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Enzymatic Synthesis of β-Sitosterol Laurate by Candida rugosa Lipase AY30 in the Water/AOT/Isooctane Reverse Micelle

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Abstract

Phytosterols are regarded as compounds able to reduce total and low-density lipoprotein cholesterol in the blood, and their esterified derivatives could help to improve the effectiveness of this function. In the present study, the water/sodium 1,4-bis-2-ethylhexylsulfosuccinate (AOT)/isooctane reverse micelle (RM) system was set up as a reaction medium for Candida rugosa lipase AY30 (CRL AY30) to synthesize β-sitosterol laurate (β-SLE). The product was identified by TLC, FT-IR, and HPLC–APCI–QqQ–MS/MS and quantified by HPLC. Through stepwise optimization, it was found that CRL AY30 had the highest activity in the water/AOT/isooctane RM system where 50 mM PBS with a pH of 7.5 was adopted as water core to carry CRL AY30, and the proportion of [CRL AY30] (mg/mL), [water] (mM), and [AOT] (mM) was set in 3:375:25, respectively, in isooctane. After screened with single-factor experiments, the esterification reaction conditions in the CRL AY30–water/AOT/isooctane RM system were further optimized by the response surface method as follows: the mole ratio of β-sitosterol to lauric acid of 1:3.5 (25 mM β-sitosterol), the enzyme load of 18% (w/w total reactants), the reaction temperature of 47 °C, and the reaction time of 48 h. As a result, the maximum esterification rate was up to 88.12 ± 0.79%.

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Abbreviations

β-SLE:

β-Sitosterol laurate

PSE:

Phytosterol ester

RM:

Reverse micelle

Y%:

Yield rate

AY30–WAI RM:

Water/AOT/isooctane RM containing the CRL AY30

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Funding

The authors acknowledge the funding support from the China National Key R&D Program during the 13th Five-year Plan Period (Grant No. 2016YFD040140402), the National Natural Science Foundation of China (Grant No. 31872890), and the Research Program of Sate Key Laboratory of Food Science and Technology in Nanchang University (Project SKLF-ZZB-201709).

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Authors and Affiliations

  1. State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047, China

    Sunni Chen, Jing Li, Hongyan Li, Bin Zhang, Liufeng Zheng & Zeyuan Deng

  2. The State Centre of Quality Supervision and Inspection for Camellia Products, Ganzhou, 341000, Jiangxi, China

    Zejian Fu

  3. Yichun Dahaigui Life Science Co., Ltd, Yichun, 336000, Jiangxi, China

    Guohua Wei

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  1. Sunni Chen
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Correspondence to Zeyuan Deng.

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Chen, S., Li, J., Fu, Z. et al. Enzymatic Synthesis of β-Sitosterol Laurate by Candida rugosa Lipase AY30 in the Water/AOT/Isooctane Reverse Micelle. Appl Biochem Biotechnol 192, 392–414 (2020). https://doi.org/10.1007/s12010-020-03302-0

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