Stability of clavulanic acid under variable pH, ionic strength and temperature conditions. A new kinetic approach

Clavulanic acid (CA) is a beta-lactam antibiotic that alone exhibits only weak antibacterial activity, but is a potent inhibitor of beta-lactamases enzymes. For this reason it is used as a therapeutic in conjunction with penicillins and cephalosporins. However, it is a well-known fact that it is unstable not only during its production phase, but also during downstream processing. Therefore, the main objective of this study was the evaluation of CA long-term stability under different conditions of pH and temperature, in the presence of variable levels of different salts, so as to suggest the best conditions to perform its simultaneous production and recovery by two-phase polymer/salt liquid-liquid extractive fermentation. To this purpose, the CA stability was investigated at different values of pH (4.0-8.0) and temperature (20-45 degrees C), and the best conditions were met at a pH 6.0-7.2 and 20 degrees C. Its stability was also investigated at 30 degrees C in the presence of NaCl, Na(2)SO(4), CaCl(2) and MgSO(4) at concentrations of 0.1 and 0.5 M in Mcllvaine buffer (pH 6.5). All salts led to increased CA instability with respect to the buffer alone, and this effect decreased in following sequence: Na(2)SO(4) > MgSO(4) > CaCl(2) > NaCl. Kinetic and thermodynamic parameters of CA degradation were calculated adopting a new model that took into consideration the equilibrium between the active and a reversibly inactivated form of CA after long-time degradation. (C) 2009 Elsevier B.V. All rights reserved.

Growth and acidification performance of probiotics in pure culture and co-culture with Streptococcus thermophilus: The effect of inulin

Inulin was used as a prebiotic to improve the quality and consistency of skim milk fermented by co-cultures and pure Cultures of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus bulgaricus and Bifidobacterium lactis with Streptococcus thermophilus. We compared, either in the presence or absence of 4 g inulin/100 g, the results of the main kinetic parameters, specifically the generation time (t(g)), the maximum acidification rate (V(max)). and the times to reach V(max) (t(max)), to attain pH 5.0 (t(pH5.0)) and to complete the fermentation (t(pH4.5)). Post-acidification, lactic acid formation and cell counts were also determined and compared, either 1 day after the fermentation was complete or after 7 day storage at 4 degrees C. In general, inulin addition to the milk increased in co-cultures V(max), decreased t(max), t(g) and t(pH4.5), favored post-acidification, exerted a bifidogenic effect, and preserved almost intact cell viability during storage. In addition, S. thermophilus was shown to stimulate the metabolism of the other lactic bacteria. Contrary to co-cultures, most of the effects in pure Cultures were not statistically significant. The most important aspect of this paper is the use of the generation time as a toot to investigate the microbial response to inulin addition. (c) 2009 Elsevier Ltd. All rights reserved.

Liquid-liquid extraction by mixed micellar systems: A new approach for clavulanic acid recovery from fermented broth

This work is the first attempt to apply aqueous two-phase mixed micellar systems (ATPMS) of the nonionic surfactant Triton X-114 and the anionic one AOT to extract clavulanic acid (CA) from broth fermented by Streptomyces clavuligerus. Cloud points were determined in McIlvane buffer pH 6.5 with or without NaCl, and diagram phases/coexistence curves were constructed. CA partition was investigated following a 2(4)-full factorial design in which AOT (0.022, 0.033 and 0.044% w/w), Triton X-114 (1.0, 3.0 and 5.0% w/w) and NaCl (0, 2.85 and 5.70% w/w) concentrations and temperature (24,26 and 28 degrees C) were selected as independent variables, and CA partition coefficient (K(CA)) and yield in the top phase (eta(CA)) as responses. CA partitioned always to the top, micelle-poor phase. The regression analysis pointed out that NaCl concentration and interaction between temperature and Triton X-114 concentration had statistically significant effects on K(CA), while eta(CA) was mainly influenced by temperature, Triton X-114 concentration and their interaction. Different ATPMS compositions were then needed to maximize these responses, specifically 0.022% (w/w) AOT, 5% (w/w) Triton X-114 for K(CA) (2.08), and 0.044% (w/w) AOT, 1% (w/w) Triton X-114 for eta(CA) (98.7%), both at 24 degrees C without NaCl. Since at 0.022% (w/w) AOT, 1% (w/w) Triton X-114 and 28 degrees C without NaCl the system was able to ensure satisfactory intermediate results (K(CA) = 1.48; eta(CA) = 86.3%), these conditions were selected as the best ones. These preliminary results are of concern for possible industrial application, because CA partition to the dilute phase can simplify the subsequent purification protocol. (C) 2011 Elsevier B.V. All rights reserved.

Thermal Behavior, Microstructure, Polymorphism, and Crystallization Properties of Zero Trans Fats from Soybean Oil and Fully Hydrogenated Soybean Oil

Blends of soybean oil (SO) and fully hydrogenated soybean oil (FHSBO), with 10, 20, 30, 40, and 50% (w/w) FHSBO content were interesterified under the following conditions: 20 min reaction time, 0.4% sodium methoxide catalyst, and 500 rpm stirring speed, at 100 A degrees C. The original and interesterified blends were examined for triacylglycerol composition, thermal behavior, microstructure, crystallization kinetics, and polymorphism. Interesterification produced substantial rearrangement of the triacylglycerol species in all the blends, reduction of trisaturated triacylglycerol content and increase in monounsaturated-disaturated and diunsaturated-monosaturated triacylglycerols. Evaluation of thermal behavior parameters showed linear relations with FHSBO content in the original blends. Blend melting and crystallization thermograms were significantly modified by the randomization. Interesterification caused significant reductions in maximum crystal diameter in all blends, in addition to modifying crystal morphology. Characterization of crystallization kinetics revealed that crystal formation induction period (tau (SFC)) and maximum solid fat content (SFC(max)) were altered according to FHSBO content in the original blends and as a result of the random rearrangement. Changes in Avrami constant (k) and exponent (n) indicated, respectively, that-as compared with the original blends-interesterification decreased crystallization velocities and modified crystallization processes, altering crystalline morphology and nucleation mechanism. X-ray diffraction analyses revealed that interesterification altered crystalline polymorphism. The interesterified blends showed a predominance of the beta` polymorph, which is of more interest for food applications.

The effect of inulin as a prebiotic on the production of probiotic fibre-enriched fermented milk

We investigated the effect of inulin as a prebiotic on the production of probiotic fibre-enriched fermented milk. The kinetics of acidification of inulin-supplemented milk (0, 0.01, 0.02 and 0.04 g/g), as well as probiotic survival, pH and firmness of fermented milk stored at 4 degrees C for 24 h and 7 days after preparation were examined. Probiotic fibre-enriched fermented milk quality was influenced both by the amount of inulin and by the co-culture composition. Depending on the co-culture, inulin addition to milk influenced acidification kinetic parameters, probiotic counts, pH and the firmness of the product.

Zero trans fats from soybean oil and fully hydrogenated soybean oil: Physico-chemical properties and food applications

Blends of soybean oil (50) and fully hydrogenated soybean oil (FHSBO), with 10%, 20%, 30%, 40% and 50% FHSBO (w/w) content were interesterified under the following conditions: 0.4% sodium methoxide, 500 rpm stirring, 100 degrees C, 20 min. The original and interesterified blends were examined for triacylglycerol composition, melting point, solid fat content (SFC) and consistency. Interesterification caused considerable rearrangement of triacylglycerol species, reduction of trisaturated triacylglycerol content and increase in monounsaturated and diunsaturated triacylglycerols, resulting in lowering of respective melting points. The interesterified blends displayed reduced SFC at all temperatures and more linear melting profiles as compared with the original blends. Yield values showed increased plasticity in the blends after the reaction. Isosolid diagrams before and after the reaction showed no eutectic interactions. The 90:10, 80:20, 70:30 and 60:40 interesterified SO:FHSBO blends displayed characteristics suited to application, respectively, as liquid shortening, table margarine, baking/confectionery fat and all-purpose shortenings/biscuit-filing base. (C) 2009 Elsevier Ltd. All rights reserved.

Effect of inulin on growth and acidification performance of different probiotic bacteria in co-cultures and mixed culture with Streptococcus thermophilus

Inulin was used as a prebiotic to improve the quality and consistency of skim milk fermented by Lactobacillus acidophilus (La), Lactobacillus rhamnosus (Lr), Lactobacillus bulgaricus (Lb) and Bifidobacterium lactis (BI) with Streptococcus thermophilus (St), either in binary co-cultures or in cocktail containing all microorganisms. We compared, either in the presence of 40 mg inulin g(-1) or not, the results of the maximum acidification rate (V(max)) and the times to reach it (t(max)), to reach pH 5.0 (t(PH5.0)) and to complete the fermentation (t(f)). Post-acidification, lactic acid formation and cell counts were also compared after either 1 day (D1) or 7 days of storage at 4 degrees C (N). In co-culture, inulin addition to the milk increased V(max), decreased t(max) and t(f), favored post-acidification and exerted a bifidogenic effect. S. thermophilus proved to stimulate the metabolism of the other lactic bacteria and enhanced the product features. After D7, a significant prebiotic effect of inulin was observed in all co-cultures. Either after D1 or D7, the enumerations of Lr and BI in mixed culture markedly decreased compared to their respective co-cultures because of greater competition for the same substrates. (C) 2008 Elsevier Ltd. All rights reserved.

Viability of Lactobacillus acidophilus La-5 added solely or in co-culture with a yoghurt starter culture and implications on physico-chemical and related properties of Minas fresh cheese during storage

The effect of a probiotic culture of Lactobacillus acidophilus (La-5), added solely or in co-culture with a starter culture of Streptococcus thermophilus, on texture, proteolysis and related properties of Minas fresh cheese during storage at 5 degrees C was investigated. Three cheese-making trials were prepared and produced with no addition of cultures (T1 - control), supplemented with La-5 (T2), and with La-5 + S. thermophilus (T3). Viable counts of La-5 remained above 6.00 log cfu g(-1) during the whole storage for T2, reaching 7.00 log cfu g(-1) on the 14th day. For T3, the counts of La-5 remained above 6.00 log cfu g(-1) after 7 days of storage. Due to the presence of S. thermophilus, T3 presented the highest proteolytic index increase and titratable acidity values. Nevertheless, these results and S. thermophilus addition had no influence on viability of La-5 which presented satisfactory populations for a probiotic food. Moreover, the use of a yoghurt culture for the production of Minas fresh cheese T3 supplemented with La-5 resulted in a good quality product, with a small rate of post-acidification, indicating that traditional yoghurt culture could be employed in co-culture with La-5 to improve the quality of this cheese. (C) 2008 Swiss Society of Food Science and Technology. Published by Elsevier Ltd. All rights reserved.

Effect of different prebiotics on the fermentation kinetics, probiotic survival and fatty acids profiles in nonfat symbiotic fermented milk

The simultaneous effects of different binary co-cultures of Lactobacillus acidophilus, Lactobacillus bulgaricus, Lactobacillus rhamnosus and Bifidobacterium lactis with Streptococcus thermophilus and of different prebiotics on the production of fermented milk were investigated in this paper. In particular, we determined and compared the kinetics of acidification of milk either as such or supplemented with 4% (w/w) maltodextrin, oligofructose and polydextrose, as well as the probiotic survival, chemical composition (pH, lactose, lactic acid and protein contents), fatty acids profile and conjugate linoleic acid (CIA) content of fermented milk after storage at 4 degrees C for 24 h. Fermented milk quality was strongly influenced both by the co-culture composition and the selected prebiotic. Depending on the co-culture, prebiotic addition to milk influenced to different extent kinetic acidification parameters. All probiotic counts were stimulated by oligofructose and polydextrose, and among these B. lactis always exhibited the highest counts in all supplemented milk samples. Polydextrose addition led to the highest post-acidification. Although the contents of the main fatty acids were only barely influenced. the highest amounts of conjugated linoleic acid (38% higher than in the control) were found in milk fermented by S. thermophilus-L. acidophilus co-culture and supplemented with maltodextrin. (C) 2008 Elsevier B.V. All rights reserved.

Separation and partitioning of Green Fluorescent Protein from Escherichia coli homogenate in poly(ethylene glycol)/sodium-poly(acrylate) aqueous two-phase systems

The partitioning of Green Fluorescent Protein (GFP) in poly(ethylene glycol)/Na-poly(acrylate) aqueous two-phase systems (PEG/NaPA-ATPS) has been investigated. The aqueous two-phase systems are formed by mixing the polymers with a salt and a protein solution. The protein partitioning in the two-phase system was investigated at 25 degrees C. The concentration of the GFP was measured by fluorimetry. It was found that the partitioning of GFP depends on the salt type, pH and concentration of PEG. The data indicates that GFP partitions more strongly to the PEG phase in presence of Na2SO4 relative to NaCl. Furthermore, the GFP partitions more to the PEG phase at higher pH. The partition to the PEG phase is strongly favoured in systems with larger tie-line lengths (i.e. systems with higher polymer concentrations). The molecular weight of PEG is important since the partition coefficient (K) of GFP gradually decreases with increasing PEG size, from K ca. 300-400 for PEG 400 to K equal to 1.19 for PEG 8000. A separation process was developed where GFP was separated from a homogenate in two extraction steps: the GFP is first partitioned to the PEG phase in a PEG 3000/NaPA 8000 system containing 3 wt% Na2SO4, where the K value of GFP was 8. The GFP is then re-extracted to a salt phase formed by mixing the previous top-phase with a Na2SO4 solution. The K-value of GFP in this back-extraction was 0.22. The total recovery based on the start material was 74%. (c) 2008 Elsevier B.V. All rights reserved.

Nisin expression production from Lactococcus lactis in milk whey medium

BACKGROUND: Nisin is a commercially available bacteriocin produced by Lactococcus lactis ATCC 11454 and used as a natural agent in the biopreservation of food. In the current investigation, milk whey, a byproduct from dairy industries was used as a fermentation substrate for the production of nisin. Lactococcus lactis ATCC 11454 was developed in a rotary shaker (30 degrees C/36 h/100 rpm) using two different media with milk whey (i) without filtration, pH 6.8, adjusted with NaOH 2 mol L-1 and without pH adjustment, both autoclaved at 121 degrees C for 30 min, and (ii) filtrated (1.20 mu m and 0.22 mu m membrane filter). These cultures were transferred five times using 5 mL aliquots of broth culture for every new volume of the respective media. RESULTS: The results showed that culture media composed of milk whey without filtration supplied L. lactis its adaptation needs better than filtrated milk whey. Nisin titers, in milk whey without filtration (pH adjusted), was 11120.13 mg L-1 in the second transfer, and up to 1628-fold higher than the filtrated milk whey, 6.83 mg.L-1 obtained in the first(t) transfer. CONCLUSIONS: Biological processing of milk byproducts (milk whey) can be considered a profitable alternative, generating high-value bioproducts and contributing to decreasing river disposals by dairy industries. (C) 2008 Society of Chemical Industry.

Moisture sorption isotherm characteristics of freeze-dried D-limonene emulsions in modified chitosan and maltodextrin

This article reports on modified chitosan as an alternative substance for protecting loss of volatile compounds during freeze drying. Moisture sorption isotherms of freeze-dried D-limonene emulsions in modified chitosan were determined at 15, 25, and 35 degrees C. The data were adjusted to the GAB model. Maltodextrin was used in a parallel experiment. Flavor released from microcapsules was measured. The monolayer humidity, the sorption heat, the diffusivity coefficients, and the surface area of freeze-dried D-limonene emulsions were determined.

Investigation of charged polymer influence on green fluorescent protein thermal stability

Methods of stabilization and formulation of proteins are important in both biopharmaceutical and biocatalysis industries. Polymers are often used as modifiers of characteristics of biological macromolecules to improve the biochemical activity and stability of proteins or drug bioavailability. Green fluorescent protein (GFP) shows remarkable structural stability and high fluorescence; its stability can be directly related to its fluorescence output, among other characteristics. GFP is stable under increasing temperatures, and its thermal denaturation is highly reproducible. Relative thermal stability was undertaken by incubation of GFP at varying temperatures and GFP fluorescence was used as a reporter for unfolding. At 80 degrees C, DEAE-dextran did not have any effect on GFP fluorescence, indicating that it does not confer stability.

LACTOBACILLUS ACIDOPHILUS AND BIFIDOBACTERIUM SP IN CO-CULTURE IMPROVE SENSORY ACCEPTANCE OF POTENTIALLY PROBIOTIC PETIT-SUISSE CHEESE

Sensory acceptance of four trials of probiotic petit-suisse cheese was investigated. Cheeses were prepared using Streptococcus thermophilus TA 040 as starter not supplemented with any probiotic culture (T1-control), Lactobacillus acidophilus La-5 (T2), Bifidobacterium animalis subsp. lactis BL04 (T3) and L. acidophilus + B. animalis subsp. lactis (T4). Sensory acceptance tests were performed after 7 and 14 days of storage at 4 +/- 1 degrees C, using a 9-point hedonic scale to evaluate flavour, texture and overall acceptability. The population of La-5 and BL04 remained at 7.0 log CFU g(-1) and at 8.0 log CFU g(-1), respectively, during storage for up to 28 days. After 7 and 14 days of storage, cheese T4 presented the highest sensory acceptance for all attributes evaluated and differed significantly from the other cheeses (P<0.05). After 14 days of storage, cheese T3 presented the lowest acceptance and differed significantly from the other cheeses (P<0.05). The supplementation of petit-suisse cheese T4 with both La-5 and BL04 in co-culture with a starter culture resulted in a product with high probiotic populations during storage and excellent sensory acceptance. The results also showed that, when added separately, La-5 and BL04 did not affect the sensory acceptability of petit-suisse cheese.

Effect of natural pigments on the oxidative stability of sausages stored under refrigeration

The objective was to use natural pigments to replace sodium erythorbate (NaEry), a synthetic compound used as an antioxidant in sausage formulations, and to evaluate the oxidative stability of the samples. Six assays were prepared in which sodium erythorbate (ERY) at 0.05 g/100 g was substituted by norbixin (NOR), lycopene (LYC), zeaxanthin (ZEA), beta-carotene (CAR) or dextrose (used as a control (CON)). Physical, chemical, color, texture and sensory parameters were measured on the first day and after 45 days of storage at 4 degrees C. All pigments used in the sausage formulations were able to maintain the oxidative stability of the sausages (MDA equivalents <038 mg/kg). Zeaxanthin and norbixin were the most efficient antioxidants of those tested. This antioxidant effect might be associated with the intermediate polarities of these two compounds, which would allow them to concentrate in the membrane lipids or emulsion interface, where lipid oxidation is most prevalent. Other volatile secondary products of oxidation besides MDA should be evaluated in further studies involving natural pigments and sensory oxidative stability. (C) 2009 Elsevier Ltd. All rights reserved.