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The Effect of Growth Medium pH towards Trypsin-Like Activity Produced by Lactic Acid Bacteria DYAH WULANSARI; BUDIASIH WAHYUNTARI; TRISMILAH TRISMILAH; ASTUTIATI NURHASANAH
Microbiology Indonesia Vol. 6 No. 2 (2012): June 2012
Publisher : Indonesian Society for microbiology

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (911.59 KB) | DOI: 10.5454/mi.6.2.1

Abstract

In cases of pancreatic disease, trypsin deficiency often occurs due to reduced expression of trypsin in the pancreas. Patients with pancreatic problem can be treated with a supplement containing digestive enzymes, including trypsin. However, most of the enzymes currently used for the treatment are derived from porcine and bovine sources. On the other hand, lactic acid bacteria are also known to show trypsin-like activity. In the previous work, our group screened 11 lactic acid bacteria isolates, which had previously been proven to show serine protease activity, for trypsin-like activity. The strains were initially grown in MRS (de Mann, Rogosa and Sharpe) medium before being transferred directly to the production medium to produce trypsin. During the previous study, the initial pH of the production medium was set at 6 (the same as the MRS medium pH), which is the optimum pH for the cell growth of lactic acid bacteria. However, most trypsin has an optimum pH of around 8. In this study, we altered the production medium pH to 8 and we harvested the lactic acid bacteria from MRS medium by centrifugation prior to their inoculation to the production medium. Observation of the culture growth and enzyme activity indicated that the new strategy improved the enzyme activity expressed by some strains.
Optimization of Trypsin-like Protease Production by Lactobacillus plantarum FNCC 0270 using Response Surface Methodology Trismilah,; Nurhasanah, Astutiati; Sumaryono, Wahono; Malik, Amarila; Sadikin, Mohamad
Makara Journal of Science
Publisher : UI Scholars Hub

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Abstract

The purpose of this study was to get optimum medium composition and agitation to trypsin-like protease production by Lactobacillus plantarum FNCC 0270. The medium composition and agitation for enzyme production was optimized using Central Composite Design and Response Surface Method with Design Expert software version 7.1.5 Fermentation was carried out in erlenmeyer flasks at initial pH 8, 37 °C, using an incubator shaker at 87.5 rpm. The best results showed an enzyme activity of 1.0 mU/mL, a protein level of 0.557 mg/mL, and desirability value of 0.740. Numerical optimization was performed to approach the ideal state of the fermentation or the desirability value of 1. The medium composition containing of 3.64% baker's yeast, 1.21% glucose, and 0.13% skim milk was used for the fermentation. The enzyme activity of 1.51 mU/mL and protein level of 0.205 mg/mL can be achieved. After numerical optimization, the fermentation process was verified in erlenmeyer flasks with incubator shaking at 77 rpm, initial pH 8, 37 °C, and 15 h fermentation. The verification results showed that the enzyme activity of 1.273 ± 0.227 mU/mL and protein level of 0.248 ± 0.012 mg/mL.
Medium Optimization for Penicillin Acylase (PAc) Production by Recombinant B. megaterium MS941 Containing pac Gene from B. thuringiensis BGSC BD1 Using Response Surface Methodology FENTRI PARAMITHA PUTRI; ASTUTIATI NURHASANAH; NIKNIK NURHAYATI; IS HELIANTI; KHASWAR SYAMSU
Microbiology Indonesia Vol. 9 No. 2 (2015): June 2015
Publisher : Indonesian Society for microbiology

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (1794.751 KB) | DOI: 10.5454/mi.9.2.3

Abstract

Penicillin G acylase (PAc) hydrolyses of the amide bond of benzylpenicillin (Pen-G) releasing PAA and 6-APA, key intermediate in the production of various semisynthetic penicillins. In this study, we optimised the production medium of PAc by RSM using two variables (xylose as inducer and CaCl2 as divalent cations) to obtain the optimum PAc specific activity from Bacillus megaterium btpacBD1. For this purpose, combinations of five different xylose concentrations (0.13 – 0.87 %) and five different CaCl2 concentrations (0.64 – 4.36 mM) were analysed, in a total of 22 experiments. CCD used for the analysis showed that in shake flask cultivations, xylose and CaCl2 showed significant effects on PAc volumetric activity and the quadratic model was in good agreement with the experimental results (R2= 0.86 (p-value < 0.0001)). The maximum specific activity (130.669 ± 50.241 units mg protein-1) was reached when xylose and CaCl2 concentrations were 0.49% and 2.4 mM, respectively, and medium pH was around 7. Under such conditions, the activity of PAc and protein concentration achieved were 1.318 ± 0.406 units mL-1 and  0.0101 ± 0.01 mg mL-1. The shake flask validation experiments demonstrated that with such medium composition the volumetric activity, protein concentration and specific activity achieved were 1.294 ± 0.171 units mL-1, 0.0102 ± 0.0003 mg mL-1 and 125.91 ± 13.309 units mg-1, respectively. When the optimum medium composition was applied in 10 L bioreactor, the optimum volumetric activity (2.0687 ± 0.0820 units mL-1) and protein concentration (0.0078 ± 0.0008 mg mL-1) were achieved 48 h after the start of the cultivation. However, the optimum PAc specific acivity (1260.52  ± 27.5711 units mg protein-1) was achieved 18 h after the start of the cultivation.