Production of nisin from Lactococcus lactis in acid-whey with nutrient supplementation.

The production of Nisin, an FDA-approved food preservative, was attempted by Lactococcus lactis subsp. lactis ATCC® 11454 using the underutilized milk industry effluent, acid-whey, as a substrate. Nisin production was further improved by studying the effect of supplementation of nutrients and non-nutritional parameters. The addition of yeast extract (6% w/v) as nitrogen source and sucrose (4% w/v) as carbon source were found to be suitable nutrients for the maximum nisin production. The changes in the medium pH due to lactic acid accumulation during batch fermentation and its influence on the production of nisin were analyzed in the optimized whey medium (OWM). The production characteristics in OWM were further compared with the nisin production in MRS media. The influence of nisin as an inducer for its own production was also studied and found that the addition of nisin at 0.22 mg/ml promote the nisin production. The analysis of consumption of various metal ions present in the OWM during the nisin production was also analyzed, and found that the copper ions are the most consumed ion. The highest nisin yield of 2.6 × 105 AU/mL was obtained with OWM.

Probing the synergistic effects of rutin and rutin ester on the oxidative stability of sardine oil.

Multicomponent antioxidant mixture is proved to be highly effective in imparting oxidative stability to the edible oil. It is believed that the high efficacy of those mixtures is due to the synergistic effect exhibited by two or more components. The current study aims to analyse the synergistic effect of a flavonoid and its corresponding ester in improving the oxidative stability of n-3 PUFA rich sardine oil. The oxidative stability of rutin, esterified rutin and their combinations at three different concentrations was studied in sardine oil stored at 37 ºC for 12 days in contact with air under darkness. The combination of rutin and rutin ester showed maximum reduction of 54.2% in oxidation at 100 mg/kg and 150 mg/kg. Perhaps this is the first report on the synergistic effect of a flavonoid and its lipophilized ester for improving the oxidative stability of n-3 PUFA rich oil.

Selective extraction of lactoferrin from acidic whey using CTAB/n-heptanol reverse micellar system.

A reverse micellar system comprising CTAB/n-heptanol, developed for extracting lactoferrin (LF) from a synthetic solution of LF, was investigated for the selective extraction of LF from synthetic whey protein solution, which was prepared by mixing the pure whey proteins. The process conditions obtained during the process was further extended to extract the LF from real acidic whey. The selective extraction of LF was improved by studying the effect of NaCl concentration (additive) and aqueous phase pH on the partitioning of LF into the micellar phase. The highest extraction of LF (98.7%) from acidic whey to micellar phase was achieved at the aqueous phase pH of 10.3 and NaCl concentration of 1.1 M. The LF was back extracted to the aqueous stripping phase with 94% extraction efficiency and 100% purity. The recycling capacity of the organic phase after the back extraction of LF was analyzed to make the process more economical.

Aqueous two phase based selective extraction of mannose/glucose specific lectin from Indian cultivar of Pisum sativum seed.

Pisum sativum lectin (Psl) being a high-value protein has marked its application in the biomedical and therapeutic field. Aqueous two phase extraction (ATPE) was implemented as a selective partitioning technique for the partial purification of Psl from its seeds. PEG/citrate based biodegradable aqueous two phase system (ATPS) was screened and the factors such as the type and concentration of citrate salts, molar mass and concentration of polyethylene glycol (PEG), tie line length (TLL) and additive (NaCl) concentration, pH, crude load and volume ratio were studied for the selective partition of Psl. The Psl was successfully extracted to the top phase in the ATPS formed with 18% PEG 6000/16% sodium citrate at 41.01% TLL, 2% NaCl and pH of 7.5. A volume ratio of 0.76 and a crude load of 20% showed maximum activity yield of 122.12% with the purification factor of 16.26. The subunits of Psl namely α and ß were identified with a molecular weight of 6 and 18 kDa respectively during the purity analysis using SDS PAGE and HPLC.

Aqueous two phase partitioning of Pisum sativum lectin in PEG/citrate salt system.

Pisum sativum lectin (Psl) is a metalloprotein which is in the center of research interest because of its HIV-1 reverse transcriptase inhibitory activity and mitogenic activity. The application of this lectin in various fields demands the economically feasible and scalable purification strategy other than affinity chromatography. The suitability of aqueous two phase system (ATPS) composed of poly ethylene glycol (PEG) with different salts (sodium citrate, potassium citrate, and ammonium citrate) was evaluated for better partitioning of Psl. The significant factors such as molar mass and concentration of PEG, type and concentration of salts, the effect of tie line length (TLL), ionic strength, and pH were studied to select a suitable system for better partitioning of Psl. ATPS comprising of 18% PEG 6000, 16% sodium citrate, 1% NaCl at the operating condition of pH 8, 40.23% of TLL, and the volume ratio of 1.32 was found to be the best system which gave a maximum partition coefficient and yield of 14.5% and 98.66%, respectively.

Reverse micellar extraction of lactoferrin from its synthetic solution using CTAB/n-heptanol system.

The partitioning of Lactoferrin (LF) into the reverse micellar phase formed by a cationic surfactant, cetyltrimethylammonium bromide (CTAB) in n-heptanol from the synthetic solution of LF was studied. The solubilization behaviour of LF into the reverse micellar phase and back extraction using a fresh stripping phase were improved by studying the effect of processing parameters, including surfactant concentration, solution pH, electrolyte salt concentration and addition of alcohol as co-solvent. Forward extraction of 100% was achieved at CTAB concentration of 50 mM in n-heptanol solvent, pH of 10 and 1 M NaCl. The electrostatic force and hydrophobic interaction have major influence on LF extraction during forward and back extraction respectively. The size of the reverse micelles and their corresponding water content were measured at different operating conditions to assess their role on the LF extraction. The present reverse micellar system has potential to solubilise almost all the LF into the reverse micelles during forward extraction and could able to allow back extraction from the reverse micellar phase with addition of small amount of co-solvent.

Low frequency sonic waves assisted cloud point extraction of polyhydroxyalkanoate from Cupriavidus necator.

Low frequency sonic waves, less than 10kHz were introduced to assist cloud point extraction of polyhydroxyalkanoate from Cupriavidus necator present within the crude broth. Process parameters including surfactant system variables and sonication parameters were studied for their effect on extraction efficiency. Introduction of low frequency sonic waves assists in the dissolution of microbial cell wall by the surfactant micelles and release of cellular content, polyhydroxyalkanoate granules released were encapsulated by the micelle core which was confirmed by crotonic acid assay. In addition, sonic waves resulted in the separation of homogeneous surfactant and broth mixture into two distinct phases, top aqueous phase and polyhydroxyalkanoate enriched bottom surfactant rich phase. Mixed surfactant systems showed higher extraction efficiency compared to that of individual Triton X-100 concentrations, owing to increase in the hydrophobicity of the micellar core and its interaction with polyhydroxyalkanoate. Addition of salts to the mixed surfactant system induces screening of charged surfactant head groups and reduces inter-micellar repulsion, presence of ammonium ions lead to electrostatic repulsion and weaker cation sodium enhances the formation of micellar network. Addition of polyethylene glycol 8000 resulted in increasing interaction with the surfactant tails of the micelle core there by reducing the purity of polyhydroxyalkanoate.

Anionic surfactant based reverse micellar extraction of L-asparaginase synthesized by Azotobacter vinelandii.

L-Asparaginase synthesized by Azotobacter vinelandii via submerged fermentation in the presence of sucrose was successfully extracted using Reverse micellar extraction. Single step enzyme purification process was developed by varying the process variables which resulted in maximum specificity and extraction of L-asparaginase. The effect of different variables, including broth pH, addition of alcohol during the forward extraction and pH of the fresh stripping aqueous phase, addition of alcohol and electrolyte during backward extraction process were studied. Lower concentration of butanol resulted in maximum activity of the enzyme during forward extraction while enzyme activity was found to increase further with the addition of higher concentrations of ammonium sulphate during backward extraction. Chromatographic analysis of L-asparaginase peak at ~7.65 min was intense for the back extracted sample confirming the maximum purity of L-asparaginase obtained. Purity of L-asparaginase was increased to about 379.68 fold.

Aqueous two phase partitioning of fish proteins: partitioning studies and ATPS evaluation.

A suitable Aqueous Two Phase System (ATPS) has been identified for the partitioning of crude fish proteins from fish processing industrial effluent. A detailed study has been performed to analyze the influence of various parameters on protein partitioning such as molecular weight of PEG, effect of different salts (MgSO4, K2HPO4, Na3C6H5O7, Na2SO4, (NH4) 2PO4, K3C6H5O7) and their concentrations, pH, temperature, Tie Line Length (TLL), effluent loading and volume ratio. PEG 2000 - sodium sulphate ATPS was found to be a most favourable system among the selected ATPS for higher partition coefficient of proteins. The binodal curve and equilibrium characteristics of PEG 2000 - sodium sulphate were established and fitted to empirical equations. The equilibrium compositions (tie line) were correlated using Othmer-Tobias and Bancroft equations.