Medium Optimization for Laccase Production by Acrophialophora sp. Z45 and Its Decolorization of Dyes

doi:10.13560/j.cnki.biotech.bull.1985.2021-1221

Abstract

Abstract:

The strain Acrophialophora sp. Z45 is a laccase-producing fungus,which was discovered in the previous study of our laboratory. In this study,based on the size of guaiacol-oxidation zone on different medium,the factors influencing the laccase production by strain Z45 were studied. The orthogonal experiments were designed to optimize the basic medium for laccase production and the difference in laccase activity before and after medium optimization was then compared. Finally,the ability of strain Z45 to decolorize eight synthetic dyes was evaluated based on the correlation between laccase and dye decolorization. The results based on the single factor experiment and orthogonal test showed that the optimized medium for laccase production was as follows：sucrose as carbon source,sodium nitrate as nitrogen source,the carbon to nitrogen ratio was 45∶1,with a pH value of 5.0. The laccase activity of strain Z45 was significantly improved after medium optimization. Strain Z45 had certain decolorization ability to bromophenol blue,neutral red,methylene blue,methyl blue and crystalline violet,among which the decolorization ability to triphenylmethane dye methyl blue was the strongest. The crude enzyme solution of strain Z45 decolorized methyl blue rapidly. On the solid medium with lower concentrations of methyl blue,the time for complete decolorization of methyl blue was not affected by the dye concentration,but at higher concentrations of methyl blue,the time for complete decolorization was gradually extended with the increase of dye concentration.

Key words: Acrophialophora sp., laccase, orthogonal test, dye decolorization

JIA Chen-bo, SU Yi-huang, MA Xiu-mei, WANG Chun-li, XU Chun-yan. Medium Optimization for Laccase Production by Acrophialophora sp. Z45 and Its Decolorization of Dyes[J]. Biotechnology Bulletin, 2022, 38(6): 252-260.

Figures/Tables 10

Fig. 1 Basic performance of strain Z45 on the G-PDA and PDA medium 1d-FS,2d-FS,4d-FS and 6d-FS indicate the growth and discoloration circles on the front side of 1,2,4 and 6 d,respectively;1d-BS,2d-BS,4d-BS and 6d-BS indicate the growth and discoloration circles on the back side of 1,2,4 and 6 d,respectively

Fig.2 Diameter of discoloration circle under different condition A：horizontal ordinate indicates the carbon source is glucose,sucrose,lactose,maltose,starch,sodium carboxymethyl cellulose,respectively;B：horizontal ordinate indicates the nitrogen source is sodium nitrate,ammonium nitrate,ammonium sulfate,peptone,yeast powder,and urea from left to right,respectively. Different letters indicates significant difference（P<0.05）

Table 1 Results of orthogonal test and the visual analysis

编号 No.	A（碳源） A（Carbon source）	B（氮源） B（Nitrogen source）	C（碳氮比） C（C/N）	D（pH） D（pH）	变色圈直径 Diameter of discoloration circle/mm
1	淀粉Starch	硝酸钠Sodium nitrate	35∶1	4.5	13.33
2	淀粉Starch	硝酸铵Ammonium nitrate	40∶1	5.0	10.00
3	淀粉Starch	硫酸铵Ammonium sulfate	45∶1	5.5	10.67
4	乳糖Lactose	硝酸钠Sodium nitrate	40∶1	5.5	14.00
5	乳糖Lactose	硝酸铵Ammonium nitrate	45∶1	4.5	12.00
6	乳糖Lactose	硫酸铵Ammonium sulfate	35∶1	5.0	9.33
7	蔗糖Sucrose	硝酸钠Sodium nitrate	45∶1	5.0	23.33
8	蔗糖Sucrose	硝酸铵Ammonium nitrate	35∶1	5.5	14.67
9	蔗糖Sucrose	硫酸铵Ammonium sulfate	40∶1	4.5	12.00
k1	11.33	16.89	12.44	12.44
k2	11.78	12.22	12.00	14.22
k3	16.67	10.67	15.33	13.11
MAX	16.67	16.89	15.33	14.22
极差R	5.34	6.22	3.33	1.78

Table 2 ANOVA of the orthogonal test

方差来源 Source of variance	III 型平方和 Type III sum of squares	自由度 Degree of freedom	均方 Mean square	F值 F value	显著性 Significance
A	144.296	2	72.148	20.292	**
B	199.407	2	99.704	28.042	**
C	64.296	2	32.148	9.042	**
D	18.074	2	9.037	2.542
误差Error	64.000	18	3.556

Fig. 3 Laccase activities in different media

Fig.4 Decolorization rate of methyl blue with different amount of crude laccase

Fig.5 Decolorization of methyl blue by strain Z45

Fig.6 Decolorization of methyl blue under different concentrations on PDA medium

Fig. 7 Decolorization of methyl blue under different conc-entrations on the optimized medium for laccase pro-duction

Fig. 8 Decolorization of methyl blue under different conc-entrations by crude enzyme solution

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