内生菌产纤维素酶辅助提取黑果枸杞花色苷工艺的优化

Optimization of anthocyanins extraction from Lycium ruthenicum Murr. assisted by cellulase from endophyte

中文关键词:  黑果枸杞  内生菌  纤维素酶  花色苷

英文关键词:Lycium ruthenicum Murr.  endophyte  cellulase  anthocyanins

中文摘要:

      目的 从黑果枸杞中筛选分离产纤维素酶菌株, 并优化纤维素酶辅助提取黑果枸杞花色苷工艺。方法 采用刚果红染色法和3,5-二硝基水杨酸比色法筛选黑果枸杞中的产纤维素酶菌株; 结合形态学与16S rRNA测序鉴定菌株; 通过单因素实验筛选菌株生长及产酶条件; 经单因素与正交实验优化酶辅助提取花色苷工艺。结果 分离筛选出8株产纤维素酶菌株, 其中菌株LRR-2辅助提取花色苷效果最佳, 经鉴定为贝莱斯芽孢杆菌(Bacillus velezensis); 其最适生长条件为: 碳源为淀粉, 氮源为酵母提取物, pH 4.0, 碳氮比1:2.5; 最适产酶条件为: 碳源为蔗糖, 氮源为酵母提取物, pH 5.0, 碳氮比1:1.0。酶辅助提取黑果枸杞果实花色苷最优工艺条件为: 液料比20:1 mL/g、乙醇浓度70%、提取温度40 ℃、醇提时间0.5 h、pH 4.0、加酶量20 U/mL、酶解时间2 h, 提取得率达(10.35±0.12) mg/g。结论 本研究为黑果枸杞花色苷高效酶辅助提取提供了具有特异性的优质菌种与优化工艺。

英文摘要:

      Objective To screen and isolate cellulase-producing strains from Lycium ruthenicum Murr. and optimize the cellulase-assisted extraction process for its anthocyanins. Methods Cellulase-producing strains were screened from Lycium ruthenicum Murr. using Congo red staining and the 3,5-dinitrosalicylic acid colorimetric assay. The isolates were identified based on morphological characteristics and 16S rRNA gene sequencing. The optimal conditions for bacterial growth and cellulase production were determined through single-factor experiments. Finally, the enzyme-assisted extraction process for anthocyanins was optimized by combining single-factor and orthogonal array designs. Results The 8 cellulase-producing strains were isolated from Lycium ruthenicum Murr.. Among them, strain LRR-2 demonstrated the highest efficiency in assisting anthocyanins extraction and was identified as Bacillus velezensis. The optimal growth conditions for LRR-2 were determined as follows: Starch as the carbon source, yeast extract as the nitrogen source, pH 4.0 and a carbon-to-nitrogen ratio of 1:2.5. Meanwhile, the optimal enzyme production conditions were: Sucrose as the carbon source, yeast extract as the nitrogen source, pH 5.0, and a carbon-to-nitrogen ratio of 1:1.0. The optimal conditions for the enzyme-assisted extraction of anthocyanins from Lycium ruthenicum Murr. fruits were determined as follows: Liquid-to-solid ratio of 20:1 mL/g, ethanol concentration of 70%, extraction temperature of 40 ℃, ethanol extraction time of 0.5 h, pH of 4.0, enzyme dosage of 20 U/mL and enzymatic hydrolysis time of 2 h. Under these optimized conditions, the anthocyanins extraction yield reached (10.35±0.12) mg/g. Conclusion This study provides a specific and high-quality microbial strain and an optimized process for the efficient enzyme-assisted extraction of anthocyanins from Lycium ruthenicum Murr..