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| 降胆固醇亚麻籽蛋白酶解肽的分离纯化及结构鉴定 |
| Isolation and identification of peptide with cholesterol lowering activity derived from flaxseed protein |
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| DOI： |
| 中文关键词: 亚麻籽蛋白 亚麻籽肽 降胆固醇活性 分离纯化 结构鉴定 |
| 英文关键词:flaxseed protein flaxseed peptides cholesterol micelle solubity isolation and purification identification |
| 基金项目:内蒙古自治区科技计划项目（2020GG0064）；国家自然科学基金地区科学基金（31860423） |
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| 中文摘要: |
| 采用Protease M对亚麻籽蛋白进行酶解，制备具有降胆固醇活性的亚麻籽蛋白酶解肽，对其进行分离纯化及结构鉴定，并合成相应多肽验证其降胆固醇活性。结果表明，经Protease M对亚麻籽蛋白酶解4 h时获得的亚麻籽蛋白酶解肽胆固醇胶束溶解度抑制率最高，为47.57%；继续采用超滤技术将其分离为相对分子质量小于等于3 kDa、3~5 kDa、5~10 kDa、10~30 kDa和大于30 kDa 5个组分，发现相对分子质量小于等于3 kDa的组分胆固醇胶束溶解度抑制率最高，为71.0%；再经大孔树脂对该组分进行吸附后经过不同体积分数乙醇溶液洗脱，发现75%的乙醇洗脱分离得到的组分胆固醇胶束溶解度抑制率最高，为79.8%；采用反相高效液相色谱（RP-HPLC）对其进一步分离纯化，收集到F9组分的胆固醇胶束溶解度抑制率最高，为85.7%；最后采用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS/MS)从F9组分中鉴定出6种降胆固醇亚麻籽肽，氨基酸序列分别为Ile-Ile-Pro-Ala-Phe(IIPAF)、Leu-Asn-Phe-Phe(LNFF)、Leu-Leu-Gly-Thr-Leu(LLGTL)、Ile-Pro-Pro-Phe(IPPF)、Ile-Ile-Phe(IIF)和Leu-Leu-Gly-Ala(LLGA)，其胆固醇胶束溶解度抑制率分别为82.8%、77.8%、88.0%、93.5%、80.3%和87.1%。 |
| 英文摘要: |
| Protease M was selected to hydrolyze flaxseed protein to prepare flaxseed protein hydrolysate peptides with cholesterol lowering activity. The peptides were isolated, purified and identified. The corresponding polypeptides were synthesized to verify the cholesterol lowering activity.The results showed that the highest inhibition rates of cholesterol micelle solubity of flaxseed protein hydrolysate peptide was 47.57% when it was hydrolyzed for 4 h by protease M, and it was further separated into five components with molecular weight ≤3 kDa, 3-5 kDa, 5-10 kDa, 10-30 kDa and >30 kDa by ultrafiltration technology, and inhibition rates of cholesterol micelle solubity of components with molecular weight ≤3 kDa was the highest,reaching 71.0%.The fraction with molecular weight ≤3 kDa was adsorbed by macroporous resin and then eluted by ethanol of different volume fraction, and it was found that the fraction eluted by 75% ethanol had the highest inhibition rates of cholesterol micelle solubity of 79.8%. RP-HPLC was used to further separate and purify the fraction,and the inhibition rates of cholesterol micelle solubity of F9 fraction was the highest of 85.7%. Finally, six cholesterol lowering flaxseed peptides were identified from F9 fraction by MALDI-TOF-MS/MS,and their amino acid sequences were Ile-Ile-Pro-Ala-Phe(IIPAF),Leu-Asn-Phe-Phe(LNFF),Leu-Leu-Gly-Thr-Leu(LLGTL),Ile-Pro-Pro-Phe(IPPF),Ile-Ile-Phe(IIF) and Leu-Leu-Gly-Ala(LLGA) with the inhibition rates of cholesterol micelle solubility 82.8%, 77.8%, 88.0%, 93.5%, 80.3% and 871%,respectively. |
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