Abstract
For the purpose of studies on the citric acid production, some experiments were carried out with isolated strains. The results obtained were as follows. 1) The optimal culture media of the strain M-80 in surface culture contained 140g of sucrose, 3.0g of (N $H_4$)$_2$S $O_4$, 1.5g of K $H_2$P $O_4$, 0.2g of MgS $O_4$.7$H_2O$, 3.0mg of F $e^{++}$, 1.0mg of Z $n^{++}$, 0.5N HCI to a pH of 5.0 and distilled water to 1.0 liter; and that of the strain M-315 in surface culture contained 140g of sucrose, 2.0g of N $H_4$N $O_3$, 1.0g of K $H_2$P $O_4$, 0.25g of MgS $O_4$. 7$H_2O$, 2.0mg of F $e^{++}$, 2.0mg of Z $n^{++}$, 0.05mg of C $u^{++}$, 0.5N HCI to a pH of 4.5 and distilled water to 1.0 liter. While that of the strain M-315 in submerged culture contained 140g of sucrose, 2.5g of N $H_4$N $O_3$, 1.5g of K $H_2$P $O_4$, 0.3g of MgS $O_4$. 7$H_2O$, 3.0mg of F $e^{++}$, 0.1mg of C $u^{++}$, 0.5N HCI to a pH of 4.5 and distilled water to 1.0 liter. The optimal temperature and size of inoculum were mostly 28-3$0^{\circ}C$, 10$^{7}$ -10$^{8}$ spores/50ml, respectively. 2) Through the course of citric acid production, the growth of strains had nearly been completed, pH value was rapidly decreased below 2.0 and the content of sugar was also reduced, while the accumulation of citric acid in media was remarkably begun in about 3-4 days. The yields of citric acid generally reached the maximum level in 8-10 days in surface or submerged fermentation process. 3) Methanol was effective citric acid production when they were added to fermentation media. In the case of surface culture, by addition of 2％ (strain M-80), 3％ (strain M-315), the yields of citric acid was increased 6.5％, 20.6%, respectively and 5.0% yield was increased by addition of 3% methanol in submerged culture media of the strain M-315. 4) Chromatography analysis of culture broth after fermentation under optimal culture conditions detected that the majority of acid in media was citric acid. 72.1mg/ml, 98.1mg/ml, of citric acid were determined in surface culture media by strains of M-80, M-315, and 59.8 mg/ml of citric acid was contained in the submerged culture media by the strain M-315. strain M-315.
전보에서 분리선정된 균의 발효조건 및 각종첨가물의 영향을 검토하고 배양물을 분석한 결과는 다음과 같다. 1. 선정균의 최적배양 조건은 표면배양의 경우, M-80은 sucrose 140g, (N $H_4$)$_2$S $O_4$3.0g, K $H_2$P $O_4$1.5g, MgS $O_4$.7$H_2O$ 0.25g, F $e^{++}$ 3.0mg, $Zn^{++}$ l.0mg, D. W. 1ι, pH 5.0 이었고, M-315는 sucrose 140g, N $H_4$N $O_3$2.0g, K $H_2$P $O_4$1.0g, MgS $O_4$. 7$H_2O$ 0.25g, F $e^{++}$ 2.0mg, $Zn^{++}$ 2.0mg, C $u^{++}$ 0.05mg, D. W. 1ι pH 4.5 이였으며, M-315의 액내배양의 경우 sucrose 140g, N $H_4$N $O_3$2.5g, K $H_2$P $O_4$1.5g, MgS $O_4$.7$H_2O$ 0.3g, F $e^{++}$ 3.0mg, $Zn^{++}$ 3.0mg, C $u^{++}$ 0.1mg, D. W. 1ι, pH 4.5이었다. 배양온도는 28~3$0^{\circ}C$, 포자접종량은 배양액에 $10^{7}$ -$10^{8}$ 개/ml 접종하였을 때 산생성이 가장 좋았다. 2. 선정균의 생육상태와 산생성관계를 경시적으로 조사한 결과 배양 3~4일 후에 균체의 증식은 거의 끝나고 pH는 2.0이하로 저하되었으며 구연산은 이때부터 급속히 축적되기 시작하여 표면배양의 경우, M-80은 8일, M-315는 9일, M-315의 액내배양의 경우는 10일간 배양하였을 때 구연산생성량이 최고치에 달하였다. 3. Methanol의 첨가는 구연산생성에 좋은 영향을 미쳤으며 표면배양의 경우 M-80, M-315의 배양액에 2.0%, 3.0% 첨가하였을 때 카곽 6.5%, 20%의 수율이 향상되었고, M-315의 액내배양액에는 3.0%의 첨가로 5.0% 의 수율이 향상되었다. 4. 최적조건하에서 발효시킨 분리균의 배양물을 분석한 결과 대부분이 구연산만을 선택적으로 축적하고 있었고, 표면배양의 경우 M-80, M-315 배양액에 각각 72.1mg/ml, 98.1mg/ml이, M-315의 액내배양액에는 59.8mg/ml 구연산이 정량되었다.
