Alkaline pretreatment optimization for Pinus roxburghii needle biomass employing response surface methodology for bioethanol production by separate hydrolysis and cofermentation

Production of bioethanol from lignocellulosics can cater to the supply of renewable fuel to transport sector besides being environment friendly. In the present work, bioethanol production potential of Pinus roxburghii needle biomass (PNB) has been studied by optimizing the thermochemical pretreatment method using alkaline conditions (NaOH) firstly by one variable at a time (OVAT) approach, followed by Response Surface Methodology (RSM) with central composite design (CCD) tool. Total reducing sugar (TRS) yield was enhanced from 22.4 g/L (OVAT) to 32.4 g/L using design of experiment (DoE) approach. Effect of pretreatment on PNB was studied by FTIR, phloroglucinol staining and estimation of phenolics which indicated lignin removal. Enzymatic hydrolysis was done by the action of commercial enzymes cellulase and pectinase with loading of 5U/g biomass. The TRS yield was further enhanced to 67.95 g/L after enzymatic hydrolysis. Using separate hydrolysis and cofermentation (SHCF) approach for fermentation of PNB hydrolysate, 28.75 g/L bioethanol was obtained when combined cultures of Saccharomyces cerevisiae (MTCC-36) and Pichia stipitis (NCIM-3498) were used.

Production of phytase from Lactobacillus paracasei strain and its probiotic profile

Phytase, as an effective enzyme for phytic acid degradation, has significance in bioremediation, poultry and aquaculture. In view of such an environmental and industrial importance, phytase producing probiotic bacteria have gained attention. Here, we screened lactic acid bacteria (LAB) for their phytase producing potential. The strain showing maximum phytase activity was studied for its probiotic characteristics. Among 20 LAB isolated from Kalarei, an edible source, two isolates K.J (a) and K exhibiting maximum enzyme activity (5.18 U/mL and 5.0 U/mL) were selected. In optimization studies for production of phytase using ‘one-variable-at-a-time’ (OVAT) approach, isolate K showed maximum (5.92 U/mL) enzyme activity. The strain was identified by ribotyping as Lactobacillus paracasei and the sequence was submitted in NCBI GenBank under accession number MK280749. Further optimization studies for phytase production were carried out using Plackett–Burman design and central composite design (CCD) of response surface methodology (RSM). Statistically optimized four significant variables: incubation temperature, initial pH, maltose and peptone resulted in an increase (6.37 U/mL) in enzyme activity. The enzyme was purified 3.97 fold with a specific activity of 278 U/mg.The molecular weight of enzyme was 70 kDa and optimum activity was determined at 37°C, pH 5.5. The strain was designated as Lactobacillus paracasei SMVDUDB1 and showed promising probiotic characteristics viz. survival rate above 80% (low pH, high bile salt concentration under simulated gastrointestinal conditions), hydrophobicity with chloroform (96.74%), autoaggregation (69.61%) and coaggregation ability (82.79%) with Bacillus subtilis.