased over recent decades, alongside with increasing consumer and researcher awareness of their health-promoting effects. This has prompted an interest toward development of novel functional food formulations. Despite fermented dairy products still remaining the most common vectors for the delivery of probiotics to humans, such other food matrices as fruits and vegetables offer a promising performance as sources and carriers of probiotic strains. Hence, these types of matrices are thoroughly reviewed here – with table olives being subjected to comprehensive discussion as a case study, owing to their suitable microstructure and unique sensory and nutritional features.

Metal contamination of a few aquatic, atmospheric, and soil ecosystems has increased ever since the industrial revolution, owing to discharge of such elements via the effluents of some industrial facilities. Their presence to excessive levels in the environment will eventually lead to serious health problems in higher animals owing to accumulation throughout the food web. Current physicochemical methods available for recovery of metal pollutants (e.g., chemical precipitation, oxidation/reduction, or physical ion exchange) are either expensive or inefficient when they are present at very low concentrations. Consequently, removal of toxic metals by microorganisms has emerged as a potentially more economical alternative. Microalgae (in terms of both living and nonliving biomass) are an example of microorganisms suitable to recover metals and able to attain noteworthy percent removals. Their relatively high metal-binding capacities arise from the intrinsic composition of their cell walls, which contain negatively charged functional groups. Consequently, microalgal cells are particularly efficient in uptake of those contaminants when at low levels. Self-defense mechanisms developed by microalgal cells to survive in metal-containing media and environmental factors that affect their removal (e.g., pH, temperature, and biomass concentration) are reviewed here in a comprehensive way and further discussed in attempts to rationalize this form of remediation vis-a-vis with conventional nonbiological alternatives.

Broa is an example of bread that is a good candidate for inclusion in functional diets, so it deserves further in-depth study of its chemical composition-namely with regard to evolution of the lipid profile throughout breadmaking, in order to assess whether mixing, fermentation, or baking affect its nutritional value (in terms of unsaturated fatty acids, UFA) based on the assumption that neutral lipids (NL) can be protein- or carbohydrate-bound. Hence, constituent fatty acids in NL of maize (Zea mays) and rye flour (Secale cereale), and in sourdough and final broa manufactured from a mixture therefrom were quantitated. Methodologies of esterification of fatty acids, as well as of transesterification of acyl lipids and sterol esters (SE) were improved. The n-hydrocarbons containing between 4 and 24 carbon atoms were then resolved and identified by gas-liquid chromatography. Regarding total neutral lipids (TNL) in all samples, 79-89% were TAGs, and 87-93% were TAGs and DAGs in the case of free lipids (FL). Furthermore, 73-85% of TNL in bound lipids (BL) and 65-80% of TNL in starch lipids (SL) were TAG and free fatty acids (FFA). Conversely, only 4-5%, 6-16%, and 7-10% of TNL in FL, BL, and SL, respectively, were SE and MAGs. TAGs and DAGs underwent partial hydrolysis during fermentation and baking; palmitic, oleic, and linoleic acids were dominant as products released. The nutritional value of broa lipids was apparent owing to their proportions of SE (4%) and DAG (9%), coupled with 52% of linoleic acid in all samples-as well as to the high contents of polyunsaturated versus monounsaturated or saturated fatty acids, and to the general dominance of UFA.

Molecular hydrogen (H 2 ) obtained from biological sources provides an alternative to bulk chemical processes that is moving towards large-scale, economical generation of clean fuel for automotive engines. This opinion article examines recent improvements in H 2 production by wild and mutant strains of Chlamydomonas reinhardtii – the green microalga currently considered the best eukaryotic H 2 producer. Here, we review various aspects of genetic and metabolic engineering of C. reinhardtii , as well as of process engineering. Additionally, we lay out possible scenarios that would lead to more efficient research approaches in the near future, as part of a consistent strategy for sustainable biohydrogen supply.

A number of enzymes have applications in the textile sector, as several natural fibres (e.g. cotton, wool and flax) can be subjected to processing using such natural biocatalysts. In the latest two decades, demand for industrial enzymes has increased considerably; however, the textile industry requires highly stable enzymes, and good performance at extreme values of pH and temperature. New strategies are continuously emerging for the immobilization of enzymes with superior efficiency and usage. Enzymatic immobilization can stabilize enzymes and extend their useful life. Additionally, reduction in effluent treatment costs and improvement of efficiency is possible. This paper briefly reviews the most recent research efforts pertaining to immobilization of enzymes with potential application in the textile industry.

Carbon-neutral renewable liquid biofuels are needed to displace petroleum-derived transport fuels in the near future - which contribute to global warming and are of a limited availability. A promising alternative is conveyed by microalgae, the oil content of which may exceed 80% (w/wDW) - as compared with 5% of the best agricultural oil crops. However, current implementation of microalga-based systems has been economically constrained by their still poor volumetric efficiencies - which lead to excessively high costs, as compared with petrofuel prices. Technological improvements of such processes are thus critical - and this will require a multiple approach, both on the biocatalyst and bioreactor levels. Several bottlenecks indeed exist at present that preclude the full industrial exploitation of microalgal cells: the number of species that have been subjected to successful genetic transformation is scarce, which hampers a global understanding (and thus a rational design) of novel blue-biotechnological processes; the mechanisms that control regulation of gene expression are not fully elucidated, as required before effective bioprocesses based on microalgae can be scaled-up; and new molecular biology tools are needed to standardize genetic modifications in microalgae - including efficient nuclear transformation, availability of promoter or selectable marker genes, and stable expression of transgenes. On the other hand, a number of pending technological issues are also present: the relatively low microalga intrinsic lipid productivity; the maximum cell concentration attainable; the efficiency of harvest and sequential recovery of bulk lipids; and the possibility of by-product upgrade. This review briefly covers the state of the art regarding microalgae toward production of biofuels, both from the point of view of the microalgal cell itself and of the supporting bioreactor; and discusses, in a critical manner, current limitations and promising perspectives in this field.

Reactive forms of oxygen can damage DNA (among other molecules), thus triggering, e.g., atherogenesis and carcinogenesis. However, such dietary antioxidants as lutein and β-carotene can effectively inactivate them; these compounds were found to high levels in a novel strain (M2-1) of the microalga Scenedesmus obliquus. The independent and combined effects of pH and temperature on its rates of growth and production of antioxidants were experimentally assessed, via a full factorial experimental design; the effects of each parameter independently, and of their interactions were accordingly quantified by ANOVA. Our results indicated that temperature plays a more important role on the maximum specific growth rate than pH; in terms of antioxidant content, pH and, to a lesser extent, temperature also have relevant effects. Consequently, the highest rate of biomass specific growth (0.294 ± 0.013 day−1) and biomass productivity (0.837 ± 0.054 mg L−1 day−1) were associated with relatively low pH (6) and relatively high temperature (30°C). Conversely, the antioxidant production rate increased with pH; hence, the highest productivity (0.638 mg L−1 day−1) was attained at pH 8 and 30°C. At the best operating conditions for antioxidant content, the levels of lutein and β-carotene were 203.57 ± 1.41 and 18.20 ± 0.33 mg mL−1, respectively; the maximum production of either one occurred at the early exponential phase. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011

This research effort was aimed at assessing the changes in extractable neutral lipids (NL) throughout the baking process of Broa, a Portuguese traditional sourdough bread. NL were accordingly isolated, purified and quantitated - starting from non-starch lipid (NSL) and starch lipid (SL) extracts of maize and rye flours, as well as fermented dough and bread. NSL accounted for the major fraction of extracted lipids; furthermore, the NSL/SL ratio evolved throughout processing in agreement with the phenomena prevailing during dough preparation, fermentation, and baking. An analytical method based on resolution by normal-phase HPLC coupled with detection by evaporative light scattering was accordingly developed for quantitation of the aforementioned NL classes. Distinct NL classes correlated well with the stage of bread making. The main NL in NSL were triacylglycerols (ca. 75% of the total), but relatively high concentrations of sterol esters and diacylglycerols were also found. Conversely, free fatty acids were the dominant component of SL, whereas monoacylglycerols and free sterols were comparable to those in NSL.

The behavior of Bifidobacterium animalis subsp. lactis Bb 12 under batch cultivation, after continuous culturing for up to 12 d, was monitored in skim milk-based media. Previous continuous culture for longer than 6 d affected the physiology of said microorganism. The minimum inhibitory concentrations of lactic and acetic acids increased from 18 to 26 g/l, whereas the molar ratio of acetic to lactic acid increased from 0.8 to 1.55, when the previous continuous culture increased its duration from 1 to 12 d. The specific lactose consumption rate decreased from 0.94 to 0.77 glactose/gcell dry mass/h within the batch culture timeframe; this was concomitant with greater amounts of acetic and formic acids, and lower amounts of lactic acid produced. The β-galactosidase activity increased as continuous culturing time increased, and reached 446 units/ml by 12 d; however, the rate of enzyme synthesis decreased concomitantly. Succinic acid was produced during the exponential growth and stationary phases of the batch culture, but the former at exponential growth phase was higher as the continuous culturing time was longer. For comparison purposes, batch cultivation of samples taken from continuous cultures by 1 and 12 d was done using a semi-synthetic medium with glucose as carbon source; a pattern similar to that observed when using skim milk-based media was observed.

Cheese is a classical dairy product, which is strongly judged by its appearance and texture; hence, a renewed interest in its microstructure has been on the rise, as sophisticated techniques of analysis become more and more informative and widely available. Processing parameters that affect microstructure play a dominant role upon the features exhibited by the final product as perceived by the consumer; rational relationships between microstructure (which includes biochemical and microbiological indicators), and quality and safety of the products are accordingly required. Subsequent to that extra fundamental knowledge, technological innovations may eventually improve current cheesemaking processes, and permit mechanistic design of novel ones. This review thus focuses on recent advances pertaining to the microstructure of cheese, and discusses them in a logical and critical manner.

The aim of this research effort was to investigate the role of various sugar substrates in the growth medium upon thermotolerance and upon survival during storage after freeze-drying of Lactobacillus bulgaricus. Addition of the sugars tested to the growth medium, and of these and sorbitol to the drying medium (skim milk) was investigated so as to determine whether a relationship exists between growth and drying media, in terms of protection of freeze-dried cells throughout storage. The lowest decrease in viability of L. bulgaricus cells after freeze-drying was obtained when that organism was grown in the presence of mannose. However, L. bulgaricus clearly survived better during storage when cells had been grown in the presence of fructose, lactose or mannose rather than glucose (the standard sugar in the growth medium). A similar effect could not be observed in terms of thermotolerance; in this case, the growth medium supplemented with lactose was found to yield cells bearing the highest heat resistance. Supplementation of the drying medium with glucose, fructose, lactose, mannose or sorbitol led in most cases to enhancement of protection during storage, to a degree that was growth medium-dependent.

The influence of light intensity upon biomass and fatty acid productivity by the microalga Pavlova lutheri was experimentally studied using a novel device. This device was designed to automatically adjust light intensity in a photobioreactor: it takes on-line measurements of biomass concentration, and was successfully tested to implement a feedback control of light based on the growth rate variation. Using said device, batch and semicontinuous cultures of P. lutheri were maintained at maximum growth rates and biomass productivities – hence avoiding photoinhibition, and consequent waste of radiant energy. Several cultures were run with said device, and their performances were compared with those of control cultures submitted to constant light intensity; the biomass levels attained, as well as the yields of eicosapentaenoic and docosahexaenoic acids were calculated – and were consistently higher than those of their uncontrolled counterpart.

BACKGROUND: Aqueous extracts of 48 herbal plants were obtained via alternative extraction protocols, and were assayed for their capacity to protect deoxyribose and DNA itself from degradation (or, conversely, for their capacity to promote DNA degradation), using electrophoresis as analytical tool. RESULTS: For a given (constant) volume of extract, deoxyribose protection ranged from 14.13 ± 1.35% (mean ± SD) inhibition by dwarf mallow powder infusion, up to 106.51 ± 15.93% inhibition by avocado powder infusion. DNA protection was tested at two extract concentrations, and was slightly greater at the higher concentration. Pro-oxidant effects were essentially absent. CONCLUSION: The anti-oxidative roles of plants upon deoxyribose and DNA displayed by our experimental results were rather promising with regards to practical applications of those plants, viz. as ingredients in the formulation of nutraceutical beverages and/or foods. Copyright © 2007 Society of Chemical Industry

BACKGROUND: Aqueous extracts of most medicinal plants traditionally employed in Portugal (at the ratio of 1 g plant: 110 mL water) have been assayed for total antioxidant capacity and phenol content, in order to elucidate their claimed medicinal features. RESULTS: The antioxidant activity was assessed by the ABTS•+ method; the ascorbic acid equivalent values ranged from 1.4280 ± 0.1261 g L-1 for avocado (Persea americana (Lauraceae)) obtained by infusion of powder, down to 0.0027 ± 0.0012 g L-1 for olive (Olea europaea (Oleaceae)) obtained by infusion of leaves. Total phenol content was determined by the Folin-Ciocalteu procedure; the gallic acid equivalent values ranged from 0.5541 ± 0.0289 g L -1 for avocado obtained by infusion of powder, down to 0.0053 ± 0.0014 g L-1 for olive obtained by boiling leaves. A good correlation between total antioxidant capacity and total phenol content was found. CONCLUSION: The method of powder infusion should be chosen if high concentration of antioxidants are sought. On the other hand, a high antioxidant capacity and a high phenol content correlate well with the empirically established (and widely publicised) capacity to treat respiratory infections.

Yacon [Smallanthus sonchifolius (Poepp. and Endl.) H. Robinson; Asteraceae] roots have been shown to be a source of prebiotic compounds. However, there are no known studies concerning processed yacon roots. The objective of this study was to investigate the potential prebiotic activity of yacon tuber flour. For this purpose, an aqueous extract was tested for selection of yacon incorporation and sterilization method and selection of the most favourable concentration to be tested for prebiotic activity. Once these conditions were identified, the potential prebiotic activity of the yacon extract was evaluated by determination of viable cell numbers and metabolic activity against four probiotic strains, namely, Enterococcus faecium 32, Bifidobacterium animalis Bo, Lactobacillus acidophilus Ki and Lactobacillus casei L26). Results showed that the best incorporation and sterilization method was to autoclave the supernatant, resultant from the yacon tuber flour suspension, at 121 degrees C for 20 min and add it to sterilized basal medium. For the confirmation of potential prebiotic activity, de Man-Rogosa-Sharpe (MRS) medium without a conventional carbon source (negative control), with 2% (w/v) glucose per se (positive control) and associated with 1% (w/v) yacon tuber flour were chosen. Yacon tuber flour revealed a potential prebiotic activity upon the growth of the probiotic strains tested, probably due to its fructooligosaccharides (FOS) content. (C) 2015 The Institution of Chemical Engineers.

Fisheries are the most important sources of feedstock for fishmeal. Only a small percentage of global fish production is indeed channeled to human consumption, with the remainder being used for fish and animal feed. Fishmeal is a protein-rich food, and sets the basis for any balanced formulation used in commercial aquaculture. For several reasons there is a definite need for a new source of nutritious fish food. Therefore, the potential use of unconventional feed ingredients, such as microalgae, as feed inputs to replace high cost feed stuffs has been increasing. Microalgae are a more reliable and less volatile source of protein, and their availability is not dependent on fish captures. This provides industry with a better control of their costs, and supports a potential for future investment due to the reduction of risk in aquaculture farming operations.

Use of probiotic bacteria and consumes in large — in novel foods to provide beneficial health effects has attracted an increasing interest by the food industry and fermented olives are an excellent example of a new generation of those foods from plant origin so as to assure maximum viability by the time of ingestion during processing and storage of food products, as well as during transit through the gastrointestinal tract. Our study focused on production, characterization and assessment of efficacy of microencapsulation upon survival of probiotic strains and sensory properties of the final olive paste throughout refrigerated storage. Microencapsulation appears to be an effective technique for strain survival, depending on the operating temperature and experimental results on tolerance to gastrointestinal-like conditions, and ability to adhere to intestinal epithelium is thereby presented and discussed. The sensory panel rated all experienced matrices as good, including overall acceptance without significant preference between them. However, the success of microencapsulation was more limited when incorporated into olive paste. Free cells of Lactobacillus plantarum 33 proved able to survive in olive paste during storage at refrigerated temperatures.

From the extracellular fluid of a novel strain of Bjerkandera sp., it was isolated, purified and identified the main enzyme responsible for Remazol Brilliant Blue R dye decolourisation. Such an enzyme is able to oxidise manganese, as well as veratryl alcohol and 2,6-dimethoxyphenol in manganese-independent reactions; hence, it can be included in the new group of versatile peroxidases. The molecular mass of said enzyme is ca. 45 kDa, and the N-terminal amino acid sequence obtained by Edman degradation is VAXPDGVNTA. The enzyme substrate range for oxidation of several phenolic and non-phenolic aromatic compounds was determined and the corresponding Michaelis–Menten kinetic constants calculated. Furthermore, spectrophotometric assays showing the Soret band and allowing observation of band shifts of the enzyme led to the conclusion that Bjerkandera strains may also synthesise at least two different versatile peroxidases, as happens with Pleurotus eryngii .

Four strains of lactic acid bacteria, previously isolated from traditional Serra da Estrela cheese and duly identified as Lactobacillus paracasei ssp. paracasei, Leuconostoc mesenteroides ssp. dextranicum, Lactococcus lactis ssp. lactis and Enterococcus faecium, were tested for their aminotransferase, oxidase and dehydrogenase activities towards five amino acids in free form: two branched-chain (leucine and valine), one sulfur-containing (methionine) and two aromatic (phenylalanine and tryptophan) amino acids. For this purpose, both resting cells (RC) and crude cell-free extracts (CFE) were considered; their lyase and demethiolase activities (towards methionine and α-keto-γ-methiolbutyric acid (KMBA), respectively) were also evaluated. Aminotransferase activity (AT) of RC was ca. 10-fold higher towards Met than towards Leu or Val. No AT activity was found in CFE, although these extracts displayed high dehydrogenase activity (DA) towards Phe, especially L. lactis ssp. lactis and E. faecium. In the case of L. paracasei ssp. paracasei and L. mesenteroides ssp. dextranicum, they showed high DA activity, which was ca. 10-fold higher towards Met than towards Leu or Val. E. faecium displayed high oxidase activity towards Met, and produced nine-fold more methanethiol from Met and five-fold more methanethiol from KMBA than the other three isolates under study.

The optimization and control of biochemical processes require the previous establishment of mathematical models that can describe the effect of process variables on their actual kinetics. Environmental temperature is a modulating factor to which the algal cells respond continuously by adjusting their rates of cellular reactions, their nutritional requirements, and, consequently, their biomass composition. Light intensity is an exhaustible resource, indispensable to autotrophic organisms. The effects of light intensity and temperature on growth of the microalga Pavlova lutheri, which have hardly been considered to date in a simultaneous fashion, were experimentally assessed using a factorial experimental design; in this way, the effects of each variable independently and their interactions could be quantified, using maximum biomass (X max) or maximum specific growth rate (μmax) as objective functions. The preliminary results produced indicated that light intensity plays a more important role on μmax than temperature; in the case of Xmax, both temperature and, to a lesser extent, light intensity do apparently play a role. The highest values of Xmax were associated with low temperatures and high light intensities; a similar behavior could be observed for μmax concerning light intensity, although the dependency on temperature did not seem to be as important. A more complex mechanistic model was then postulated, incorporating light and temperature as input variables, which was successfully fitted to the experimental data generated during batch cultivation of P. lutheri.

An intracellular hydrolase able to cleave l -lysine-p-nitroanilide was purified from Lactobacillus plantarum strain ESB5004 via two steps of precipitation with ammonium sulfate (at 30 and 50% (w/v)), followed by hydrophobic interaction and ion-exchange chromatographies. The aminopeptidase was purified up to 11-fold, with a final yield of ca. 1%. Its native molecular weight is ca. 70 kDa, and it is apparently composed of two subunits, the molecular weight of which is 34 kDa. The enzyme was assayed using a wide variety of p-nitroanilide (pNA) derivatives as substrates: it hydrolyzed preferentially pNA adducts of hydrophobic and basic amino acid residues; no hydrolysis was in particular observed of Glu-pNA, Gly-pNA or Pro-pNA. The enzyme activity was removed by the metal-chelating agent EDTA, thus suggesting that it is a metallo-enzyme; however, the EDTA-inhibited enzyme was reactivated in the presence of Co2+. Optimal aminopeptidase activity was obtained at 28 °C (pH 7.0) and pH 6.5 (37 °C). The enzyme was inhibited by 10 mM CaCl2 or MgCl2.

The high commercial values of eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids have driven a strain-improvement program, aimed at increasing the content of those fatty acids in the microalga Pavlova lutheri (SMBA 60) as parent strain. After a round of mutation using UV-light as mutagenic agent, an isolate strain (tentatively called II#2) was obtained, the EPA and DHA contents of which (in % dry biomass) were 32.8% and 32.9% higher than those of the control, native strain. The final EPA yields, when the cultures were maintained under appropriate conditions, were 17.4 and 23.1 mg. g(-1) dry biomass, for the wild-type and the II#2 strain, respectively, whereas the final DHA yields were 8.0 and 10.6 mg. g(-1) dry biomass, respectively. These results suggest that random mutagenesis can successfully be applied to increase the yield of n-3 fatty acids by microalgae.

No significant differences were observed in the viability of Lactobacillus plantarum and Lactobacillus rhamnosus cells during freeze-drying in the presence or absence of inositol, sorbitol, fructose, trehalose, monosodium glutamate and propyl gallate. However, survival was higher during storage when drying took place in the presence of these compounds. Sorbitol produced more significant effects than the other compounds toward maintaining viability of freeze-dried L. plantarum and L. rhamnosus.

This paper reports experimental and modeling work concerning alcoholysis reactions between butanol and ethyl butanoate, catalyzed by Lipozyme™ in n -hexane, using a batch stirred system at 60°C. Description of the reaction kinetics was based on a postulated multisubstrate Ping Pong Bi Bi mechanism, and appropriate rate expressions were derived for all components in the reaction medium. Simplified models were fitted by non-linear multiresponse regression analysis to data (experimental or calculated from mass balances, as appropriate) encompassing the concentrations of free butanol, ethyl butanoate, ethanol and butyl butanoate. Finally, incremental F -tests were performed to assess the simplest model form that was able to provide a statistically good fit throughout the entire reaction time frame.

Hydrolysis of whey proteins may produce peptide mixtures with better functional properties than the original protein mixture, viz. higher solubilites and lower allergenic effects. Cynara cardunculus is a wild plant that possesses (aspartic) proteases in its flower cells; those enzymes exhibit general proteolytic and specific milk clotting activities, which are rather useful in traditional cheesemaking. This study was thus aimed at characterizing the enzymatic action of crude extracts of said plant after preliminary purification by salting out with ammonium sulfate at two different concentration levels, viz. 30% and 70% saturation. The coagulant activity on milk, and the proteolytic activity using casein and azocasein as substrates, of the crude extract and of each precipitated fraction were measured at 37°C and pH 5.2. The profile of hydrolysis of the major whey proteins, i.e. α-lactalbumin (α-La), β-lactoglobulin (β-Lg) and bovine serum albumin (BSA) was characterized by gel permeation chromatography and polyacrylamide gel electrophoresis (PAGE) in the presence of sodium dodecyl sulfate. The 30% and 70% saturation fractions exhibited lower coagulant and proteolytic specific activities than the crude extract. However, the relative ratio of coagulant to proteolytic activity, which is a useful indicator of appropriateness for cheesemaking, was higher for the partially purified fractions. The extents of hydrolysis of whey proteins brought about by the partially purified extracts were above those by their crude counterpart, but qualitative hydrolysis patterns were essentially identical to each other; by 24 h, α-La was substantially depleted, whereas β-Lg was very poorly hydrolyzed and BSA was only slightly hydrolyzed. The native proteins were converted to lower and lower molecular weight peptides.

Blends of cardosins A and B, enzymes present in aqueous extracts of the flowers of the thistle (Cynara cardunculus L.), have for long been used as rennets by the cheesemaking industry in the Iberian Peninsula. These dimeric proteases are present in the stigmae and stylets of said flowers, and are thought to play a role in sexual reproduction of the plant. In the present research effort, production of cardosin derivatives (starting from a crude extract), encompassing full stabilization of their dimeric structure, has been attempted via covalent, multi-subunit immobilization onto highly activated agarose-glutaraldehyde supports. Boiling such enzyme derivatives in the presence of sodium dodecyl sulfate and beta-mercaptoethanol did not lead to leaching of enzyme, thus proving the effectiveness of the attachment procedure. Furthermore, derivatives prepared under optimal conditions presented ca. half the specific activity of the enzyme in soluble form, and were successfully employed at lab-scale trials to perform (selective) hydrolysis of alpha-lactalbumin, one of the major proteins in bovine whey.

The rates and extents of hydrolysis of αS- and β-caseins from bovine, caprine, and ovine sodium caseinates produced by an enzymatic extract of the fruit of Opuntia ficus-indica, (L.). Miller were evaluated and compared with those produced by a commercial animal rennet. A mechanistic model based on a pseudo-first-order enzymatic reaction, in the presence of first-order deactivation of the enzyme, was postulated and successfully fitted to the experimental data. The animal rennet exhibited higher enzymatic efficiency than the fruit extract, irrespective of the source (i.e., bovine, caprine, or ovine) and the type (i.e., αS- or β-casein) of substrate. The enzymatic efficiency (kcat/Km) for αS-casein ranged from 72 to 220 and from 43 to 65 L g-1 h-1, and for β-casein from 242 to 742 and from 55 to 164 L g-1 h-1, for the animal rennet and the enzymatic extract of O. ficus-indica, respectively. Finally, it was observed that β-casein from caprine and ovine caseinates was degraded by O. ficus-indica faster than its αS counterpart, but the reverse was observed for bovine caseinate.

Direct hydrolysis of whey leads to peptides that possess higher digestibility and better functional properties, so they may to advantage be incorporated in food formulae to improve their performance. Incubation of pure bovine α-La and β-Lg, as well as of caprine, bovine and ovine wheys with trypsin and pepsin led to production of various hydrolysates, which absorb at 280 nm and are characterized by molecular weights ranging from ca. 8000 Da to less than 500 Da. Bovine α-La was slowly hydrolyzed by trypsin but rapidly by pepsin, in either pure form or in whole whey. Bovine β-Lg was more rapidly broken down by trypsin, and less rapidly by pepsin than α-La, and a similar performance was observed when β-Lg was tested on whole whey. In most whey digests, a peak corresponding to a molecular weight comprised between 3000 and 4000 Da was observed by gel permeation chromatography; it was detected mainly in ovine and caprine wheys, and grew slowly with incubation time in bovine whey but fast in wheys from the small ruminants. Incubation of the fraction corresponding to the unknown peak with pepsin did not produce any effect detectable by chromatography, yet incubation with trypsin led to a decrease of the area of such peak and concomitant rise of the areas accounted for by low molecular weight peptides.

The effects of modified atmosphere packaging on growth of adventitious microorganisms in Portuguese whey cheese (Requeijao) were studied following a response surface methodology using storage time (2, 6, 10 and 15 days), storage temperature (4, 12 and 18C) and fraction of CO 2 in the overhead gaseous mixture also containing nitrogen (0, 50 and 100%) as manipulated variables. The viable numbers of Enterobacteriacea, staphylococci, yeasts and spore-forming bacteria in the experimental whey cheeses did not increase within 15 days when storage was at 4C under 100% CO 2 : those of enterococci increased significantly after 6 days under similar conditions, and a similar inhibiting effect was observed against Bacillus, pseudomonads, lactobacilli and streptococci. It was observed that 100% N 2 at 4C was able to completely inhibit growth of staphylococci, lactobacilli and Bacillus for 2 days. The loci (and the nature) of the optima in terms of manipulated variables were obtained for all microbial groups studied. No true overall minimum was found, but storage conditions preset at 4C and 100% CO 2 led to a 15 day extension of the shelf-life of Requeijao.

The concentration of total free amino acids (FAA) in Picante cheese increased with ripening time irrespective of the particular protocol used for manufacture (ie ratio of caprine to ovine milks, animal or plant rennet and number of salting steps). The experimental cheeses manufactured with 20% (v/v) caprine milk, coagulated with animal rennet and salted only once exhibited the highest content of total FAA by 120 days of ripening. All four manufacture parameters were statistically significant on the 0.5% level of significance in terms of total concentration of FAA. The dominating free amino acids present in the various experimental cheeses throughout the ripening period were valine, leucine and phenylalanine, each one representing more than 10% (w/w) of the total concentration of FAA. All four manufacture parameters were, in general, statistically significant with respect to the content of every single FAA, with particular emphasis on salting and ripening time.

The present communication describes the chemical modification of anhydrous butterfat by interes- terification with oleic acid catalyzed by a lipase of Mucor javanicus. Two reactor configurations were tested, a batch-stirred tank reactor containing suspended lipase and a batch-stirred tank reactor in combination with a hollow-fiber membrane module containing adsorbed li- pase. The goal of this research was to assess the advan- tage of using a (hydrophobic) porous support to immo- bilize the lipase in attempts to engineer butterfat with increased levels of unsaturated fatty acid residues (oleic acid) at the expense of medium-to-long chain saturated fatty acids (myristic and palmitic acids). Reactions were carried out at 40°C in the absence of solvent under con- trolled water activity, and were monitored by chromato- graphic assays for free fatty acids. The results obtained indicate that the rate of interesterification using the pro- posed reactor configuration is enhanced by a factor above 100 relative to that using suspended lipase, for the same protein mass basis. Although hydrolysis of butter- fat occurred to some degree, the enzymatic process that uses the hollow-fiber reactor was technically superior to the stirred tank system. © 1998 John Wiley & Sons, Inc. Bio- technol Bioeng 60: 114-123, 1998.

The evolution of concentration of free amino acids in Picante cheese throughout ripening was studied for several volumetric ratios of ewe's and goat's milks. The concentrations of all free amino acids, except asparagine, γ-aminobutyric acid, and cysteine, generally increased as ripening time elapsed. Analyses of variance have indicated that ripening time and, to a lesser extent, milk composition have significant effects on the overall concentration of free amino acids. The major free amino acids present in the various cheeses along the ripening period were valine, leucine, and phenylalanine; these three amino acids accounted for 50, 49, 57, 46 and 42% of total free amino acids at 0 days and 42, 42, 43, 39 and 36% of total free amino acids at 180 days for cheeses manufactured with 0, 25, 50, 75 and 100% goat's milk, respectively. Significant differences could be detected in terms of amino acid profile when the relative proportions of ewe's and goat's milks were altered (eg valine changed from 251.79 ± 0.99 to 352.20 ± 16.49 mg/100 g of dry matter, leucine from 181.48 ± 1.77 to 226.00 ± 11.60 mg/100 g of dry matter, and phenylalanine from 120.39 ± 1.44 to 155.36 ± 8.39 mg/100 g of dry matter in 140-day ripened cheeses when plain ewe's milk was replaced by plain goat's milk). The correlation coefficients between the concentrations of valine, isoleucine, leucine and phenylalanine, on the one hand, and ripening time, on the other, were greater than 0.93.

The modification of anhydrous butterfat via interesterification reactions with oleic acid catalyzed by a lipase from Mucor circinelloides immobilized by adsorption onto hydrophobic hollow fibers is described. A reasonable degree of incorporation of free (externally added) oleic acid into the triacylglycerols of butterfat has been achieved while short-chain fatty acid residues remained virtually unaffected. Total saturated triacylglycerols decreased by 27%, and triacylglycerols with 32–44 acyl carbons (which contained two or three lauric, myristic, or palmitic acid residues) decreased by 33%. Total monoene and polyene triacylglycerols increased by 21% and 17%, respectively. The triacylglycerols (TAG) of interesterified butterfat had approximately 27% more oleic acid residues and approximately 8% less lauric, 6% less myristic, and 6% less palmitic acid residues than those of the original butterfat; the fraction of low-melting TAG peak increased by 19% whereas that of high-melting TAG decreased by 83%. Although a certain degree of butterfat hydrolysis was observed, enzymatic acidolysis was technically feasible and able to produce a modified butterfat with a stronger nutraceutical character.

Lipases have become powerful tools in the manufacture of structured fats either via randomization of their glyceride composition or incorporation of externally supplied fatty acid residues in such glycerides. The present communication reports on changes that occurred in the fatty acid pool of anhydrous butterfat subject to interesterification and to acidolysis with oleic acid catalyzed by a commercial lipase immobilized by plain physical adsorption onto hydrophobic hollow fibers at 40°C under controlled water activity. The main goal of this research effort was to engineer butterfat so as to increase its level of unsaturated fatty acid residues and concomitantly decrease its level of medium- and long-chain saturated fatty acid residues (viz. lauric and myristic acids). Although a certain degree of net hydrolysis of butterfat was observed, the triacylglycerols of butterfat subject to acidolysis were found to possess more (approximately 30% w/w) oleic acid and significantly less (8% w/w) lauric acid and less (2% w/w) myristic acid than those of the original butterfat.

Primary proteolysis of ovine and caprine Na-caseinate at 30°C in phosphate buffer at pH 6.5 or 5.5 in the absence of NaCl and at pH 5.2 with 5% (w/v) NaCl by cardosins in aqueous extracts of Cynara cardunculus flowers was investigated using urea-polyacrylamide gel electrophoresis and reversed-phase high performance liquid chromatography. Caprine caseinate underwent more extensive degradation than ovine caseinate under the same conditions (pH 6.5 and pH 5.5); proteolysis of β- and αs-caseins in ovine and, to a lesser extent, in caprine caseinates was reduced in the presence of 5% (w/v) NaCl. Peptide profiles of the pH 4.6-soluble extract had different patterns throughout ripening arising from the different specificity of cardosins toward ovine and caprine Na-caseinates. The major cleavage sites in ovine (caprine) caseinate were Phe105-Met106 (Lys116-Thr117) for κ-casein, Leu127-Thr128 and Leu190-Tyr191 (Glu100-Thr101, Leu127-Thr128, Leu136-Pro137 and Leu190-Tyr191) for β-casein, Phe23-Val24 (Phe23-Val24, Trp164-Tyr165 and Tyr173-Thr174) for αs1-casein and Phe88-Tyr89 (Ser9-Ser10, Phe88-Tyr89 and Tyr179-Leu180) for αs2-casein.

Decreasing consumption of high fat milk and dairy products is driving the dairy industry to seek other uses for increasing surplus of milkfat. Enzyme catalyzed modification of milkfat using lipases is receiving particular attention. This review examines lipase-mediated modification of milkfat. Especial attention is given to industrial applications of lipases for producing structured and modified milkfat for improved physical properties and digestibility, reduced caloric value, and flavor enhancement. Features associated with reactions such as hydrolysis, transesterification, alcoholysis and acidolysis are presented with emphasis on industrial feasibility, marketability and environmental concerns. Future prospects for enzyme catalyzed modification of milk fat are discussed.

The present communication reports experimental and modelling work pertaining to the independent roles of pH and temperature on deactivation of a crude lipase from Mucor juvanicus. Experimental data of lipolytic activities were generated by a classic pH-stat assay on a triolein emulsion following incubation at several pH values for a fixed time, or at several temperatures for various times; postulated models were then fitted by nonlinear fitting to such data. The pH-dependence data were best fit by assumption of three forms of enzyme with increasing states of protonation, with pKa values of 6.2 and 11.3, respectively, where only the intermediate form is stable within the time frame considered. The thermal-dependence data were best fit by assumption of parallel steps of deactivation and rearrangement, with activation energies of 228.8 and 221.7 kJ mol-1, respectively.

Seasonal changes of the ash content and mineral concentrations in Serra cheese were studied over a typical 35-day ripening period. Statistically significant differences (at the 5% level) exist between the ash content and the concentrations of Na, K, Ca, P, Mg and Zn in cheeses during ripening. The highest concentrations of Na was obtained in cheese ripened for 7 days, whereas the concentrations of K, Ca, P, Ng and Zn decreased significantly during ripening. For 35-day-old cheeses, concentrations of Na, K and Cu were lowest and concentration of P was highest for cheeses manufactured in May. The concentration of Ca was lowest for cheeses manufactured in February. On average, the most concentrated minerals (in g kg -1 of total solids, TS) in 35-day-old Serra cheese were Na (18.56), Ca (9.70) and P (7.92) and, at a lower level, K (1.70) and Mg (0.96). Only trace levels (in mg kg τs -1 ) of Zn (94.33), Cu (2.26) and Mn (1.25) were detected. A high mineral nutrition quality was thus ascribed to 35-day-old Serra cheese based on the average nutritional densities: 4.8 for Ca, 4.0 for P, 1.1 for Mg, 3.4 for Na, 2.4 for Zn, 0.4 for Cu, 0.2 for Mn and 0.2 for K.

A food can be considered as functional if, beyond its nutritional outcomes, it provides benefits upon one or more physiological functions, thus improving health while reducing the risk of illness [1]. This definition – originally proposed by the European Commission Concerted Action on Functional Food Science in Europe (FuFoSE), should be refined in that: (i) the functional effect is different from the nutritional one; and (ii) the benefit provided requires scientific consubstantiation in terms of improvement of physiological functions, or reduction of occurrence of pathological conditions. The concept of functional food emerged in Japan during the 80's, chiefly because of the need to improve the quality of life of a growing elderly population – who typically incurs in much higher health costs [2]. Nowadays, a growing consumer awareness of the relationship between nutrition and health has made the market of functional foods to boom.

Gastrointestinal cancers rank second in overall cancer-related deaths. Carotenoids, sulfated polysaccharides, and polyunsaturated fatty acids (PUFAs) from microalgae exhibit cancer chemopreventive features at different stages of carcinogenesis. For instance, sulfated polysaccharides bear a prophylactic potential via blocking adhesion of pathogens to the gastric surface, whereas carotenoids are effective against Helicobacter pylori infection. This effect is notable because H. pylori has been targeted as the primary cause of gastric cancer. Recent results on antitumor and antibacterial compounds synthesized by microalgae are reviewed here, with an emphasis on their impact upon H. pylori infection and derived pathologies accompanying the progression of gastric carcinogenesis.

Response surface methods were utilized in the statistical optimization of several quality factors pertaining to the preservation of carrot slices using a brine containing lactic acid produced by in situ fermentation. These factors were the concentrations of salt, acid, and reducing sugars, and the texture of the carrot material, and the pH, concentrations of acid, reducing sugars, and biomass (at two different times) of the brine. The processing variables considered were the temperature, the salt concentration of the brine, and the ratio of volumes of brine to the carrot material. The starting (corner and center) factorial design has indicated that, within the experimental range of practical interest, a linear model failed to provide a good fit to the experimental data ; hence, this design was complemented with an axial design. Checks of the residuals and estimates of third-order parameters have indicated that no apparent reason existed to question the statistical adequacy of the quadratic empiric model. The loci of the optima (and the characterization of such optima) were then obtained for this model, and the general directions for the variation of the values of the processing parameters were presented. This study has indicated that temperature may be manipulated to give rise to any desired increase or decrease of the quality factor chosen, a conclusion that may be relevant in attempts to industrially improve carrot preservation processes based on lactic acid fermentation.

Scope: The aim of this study was to investigate the antihypertensive effect of a peptide fraction (PepC) obtained from a whey protein concentrate following hydrolysis by Cynara cardunculus, as well as of its fraction with MW below 3 kDa (PepCF). Both these concentrates encompassed peptides that exhibited potent in vitro inhibition of angiotensin-converting enzyme (ACE): two were released from a-lactalbumin – KGYGGVSLPEW and DKVGINYW, and one from b-lactoglobulin – DAQSAPLRVY. Methods and results: Upon oral administration, by gastric intubation, of 400 mg/kg body weight (bw) of those peptide concentrates, or 5 mg/kg bw of the corresponding synthetic peptides, to 12 wk-old spontaneously hypertensive rats (SHR), the systolic and diastolic blood pressures were monitored by the tail-cuff method – before, and 2, 4, 6, 8 and 24 h afterwards. Water and zofenopril (5 mg/kg bw) – a known ACE-inhibitor, were used as negative and positive controls, respectively. Acute administration of PepC, PepCF, KGYGGVSLPEW, DKVGINYW and DAQSAPLRVY caused antihypertensive effects in SHR; the maximum effect occurred by 4 h and 6 h after administration of the peptide concentrates and the synthetic peptides, respectively. PepC and KGYGGVSLPEW also showed ACE-inhibitory activity in vivo: the pressor effect of angiotensin I was significantly lower, and the response to bradykinin increased when the rats were pre-treated with either product. Conclusion: Our results strongly suggest that PepC will be effective as nutraceutical ingredient for the formulation of functional foods aimed at hypertension control.

Microalgae have much higher lipid yields than those of agricultural oleaginosous crops, and they do not compromise arable land. Despite this, current microalga-based processes suffer from several constraints pertaining to the biocatalyst and the bioreactor, which hamper technologically and economically feasible scale-up. Here, we briefly review recent active research and development efforts worldwide, and discuss the most relevant shortcomings of microalgal biofuels. This review goes one step further relative to related studies, because it tackles otherwise scarcely mentioned issues – for example, heterotrophic versus autotrophic metabolism, alkane versus glyceride synthesis, conduction versus bubbling of CO 2 , and excretion versus accumulation of lipids. Besides promising solutions that have been hypothesized and arise from multidisciplinary approaches, we also consider less conventional ones. Microalgae and biofuels hold indeed a promising partnership, but a fully competitive technology is not expected to be available before the end of this decade, because the need for one order of magnitude increase in productivity requires development of novel apparatuses and transformed cells.

A model based on transient substrate and population balances coupled with Monod kinetics for growth has been formulated using several simplifying assumptions for batch cultures of cells which form biomass aggregates. The functional form of the analytical solution of this system of equations in its most generic aspect has been obtained and applied in the calculation of the time period corresponding to the maximum average rate of biomass increase.

A lipase from Aspergillus niger, immobilized by adsorption on microporous, polypropylene hollow fibers, was used to effect the hydrolysis of the glycerides of melted butterfat at 40°C and pH values of 3.0, 4.0, 5.0, 7.0, 8.0, and 9.0. The aqueous phase and the melted butterfat were pumped in a cocurrent fashion through the lumen and the shell sides, respectively, of a hollow fiber reactor. Nonlinear regression methods were employed to independently fit the data set obtained at each pH to models based on combinations of three nested rate expressions for the hydrolysis and three nested rate expressions for thermal deactivation of the enzyme. A rate expression containing 4 lumped parameters is sufficient to model the release of free fatty acids at each of the six pH values as a function of reactor space time and time elapsed after immobilization. Nonlinear regression methods were also employed in global fits of the data to rate expressions containing an explicit dependence on pH. For the reaction conditions ...

Unbalanced production of atmospheric CO(2) constitutes a major challenge to global sustainability. Technologies have thus been developed for enhanced biological carbon fixation (also referred to as CO(2) mitigation), and one of the most promising capitalizes on microalgae. However, the "best bioreactor", which would be able to achieve maximum productivity and maximum energy efficiency under a given set of operational costs, does not exist. This review briefly examines the current technologies available for enhanced microalgal CO(2) fixation, and specifically explores the possibility of coupling wastewater treatment with microalgal growth for eventual production of biofuels and/or added-value products, with an emphasis on productivity. In addition, an overview of reactor configurations for CO(2) fixation and bottlenecks associated with the underlying technology are provided. Finally, a review of life cycle analysis studies is presented, and routes for improvement of existing processes are suggested.