脂肪酶催化合成α-亚麻酸植物甾醇酯的研究
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摘要
植物甾醇和α-亚麻酸均能够降低人体的血脂含量,减少心血管疾病的出现几率,但是植物甾醇脂溶性差,在油脂中溶解度很低,而α-亚麻酸氧化稳定性较差,两者经过酯化交联改性以后能改善上述特性,扩大其应用范围。
酶法催化酯化反应具有条件温和、产品安全性高的优点,因此应用广泛,但是在催化制备α-亚麻酸甾醇酯产品目前国内外还无人报道。本论文旨在尝试采用脂肪酶催化法合成α-亚麻酸植物甾醇酯,并对所得产品进行结构表征和理化特性研究。
主要研究结果如下:
1.酶催化合成α-亚麻酸植物甾醇酯的工艺研究
通过单因素和正交试验探讨酶法合成α-亚麻酸植物甾醇酯的影响因素,表明对反应酯化率的影响大小依次为:反应温度＞物料比＞酶添加量,最佳工艺条件为:选取Novozyme 435为反应催化剂,添加量5%,混合溶剂(异辛烷:正己烷:环己烷=6:2:2,v/v/v)为反应溶剂,添加量为1.6:1(溶剂:底物,ml/g),反应温度55℃,α-亚麻酸和植物甾醇的质量比为3:1的时候,在此条件下酯化率为40.56%,而经进一步纯化后纯度可高达75%。
2.α-亚麻酸植物甾醇酯的结构表征
采取薄层色谱法、红外分光光度法、紫外分光光度法、气质联用色谱法和核磁共振法对所得到产品进行结构分析,通过与植物甾醇的图谱进行对比,表明纯化后反应产物不含有机酸、醛和游离羟基,通过多种手段均观察到有酯键的生成,证明纯化后产品为α-亚麻酸植物甾醇酯。GC-MS法分析表明酶法合成α-亚麻酸植物甾醇酯产品中反式脂肪酸含量小于0.1%。
3.α-亚麻酸植物甾醇酯的理化特性研究
对α-亚麻酸植物甾醇酯在不同植物油脂中的溶解度测定结果表明,植物甾醇在经过酯化改性后,其脂溶性提高了20-30倍;采取Schaal烘箱法和Rancimat诱导期测定法分别对α-亚麻酸植物甾醇酯和添加α-亚麻酸植物甾醇酯植物油脂的氧化稳定性进行评价,结果表明,α-亚麻酸植物甾醇酯诱导时间为15.46h,α-亚麻酸乙酯诱导时间2.93h,因此α-亚麻酸植物甾醇酯稳定性良好,但是其对植物油脂的酸价、过氧化值和诱导时间都存在一定不利影响;研究了α-亚麻酸植物甾醇酯和添加α-亚麻酸植物甾醇酯的植物油脂的结晶特性,结果表明,α-亚麻酸植物甾醇酯的结晶温度区间为-25.9℃--29.6℃,能降低大豆油和菜籽油的结晶温度6℃-10℃,扩大了其应用温度范围。
Plant sterol andα-linolenic both possessed activities of hypolipidemic activity and preventing cardiovascular diseases.However poor solubility of plant sterol in lipid and oxidation instability ofα-linolenic limited their application in food industry, which could be improved by converting the two compounds into phytosterolsα-linolenate.
Phytosterolsα-linolenate with high security could be obtained by lipase catalyzing esterification in gentle condition,but there were no reports on the research. In the paper,researches including synthesis of phytosterolsα-linolenate by lipase catalysis,physical and chemical properties,and structure of product were carried out.
The main conclusions were obtained as follows:
1.Synthesis technology of phytosterolsα-linolenate by lipase catalysis
We studied factors affecting esterification rate by single factor and orthogonal test,results showed that effect of temperature was the greatest,followed by substrate ratio and enzyme addition.Finally,we got the appropriate esterification conditions as follows:Novozyme 435 addition was 5%,mixed solvent proportion(iso-octane: hexane:cyclohexane) was 6:2:2(v/v/v),solvent and substrate ratio was 1.6:1 (ml/g),temperature was 55℃,and substrate ratio(α-linolenic:phytosterol) was 3:1. In the conditions,the esterification rate reached to 40.56%and finally to 75%by purification.
2.Structure of phytosterolsα-linolenate
Main ingredient in the products was confirmed as phytosterolsα-linolenate by TLC,FTIR,UV-VIS and NMR.Results showed that there were no organic acid、aldehyde and free hydroxyl existed in the product,and formation of ester bond was observed.The content of trans fatty acid in product was less than 0.1%by GC-MS.
3.Physical and chemical characteristics of phytosterolα-linolenate
Fat-solubility of plant sterol was increased by 20-30 times in plant oils after etserification.Inductive time of ethylα-linolenate was 15.46h and phytosterolsα-linolenate were 15.46h and 2.93h respectively,but phytosterolsα-linolenate made oxidation stability of plant oil decreased,which were observed by using Schaal-oven method and Rancimat instruments.Crystallization behavior was studied by DSC, crystallization temperature of phytosterolsα-linolenate was -25.9℃--29.6℃,and phytosterolsα-linolenate made the crystallization temperature of plant oil decreased 6℃-10℃.
酶法催化酯化反应具有条件温和、产品安全性高的优点,因此应用广泛,但是在催化制备α-亚麻酸甾醇酯产品目前国内外还无人报道。本论文旨在尝试采用脂肪酶催化法合成α-亚麻酸植物甾醇酯,并对所得产品进行结构表征和理化特性研究。
主要研究结果如下:
1.酶催化合成α-亚麻酸植物甾醇酯的工艺研究
通过单因素和正交试验探讨酶法合成α-亚麻酸植物甾醇酯的影响因素,表明对反应酯化率的影响大小依次为:反应温度＞物料比＞酶添加量,最佳工艺条件为:选取Novozyme 435为反应催化剂,添加量5%,混合溶剂(异辛烷:正己烷:环己烷=6:2:2,v/v/v)为反应溶剂,添加量为1.6:1(溶剂:底物,ml/g),反应温度55℃,α-亚麻酸和植物甾醇的质量比为3:1的时候,在此条件下酯化率为40.56%,而经进一步纯化后纯度可高达75%。
2.α-亚麻酸植物甾醇酯的结构表征
采取薄层色谱法、红外分光光度法、紫外分光光度法、气质联用色谱法和核磁共振法对所得到产品进行结构分析,通过与植物甾醇的图谱进行对比,表明纯化后反应产物不含有机酸、醛和游离羟基,通过多种手段均观察到有酯键的生成,证明纯化后产品为α-亚麻酸植物甾醇酯。GC-MS法分析表明酶法合成α-亚麻酸植物甾醇酯产品中反式脂肪酸含量小于0.1%。
3.α-亚麻酸植物甾醇酯的理化特性研究
对α-亚麻酸植物甾醇酯在不同植物油脂中的溶解度测定结果表明,植物甾醇在经过酯化改性后,其脂溶性提高了20-30倍;采取Schaal烘箱法和Rancimat诱导期测定法分别对α-亚麻酸植物甾醇酯和添加α-亚麻酸植物甾醇酯植物油脂的氧化稳定性进行评价,结果表明,α-亚麻酸植物甾醇酯诱导时间为15.46h,α-亚麻酸乙酯诱导时间2.93h,因此α-亚麻酸植物甾醇酯稳定性良好,但是其对植物油脂的酸价、过氧化值和诱导时间都存在一定不利影响;研究了α-亚麻酸植物甾醇酯和添加α-亚麻酸植物甾醇酯的植物油脂的结晶特性,结果表明,α-亚麻酸植物甾醇酯的结晶温度区间为-25.9℃--29.6℃,能降低大豆油和菜籽油的结晶温度6℃-10℃,扩大了其应用温度范围。
Plant sterol andα-linolenic both possessed activities of hypolipidemic activity and preventing cardiovascular diseases.However poor solubility of plant sterol in lipid and oxidation instability ofα-linolenic limited their application in food industry, which could be improved by converting the two compounds into phytosterolsα-linolenate.
Phytosterolsα-linolenate with high security could be obtained by lipase catalyzing esterification in gentle condition,but there were no reports on the research. In the paper,researches including synthesis of phytosterolsα-linolenate by lipase catalysis,physical and chemical properties,and structure of product were carried out.
The main conclusions were obtained as follows:
1.Synthesis technology of phytosterolsα-linolenate by lipase catalysis
We studied factors affecting esterification rate by single factor and orthogonal test,results showed that effect of temperature was the greatest,followed by substrate ratio and enzyme addition.Finally,we got the appropriate esterification conditions as follows:Novozyme 435 addition was 5%,mixed solvent proportion(iso-octane: hexane:cyclohexane) was 6:2:2(v/v/v),solvent and substrate ratio was 1.6:1 (ml/g),temperature was 55℃,and substrate ratio(α-linolenic:phytosterol) was 3:1. In the conditions,the esterification rate reached to 40.56%and finally to 75%by purification.
2.Structure of phytosterolsα-linolenate
Main ingredient in the products was confirmed as phytosterolsα-linolenate by TLC,FTIR,UV-VIS and NMR.Results showed that there were no organic acid、aldehyde and free hydroxyl existed in the product,and formation of ester bond was observed.The content of trans fatty acid in product was less than 0.1%by GC-MS.
3.Physical and chemical characteristics of phytosterolα-linolenate
Fat-solubility of plant sterol was increased by 20-30 times in plant oils after etserification.Inductive time of ethylα-linolenate was 15.46h and phytosterolsα-linolenate were 15.46h and 2.93h respectively,but phytosterolsα-linolenate made oxidation stability of plant oil decreased,which were observed by using Schaal-oven method and Rancimat instruments.Crystallization behavior was studied by DSC, crystallization temperature of phytosterolsα-linolenate was -25.9℃--29.6℃,and phytosterolsα-linolenate made the crystallization temperature of plant oil decreased 6℃-10℃.
引文
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