Raja Mahanama

Efficient media for high menaquinone-7 production: response surface methodology approach

The aim of this study was to assess the effect of nutrients on the production of menaquinone-7 and determine the optimum conditions to achieve a high concentration of this product. Bacillus subtilis natto was fermented at 40°C for a period of six days. Design of experiments was used for screening the most effective nutrients, and central composite face design was employed for the optimization. The optimum media consisted of 5% (w/v) yeast extract; 18.9% (w/v) soy peptone; 5% (w/v) glycerol and 0.06% (w/v) K(2)HPO(4). The pH, bacterial growth, concentrations of amino acids, glycerol and menaquinone-7 were measured at the optimum fermentation media each day. Total free amino acids concentration increased 1.7-fold during the fermentation. Lysine and glutamic acid were the most abundant whereas arginine, asparagine and serine were the limiting amino acids at the end of fermentation period. The menaquinone-7 concentration approached 86% of the final value in the third day of fermentation, where the bacteria growth was at exponential phase. At this condition the concentration of glycerol as carbon source and asparagine, serine and arginine as the amino acid sources were dramatically diminished in the fermentation media. The optimum menaquinone-7 concentration was in good agreement with the predicted value by the model (96% validity). The maximum menaquinone-7 concentration of 62.32 ± 0.34 mg/L was achieved after six days of fermentation; this value is the highest concentration reported in the literature.

Vitamin K series: current status and future prospects

Abstract The Vitamin K series, particularly menaquinone, have been attracting research attention, due to the potential in reducing both osteoporosis and cardiovascular diseases. This review provides an overview of the types of vitamin K and their health benefits. This is followed by a critical review of the various biotechnological approaches used in the production of menaquinone, including solid and liquid state fermentations, extraction and recovery. The currently available market information is summarized and future growth prospects are discussed. Recommendations are also given for areas of future research in order to improve the production process for menaquinone and reduce costs.

Enhanced Production of Menaquinone 7 via Solid Substrate Fermentation from Bacillus subtilis

Natto, the richest known source of menaquinone 7 (MK7), is traditionally produced via Solid Substrate Fermentation (SSF) by Bacillus subtilis natto on cooked soy beans. In this work we report a threefold increase in MK7 concentration through the use of a mixture of soy protein granules and nixtamalized corn grits. The effects of fermentation processing factors were investigated and optimized in laboratory scale. These factors include initial moisture content, incubation temperature, incubation time, α-amylase pretreatment and solid substrate medium. Response surface methodology (RSM) was used to develop a mathematical model to identify the optimum values of key process variables to increase MK7 concentration; the model was also validated experimentally. The polynomial model fitted the experimental data well with R2 = 0.89 and R2 (adj) = 0.83. MK7 production was increased from 57.78 ± 1.57 mg/kg to 67.01 ± 0.18 mg/kg when the strain was cultivated at optimum conditions predicted by statistical approach (70% initial moisture, 35°C, 4 days, 10 µL/g amylase and equal substrate mix) as compared to basal conditions (60% initial moisture, 40°C, 6 days, 10 µL/g amylase and equal substrate mix).

Nattokinase production: Medium components and feeding strategy studies

In the present study, the effect of nutrients on nattokinase activity during the fermentation of Bacillus subtilis natto was investigated. The highest nattokinase activity of 587 U/mL was obtained in fermenter for a media consisting of yeast extract (6%, w/v), soy peptone (1.2%, w/v) and glycerol (6%, w/v). The second order polynomial equation was fitted to the results by using central composite face design. The polynomial model fitted the experimental data well with R2 = 0.939 and R2(Adj) = 0.861. In addition, contribution of fed-batch glycerol addition on the nattokinase production pathway was further investigated. Maximum nattokinase activity was resulted when 3% (w/v) glycerol was added to the fermentation media during the cell growth phase. Results demonstrate 25 fold increase of nattokinase activity compared to the batch and not optimized culture. This study provides valuable data on the key nutrients and feeding strategy for further investigations and scale up of nattokinase production process.