| Abstract: | 本研究主要目的是從台灣的酸菜產品中篩選出具有高γ-胺基丁酸(γ-aminobutyric acid；GABA)生產能力的菌種，並研究此菌種的特性及產GABA的最適條件。首先選出GABA產量最高之酸菜發酵液，其產量為1,236 mg/L，接著從此發酵液中分離出14種菌株並選出其中GABA產量最高的單一菌株SF-1，產量為882.5 mg/L，此株菌在生化測試顯示為革蘭氏陽性桿菌、不具有觸?，具有凝乳效果以及在選擇性培養基中具有透明環，上述結果指出此株菌疑似為乳酸菌。其次以反應曲面法探討培養溫度及起始pH值之最佳組合；結果最適培養條件為培養溫度31.8 ℃及起始pH 6.6，預測之GABA產量為10,659 mg/L，實際產量為10,481 mg/L。接著探討麩胺酸添加量、5'-磷酸?哆醛 (Pyridoxal 5’- phosphate；PLP)以及菌數接種量對於GABA產量之影響；在麩胺酸添加量方面，最佳濃度為400 mM，菌數接種量則是以108 CFU為最佳，而添加PLP不影響GABA產量。在上述最佳條件下培養SF-1四天之GABA產量為17,742 mg/L。為了得知此菌株是否有產出更多GABA的可能性，因此進一步延長培養時間及增加麩胺酸添加量；結果以添加500 mM麩胺酸及培養六天之產量最高，可達43,286 mg/L。最後以API 50 CHL套組及16S rDNA基因定序進行菌種鑑定，確認SF-1為Lactobacillus brevis，至於是否為新的亞種，未來有待進一步進行基因比對。
The aims of this study were to screen high γ-aminobutyric acid-producing bacteria from Mstard Pickle in Taiwan and to investigate its characteristic and opitimal conditions of GABA production. At first, one out of ten fermentation broth from Mustard Pickle was selected due to its highest GABA yield of 1,236 mg/L. Then the single colony named SF-1, which was isolated from 14 colonies in the fermentation broth, had the highest GABA yield of 882.5 mg/L. Base on biochemical test, SF-1 was Gram positive rod, catalase test negative, milk-clotting activity positive and clear zone developing on selective media. The above results showed that SF-1 was similar to lactic acid bacteria. Secondly, the opitimal incubation temperature and initial pH for SF-1 were investigated by Response surface methodology. The result indicated that opitimal conditions were incubation temperature 31.8 ℃ and initial pH 6.6. The prediction and observation GABA yields were 10,659 mg/L and 10,481 mg/L, respectively. Furthermore, the effect of glutamic acid concentration, pyridoxal 5’- phosphate (PLP) and inoculum size on GABA production was also investigated. The opitimal glutamic acid concentration and inoculum size was 400 mM and 108 CFU, respectively, PLP was no effect on GABA yield. According to the above opitimal conditions, SF-1 had the GABA yield of 17,742 mg/L. To enhance the GABA yield, increase of incubation time and glutamic acid concentration was conducted. The results showed the maximum GABA yield of 43,286 mg/L was achieved by increasing incubation time to 6 days and glutamic acid concentration to 500 mM. Finally, identification of the strain was completed by API 50 CHL Kit and 16S rDNA analysis. The result showed SF-1 should be Lactobacillus brevis. However, further gene comparison is required to know whether it is a new subspecies or not in the future. |
