摘　要：本文通过采用响应曲面法优化小球藻蛋白质溶出工艺，并对其二级结构进行表征。以小球藻蛋白质的溶出率为指标，考察超声功率、超声时间、NaOH质量分数及水浴时间对提取率的影响。在Plackett-Burman试验的基础上，采用4因素3水平的响应面分析法确定小球藻蛋白溶出工艺。结果表明，小球藻蛋白质最佳溶出工艺条件为超声功率460.08 W、超声时间16.05 min、NaOH质量分数14.03%、水浴时间37.18 min，溶出率达87.64%。在最优条件下溶出的小球藻蛋白经等电点法提取且纯化后进行傅立叶变换红外光谱分析、微观结构分析以及氨基酸分析，结果表明，小球藻蛋白二级结构中主要为β-折叠，含量为30.00%；微观结构显示，小球藻蛋白表面光滑且有孔洞结构；含有5种必需氨基酸，占总量的42.71%。综上，超声辅助碱法破碎小球藻蛋白的破壁程度显著，蛋白溶出率高，且操作简单、成本低，具有较好的工业化应用前景。该研究为小球藻蛋白的食品应用提供了相关参考。
关键词：小球藻破壁；蛋白溶出工艺；超声处理；碱法破碎；氨基酸
中图分类号：TS254                              文献标志码：ADOI：10.3969/j.issn.1007-7146.2024.04.007

Abstract: This study optimized the extraction process of Chlorella protein using response surface methodology and characterized its secondary structure. With the protein solubility rate of Chlorella as the indicator, the effects of ultrasonic power, ultrasonic time, NaOH mass fraction, and water bath time on the extraction rate were investigated. Based on the Plackett-Burman design, a four-factor three-level response surface analysis was conducted to determine the extraction process of Chlorella protein. The results showed that the optimal extraction conditions for Chlorella protein were ultrasonic power of 460.08 W, ultrasonic time of 16.05 min, NaOH mass fraction of 14.03%, and water bath time of 37.18 min, with a solubility rate of 87.64%. The extracted Chlorella protein was purified using isoelectric focusing under optimal conditions and then subjected to FTIR analysis, microscopic structure analysis, and amino acid analysis. The results indicated that the secondary structure of Chlorella protein mainly consisted of β-sheet, accounting for 30.00% of the total content. The microscopic structure revealed a smooth surface with pore structures, and it contained five essential amino acids, accounting for 42.71% of the total content. In conclusion, the ultrasonic-assisted alkaline method demonstrates significantly improved protein wall-breaking efficiency and high protein extraction rate for Chlorella, with simple operation and low cost. It has promising prospects for industrial applications and provides relevant references for the food application of Chlorella protein.
Key words: Chlorella breaks the wall; protein dissolution process; ultrasonic treatment; alkaline crushing; amino acid
（Acta Laser Biology Sinica, 2024, 33(4): 347-356）