Isolation and Enzymatic Characterization of Fungus Degrading Salvia miltiorrhiza Residue Lignin

doi:10.13560/j.cnki.biotech.bull.1985.2024-0419

Abstract

Abstract:

【Objective】 Selecting fungus that degrades Salvia miltiorrhiza residue（SMR）lignin and exploring its enzymatic characteristics is for having a strain to degrade SMR lignin for improving the enzymatic hydrolysis efficiency of cellulase. 【Method】 The strains naturally growing on the surface of SMR were selected as the screening source. Basic lignin medium and guaiacol medium were used for preliminary screening, and the medium added with SMR was used for re-screening to select the laccase-producing strains. The strains were identified by ITS sequence analysis. The enzymatic characteristics of laccase from isolated strains were studied. The content of lignocellulose was determined by NREL method, and the enzymolysis effect was investigated by commercial cellulase. 【Result】 A laccase-producing and SMR-tolerant fungus MY-5 was selected and identified as Trichoderma erinaceum by ITS sequence. The enzymatic properties of laccase were investigated. The optimal reaction temperature was 50℃, the enzyme activity was maintained at 85.3% after 24 h at 60℃, and 80.7% after 25 d of low temperature storage at 4℃. The optimal pH was 5, and the activity remained above 90% at pH 4-8 for 24 h. Adding Cu²⁺ and Mg²⁺ and vanillin increased the activity of laccase, and 5 mmol/L Cu²⁺ increased to 1.68 times. Adding 4% SMR and holding it for 24 h, 90.6% laccase activity still was maintained. After 20 d of liquid fermentation, SMR lignin degradation rate reached 36.3%, and the glucose yield and maximum yield increased by 62.6% and 81.0%, respectively. 【Conclusion】 A laccase-producing and SMR-tolerant T. erinaceum MY-5 had been isolated, which had good thermal stability, low temperature storage stability, pH stability and resistance to SMR, which may effectively degrade SMR lignin and improve the efficiency of cellulolytic enzyme hydrolysis, providing a basis for the bio-utilization of Chinese medicinal residue.

Key words: Salvia miltiorrhiza residue, Trichoderma erinaceum, laccase, enzymatic characterization, delignification

YANG Bing-qian, YUN Chen-ke, CHANG Si-yuan, GUO Sheng, ZHANG Sen. Isolation and Enzymatic Characterization of Fungus Degrading Salvia miltiorrhiza Residue Lignin[J]. Biotechnology Bulletin, 2024, 40(11): 269-276.

Figures/Tables 9

Fig. 1 Colorimetric reactions of 4 fungi A: YY3. B: YY3-A1. C: BQ-A3. D: MY-5

Fig. 2 Laccase activity in 4 fungi fermenting in the corn straw and SMR fermentation The error line in the figure refers to the standard deviation. The lowercase letters indicate that the difference among different groups reached a significant level（P < 0.05）. The same below

Fig. 3 Identification of strain MY-5 A: MY-5 colony morphology. B: MY-5 phylogenetic tree

Fig. 4 Effect of temperature on laccase activity A: Optimal laccase temperature. B: Laccase thermal stability

Fig. 5 Effect of pH on laccase activity A: Optimal laccase pH. B: Laccase pH stability

Fig. 6 Effects of metal ions and inducers on laccase activity Group 1: Metal ion concentration is 1 mmol/L and vanillin concentration is 0.01 g/L. Group 2: Metal ion concentration is 5 mmol/L and vanillin concentration is 0.05 g/L

Fig. 7 Effect of SMR on laccase activity

Table 1 Lignocellulose content changes in SMR before and after fermentation

组别Group	酸溶性木质素Acid soluble lignin/%	酸不溶性木质素Acid insoluble lignin/%	纤维素Cellulose/%	半纤维素Hemicellulose/%
未处理Untreated	4.53±1.06a	10.58±1.36a	17.46±1.27a	14.02±1.67a
处理后Treated	3.07±0.59b	6.56±0.47b	15.06±0.92b	7.32±0.71b

Fig. 8 Glucose yield and enzymatic hydrolysis efficiency of SMR before and after treatment

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