Identification of a High-yielding Vitamin B 13Acinetobacter Strain and Optimization of Its Fermentation Conditions

doi:10.13560/j.cnki.biotech.bull.1985.2025-0738

Abstract

Abstract:

Objective To isolate and screen a strain of Acinetobacter sp. WJ01 capable of naturally synthesizing vitamin B₁₃ from the microbial resources of the coastal waters in Tianjin Binhai New Area, and to establish its optimal fermentation conditions. Method Strain WJ01 was subjected to screening, identification, and analysis of physiological and biochemical characteristics. The acid-producing substances were analyzed by high-performance liquid chromatography (HPLC), ultraviolet (UV) scanning spectroscopy, infrared (IR) spectroscopy, and Raman spectroscopy. Single-factor experiments and orthogonal tests were performed to optimize the seed culture medium and fermentation medium. Finally, a scale-up experiment was conducted in a 5-L fermenter. Result Acinetobacter sp. WJ01 was identified as a Gram-negative bacterium highly sensitive to the antibiotics ciprofloxacin (CIP), levofloxacin (LEV), minocycline (MI), and doxycycline (DO). Multiple spectroscopic analyses combined with the molecular formula C₅H₄N₂O₄ confirmed that strain WJ01 is an Acinetobacter strain capable of producing VB₁₃. The optimal seed culture parameters were: pH 7.0, temperature 37 ℃, shaking speed 180 r/min, and seed growth time 24 h. The optimal fermentation parameters were: CaCO₃ 6%, inoculum size 2%, temperature 35 ℃, and shaking speed 200 r/min. Using a 5-L fermenter, the VB₁₃ yield reached 112.46 g/L at 80 h, with a fermentation intensity of 1.49 g/L/h and a sugar-acid conversion rate of 0.76 g/g. Conclusion Acinetobacter sp. WJ01 provides an excellent microbial strain for the production of VB₁₃ via fermentation.

Key words: Vitamin B₁₃, HPLC, culture medium optimization, culture condition optimization, fermenter cultivation

ZHANG Yan-hua, QU Wen-long, DAI Wen-jing, ZHANG Rui-ning, LIU Yu-hong, WANG De-pei, XUE Xian-li. Identification of a High-yielding Vitamin B ₁₃Acinetobacter Strain and Optimization of Its Fermentation Conditions[J]. Biotechnology Bulletin, 2025, 41(11): 153-165.

Figures/Tables 15

References 36

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水平 Level	KH₂PO₄ （mg/L）	Na₂SO₄ （mg/L）	CaCl₂ （mg/L）	MgSO₄ （mg/L）
1	80	80	80	80
2	100	100	100	100
3	120	120	120	120

水平 Level	KH₂PO₄ （mg/L）	Na₂SO₄ （mg/L）	CaCl₂ （mg/L）	MgSO₄ （mg/L）
1	80	80	80	80
2	100	100	100	100
3	120	120	120	120

编号 Number	抗生素种类 Types of antibiotics	WJ01 （mm）	编号 Number	抗生素种类 Type of antibiotics	WJ01 （mm）
1	左氧氟沙星（LEV）	30（+++）	16	四环素（TET）	18（++）
2	环丙沙星（CIP）	27（+++）	17	万古霉素（VAN）	X
3	头孢氨苄（CN）	X	18	米诺环素（MI）	21（+++）
4	诺氟沙星（NOR）	20（++）	19	氨苄西林（AMP）	X
5	头孢呋辛钠（CXM）	9（+）	20	哌拉西林（PIP）	15（++）
6	头孢曲松（CTR）	15（++）	21	氟苯尼考（FFC）	X
7	头孢唑林（CZ）	X	22	多黏菌素（PB）	20（++）
8	头孢他啶（CAZ）	18（++）	23	林可霉素（MY）	X
9	青霉素（PEN）	X	24	复方新诺明（SXT）	12（++）
10	头孢哌酮（CPZ）	10（+）	25	庆大霉素（GEN）	X
11	苯唑西林（OX）	X	26	卡那霉素（KAN）	X
12	亚胺培南（IPM）	X	27	克林霉素（CC）	X
13	氯霉素（C）	X	28	多西环素（DO）	26（+++）
14	链霉素（S）	10（+）	29	阿米卡星（AMK）	X
15	红霉素（E）	X	30	阿奇霉素（AZI）	X

编号 Number	抗生素种类 Types of antibiotics	WJ01 （mm）	编号 Number	抗生素种类 Type of antibiotics	WJ01 （mm）
1	左氧氟沙星（LEV）	30（+++）	16	四环素（TET）	18（++）
2	环丙沙星（CIP）	27（+++）	17	万古霉素（VAN）	X
3	头孢氨苄（CN）	X	18	米诺环素（MI）	21（+++）
4	诺氟沙星（NOR）	20（++）	19	氨苄西林（AMP）	X
5	头孢呋辛钠（CXM）	9（+）	20	哌拉西林（PIP）	15（++）
6	头孢曲松（CTR）	15（++）	21	氟苯尼考（FFC）	X
7	头孢唑林（CZ）	X	22	多黏菌素（PB）	20（++）
8	头孢他啶（CAZ）	18（++）	23	林可霉素（MY）	X
9	青霉素（PEN）	X	24	复方新诺明（SXT）	12（++）
10	头孢哌酮（CPZ）	10（+）	25	庆大霉素（GEN）	X
11	苯唑西林（OX）	X	26	卡那霉素（KAN）	X
12	亚胺培南（IPM）	X	27	克林霉素（CC）	X
13	氯霉素（C）	X	28	多西环素（DO）	26（+++）
14	链霉素（S）	10（+）	29	阿米卡星（AMK）	X
15	红霉素（E）	X	30	阿奇霉素（AZI）	X

实验组 Experimental group	A	B	C	实验结果 Experimental results
实验1	1	1	1	0.434
实验2	1	2	2	0.444
实验3	1	3	3	0.654
实验4	2	1	2	0.419 5
实验5	2	2	3	0.336 5
实验6	2	3	1	0.614 5
实验7	3	1	3	0.297
实验8	3	2	1	0.615
实验9	3	3	2	0.614
均值1	0.511	0.383	0.554
均值2	0.457	0.465	0.493
均值3	0.509	0.627	0.429
极差	0.054	0.244	0.125

Identification of a High-yielding Vitamin B ₁₃Acinetobacter Strain and Optimization of Its Fermentation Conditions

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实验组 Experimental group	A	B	C	D	实验结果 Experimental results
实验1	1	1	1	1	32.91
实验2	1	2	2	2	13.61
实验3	1	3	3	3	36.73
实验4	2	1	2	3	55.98
实验5	2	2	3	1	50.25
实验6	2	3	1	2	52.26
实验7	3	1	3	2	73.83
实验8	3	2	1	3	72.11
实验9	3	3	2	1	68.47
均值1	27.75	54.24	52.427	50.543
均值2	52.83	45.323	46.02	46.567
均值3	71.47	52.487	53.603	54.94
极差	43.72	8.917	7.583	8.373

实验组 Experimental group	A	B	C	D	实验结果 Experimental results
实验1	1	1	1	1	83.55
实验2	1	2	2	2	84.97
实验3	1	3	3	3	46.58
实验4	2	1	2	3	47.32
实验5	2	2	3	1	82.69
实验6	2	3	1	2	83.77
实验7	3	1	3	2	84.24
实验8	3	2	1	3	82.66
实验9	3	3	2	1	79.3
均值1	71.7	71.703	83.327	81.847
均值2	71.26	83.44	70.53	84.327
均值3	82.067	69.883	71.17	58.853
极差	10.807	13.557	12.797	25.474

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