Antibacterial activities and optimization of fermentation conditions of lipopeptides produced by Aureobasidium pullulans PA-2

doi:10.3969/j.issn.1004-1524.2021.03.13

Acta Agriculturae Zhejiangensis ›› 2021, Vol. 33 ›› Issue (3): 479-489.DOI: 10.3969/j.issn.1004-1524.2021.03.13

• Plant Protection • Previous Articles Next Articles

Antibacterial activities and optimization of fermentation conditions of lipopeptides produced by Aureobasidium pullulans PA-2

YUE Denggao, CHENG Liang^*(), GUO Qingyun

Academy of Agriculture and Forestry Science of Qinghai University, Key Laboratory of Agricultural Integrated Pest Management of Qinghai Province, Scientific Observing and Experimental Station of Crop Pest of Ministry of Agriculture and Rural Affairs, Xining 810016, China

Received:2020-07-26 Online:2021-03-25 Published:2021-03-25
Contact: CHENG Liang

Abstract

Abstract:

The antibacterial activities of lipopeptides from Aureobasidium pullulans PA-2 was explored, and the fermentation conditions were optimized with the output of lipopeptides as the response value, laying a theoretical foundation for the production and application of lipopeptides. The lipopeptides was crudely extracted by acid precipitation method. The crude extract of lipopeptide was qualitatively detected by in-situ acid hydrolysis and ninhydrin colorimetry. Antibacterial activities were measured by the method of agar drilling diffusion. The fermentation conditions were optimized by central composite design (CCD). Finally, the crude extract was identified as cyclic lipopeptide. The lipopeptides had obvious effects on Staphylococcus aureus, Saccharomyces cerevisiae, Escherichia coli, Mycosphaerella cerasella, Pyrenophorateres drechsler and antibacterial circle were 3.65, 1.95, 2.15, 1.35 and 2.18 cm respectively. The optimal fermentation conditions were: inoculation amount 6.8%, rotation speed 216 r·min ^-1, culture temperature 26 ℃, liquid volume 125 mL and initial pH 7.Under this condition, predicting yield of antibacterial lipopeptides by the model was 0.94 g·L ^-1, actually 0.92 g·L ^-1, which was 51% more than that of before(0.61 g·L ^-1). The optimized fermentation conditions increase the yield of the lipopeptide and reduce the fermentation cost, which may be applied to the control of strains with inhibitory effect above

Key words: Aureobasidium pullulans PA-2, lipopeptides, antibacterial activity, optimization of fermentation

CLC Number:

S476.8

YUE Denggao, CHENG Liang, GUO Qingyun. Antibacterial activities and optimization of fermentation conditions of lipopeptides produced by Aureobasidium pullulans PA-2[J]. Acta Agriculturae Zhejiangensis, 2021, 33(3): 479-489.

Figures/Tables 8

References 29

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试验因素 Factor	试验水平Level
试验因素 Factor	-2.378	-1	0	+1	+2.378
接种量Inoculation amount (A)/%	2.24	5	7	9	11.76
转速Rotation speed(B)/(r·min^-1)	118.66	160	190	220	261.34
温度Temperature(C)/℃	20.24	23	25	27	29.80
装液量Liquid volume (D)/mL	31.10	100	150	200	268.90
初始pH Initial pH(E)	4.62	6	7	8	9.38

试验因素 Factor	试验水平Level
试验因素 Factor	-2.378	-1	0	+1	+2.378
接种量Inoculation amount (A)/%	2.24	5	7	9	11.76
转速Rotation speed(B)/(r·min^-1)	118.66	160	190	220	261.34
温度Temperature(C)/℃	20.24	23	25	27	29.80
装液量Liquid volume (D)/mL	31.10	100	150	200	268.90
初始pH Initial pH(E)	4.62	6	7	8	9.38

靶标菌 Target bacteria	平均抑菌圈直径 Antibacterial circle diameter/cm
金黄色葡萄球菌Staphylococcus aureus	3.65
酿酒酵母菌Saccharomyces cerevisiae	1.95
大肠埃希菌Escherichia coli	2.15
白色念珠菌Candida albicans	0
樱桃球腔菌Mycosphaerella cerasella	1.35
大麦网斑病Pyrenophora teres	2.18

靶标菌 Target bacteria	平均抑菌圈直径 Antibacterial circle diameter/cm
金黄色葡萄球菌Staphylococcus aureus	3.65
酿酒酵母菌Saccharomyces cerevisiae	1.95
大肠埃希菌Escherichia coli	2.15
白色念珠菌Candida albicans	0
樱桃球腔菌Mycosphaerella cerasella	1.35
大麦网斑病Pyrenophora teres	2.18

方差来源 Soruces of variation	平方和 Sum of squares	自由度 Degree of freedom	方差 Variance	F值 F value	P值 P value	显著性 Significance
回归模型Regression model	2.06	20	0.10	89.28	<0.000 1	**
A	1.426×10^-3	1	1.426×10^-3	1.23	0.275 6
B	0.51	1	0.51	439.53	<0.000 1	**
C	0.029	1	0.029	24.76	<0.000 1	**
D	0.028	1	0.028	24.26	<0.000 1	**
E	3.887×10^-3	1	3.887×10^-3	3.37	0.076 9
AB	1.125×10^-4	1	1.125×10^-4	0.097	0.757 2
AC	0.025	1	0.025	21.92	<0.000 1	**
AD	1.800×10^-3	1	1.800×10^-3	1.56	0.221 9
AE	3.613×10^-3	1	3.613×10^-3	3.13	0.087 5
BC	0.016	1	0.016	14.03	0.000 8	**
BD	0.019	1	0.019	16.46	0.000 3	**
BE	2.450×10^-3	1	2.450×10^-3	2.12	0.156 0
CD	0.20	1	0.20	169.10	<0.000 1	**
CE	4.500×10^-4	1	4.500×10^-4	0.39	0.537 4
DE	1.125×10^-4	1	1.125×10^-4	0.097	0.757 2
A2	0.32	1	0.32	275.38	<0.000 1	**
B2	0.15	1	0.15	129.08	<0.000 1	**
C2	0.30	1	0.30	256.42	<0.000 1	**
D2	0.54	1	0.54	468.04	<0.000 1	**
E2	0.51	1	0.51	443.22	<0.000 1	**
残差Residual	0.033	29	1.155×10^-3
失拟项Lack of fit	0.026	22	1.161×10^-3	1.02	0.528 6
纯误差Pure error	7.950×10^-3	7	1.136×10^-3
合计Cor total	2.10	49

Antibacterial activities and optimization of fermentation conditions of lipopeptides produced by Aureobasidium pullulans PA-2

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序号 No.	A	B	C	D	E	脂肽产量 Lipopeptide yield/(g·L^-1)	预测值 Predicted value/(g·L^-1)
1	0	-2.38	0	0	0	0.32	0.31
2	0	2.38	0	0	0	0.82	0.83
3	1	-1	1	1	-1	0.26	0.25
4	1	-1	1	1	1	0.25	0.26
5	0	0	0	0	0	0.83	0.86
6	0	0	0	2.38	0	0.25	0.24
7	0	0	0	0	0	0.89	0.86
8	0	0	0	0	0	0.82	0.86
9	-1	-1	1	-1	1	0.45	0.48
10	1	-1	-1	1	-1	0.46	0.45
11	-1	1	1	-1	-1	0.77	0.76
12	-1	-1	-1	1	1	0.45	0.44
13	-1	-1	-1	-1	1	0.28	0.27
14	1	1	1	1	1	0.48	0.45
15	1	1	-1	-1	1	0.52	0.53
16	-1	1	-1	1	-1	0.56	0.51
17	1	1	-1	1	-1	0.59	0.58
18	0	0	0	0	2.38	0.36	0.34
19	-1	1	1	1	-1	0.50	0.52
20	1	1	1	-1	-1	0.78	0.75
21	-2.38	0	0	0	0	0.45	0.42
22	-1	-1	1	1	-1	0.34	0.28
23	-1	1	-1	1	1	0.54	0.55
24	-1	-1	1	1	1	0.33	0.34
25	-1	1	-1	-1	1	0.45	0.47
26	0	0	0	-2.38	0	0.36	0.36
27	-1	-1	1	-1	-1	0.39	0.43
28	1	-1	-1	-1	1	0.33	0.33
29	-1	1	1	-1	1	0.81	0.77
30	-1	1	-1	-1	-1	0.43	0.45
31	-1	1	1	1	1	0.49	0.54
32	0	0	-2.38	0	0	0.38	0.39
33	1	-1	1	-1	-1	0.42	0.43
34	1	1	1	1	-1	0.45	0.48
35	1	-1	-1	-1	-1	0.34	0.31
36	0	0	2.38	0	0	0.52	0.51
37	2.38	0	0	0	0	0.42	0.45
38	0	0	0	0	-2.38	0.28	0.30
39	1	-1	1	-1	1	0.44	0.43
40	0	0	0	0	0	0.84	0.86
41	1	-1	-1	1	1	0.46	0.47
42	-1	-1	-1	-1	-1	0.23	0.21
43	0	0	0	0	0	0.85	0.86
44	-1	-1	-1	1	-1	0.30	0.37
45	1	1	1	-1	1	0.71	0.72
46	1	1	-1	1	1	0.60	0.58
47	0	0	0	0	0	0.86	0.86
48	0	0	0	0	0	0.90	0.86
49	0	0	0	0	0	0.91	0.86
50	1	1	-1	-1	-1	0.55	0.55

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