霉菌酯化酶作用规律及应用技术研究
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摘要
酯化酶是一类能使呈香前体物质转化为香味物质的酶的总称,其酶制剂应用于白酒生产中,起到增加白酒酯类含量,从而提高白酒优质率的作用,但在实际应用中存在酶活低、生香效果不稳定、增香针对性不强、使用范围不广等问题。本论文以明确霉菌酯化酶的生香作用规律和条件,提高其在白酒生产中应用效果和范围为目标,研究了根霉酯化酶、红曲酯化酶、功能性大曲三种酯化酶的不同酸底物特异性、水相合成己酸乙酯技术和黄水酯化液制备技术条件,主要结果和结论如下:
1.霉菌酯化酶不同酸底物特异性研究:三种霉菌酯化酶对乙酸、丁酸、戊酸、己酸、庚酸、辛酸与乙醇酯化合成相应乙酯的酯化率随所选酸底物碳原子数的增加而增加,但都对乳酸乙酯的生成有一定的抑制作用,原因可能与乳酸的化学结构有关。不同霉菌酯化酶对同一脂肪酸的酯化作用效果不同,其中根霉酯化酶对乙酸、乳酸、庚酸具有较高的酯化率;红曲酯化酶对辛酸有较高的酯化率;功能性大曲对丁酸、戊酸、己酸有较高的酯化率。混合酸与乙醇在不同霉菌酯化酶的作用下效果不同,其中根霉酯化酶、红曲酯化酶和功能性大曲都对己酸有较高的酯化率。
2.霉菌酯化酶水相己酸乙酯合成技术的研究:三种霉菌酯化酶的适宜催化温度范围为35～40℃,适宜催化己酸浓度为1mL/100mL;根霉酯化酶和功能性大曲合成己酸乙酯适宜的乙醇浓度为10mL/100mL,红曲酯化酶对乙醇有较强耐受性,其适宜乙醇浓度为12mL/100mL;红曲酯化酶和功能性大曲的适宜酯化酶用量是7g/100mL,而根霉酯化酶适宜用量为9g/100mL。通过正交实验优化功能性大曲在水相中合成己酸乙酯条件为:己酸浓度为1.5mL/100mL,乙醇浓度为10mL/100mL,功能性大曲用量9g/100mL,催化温度为40℃。
3.黄水酯化液制备技术的研究:浓香型白酒生产过程中的副产物黄水和酒尾中含有大量的酯类前体物质,以黄水、酒尾为主要原料,采用酯化酶酶法制备酯化液,优化得出的酯化液酶法制备参数为:黄水、水与酒尾的比例为2:2:1,乙醇浓度10mL/100mL,己酸添加量1.5mL/100mL,功能性大曲用量7g/100mL,在温度30℃的条件下,酯化7d,得到的酯化液中香味物质含量高,比例适宜,风味协调,经处理可应用于提高浓香型白酒主体香味物质的含量,提高白酒优质率。
Esterifying enzyme is a kind of enzyme which can make precursor substance form flavor components.Esterifying enzyme of Rhizopus sp, esterifying enzyme of Monascus sp and functional daqu are mainly used in liquor-making industry to increase the main fragrance of luzhou-flavour liquor, but there are many practical problems, such as low enzyme activity, unstable effect and unwide usage and so on. In order to definite the action rules and improve application of mold esterifying enzymes, this text studied the substrate specifity of mold esterifying enzymes, the technique of synthesis of ethyl hexanoate in aqueous medium and the preparation technique of esterifying liquid . Primarily study results as follows:
1. Study on the Substrate Specifity of Mold Esterifying Enzymes: The esterification rate of acetic acid, butyric acid, valeric acid, hexanoic acid, heptanoic acid and octanoic acid increased with the increase of acid carbon number by the three different esterifying enzymes. The three esterifying enzymes inhibited the formation of ethyl lactate.The reason might be related to the chemical structure of lactic acid.There were different synthesis effects of different enzymes. The esterification rates of acetic acid, lactic acid and heptanoic acid were higher by esterifying enzyme of Rhizopus sp.The esterification rate of octanoic acid was higher by esterifying enzyme of Monascus sp.The esterification rates of butyric acid, valeric acid and hexanoic acid were higher by functional daqu. But ethyl hexanoate was highly synthesized by all different enzymes in the reaction system of four-mixed acid and ethanol
2. Research on the Technique of Synthesis of Ethyl Hexanoate in Aqueous Medium:The conditions of synthesis of ethyl hexanoate in aqueous medium were different by different kinds of esterifying enzymes.The appropiate conditions for three esterifying enzymes were : the temperature 35~40℃; the concentration of hexanoic acid 1mL/100mL; the concentration of ethanol 10~12mL/100mL;the dosage of esterifying enzyme7~9g/100mL.The optimum conditions for the synthesis of ethyl hexanoate by mixed esterifying enzyme were:the concentration of ethanol 10mL/100mL,the concentration of hexanoic acid 1.5mL/100mL,the dosage of functional daqu 9g/100mL and temperature 40℃。
3. Research on the Preparation Technique of Esterifying Liquid:The preparation technique of esterifying liquid was studied by using yellow water, ending liquor and functional daqu as main materals.With the aid of single factor experiments and orthogonal experiment, the optimal preparation technique of esterifying liquid was the concentration of ethanol 10mL/100mL, the addition of hexanoic acid 1.5mL/100mL,the dosage of functional daqu 7g/100mL and temperature 30℃.
1.霉菌酯化酶不同酸底物特异性研究:三种霉菌酯化酶对乙酸、丁酸、戊酸、己酸、庚酸、辛酸与乙醇酯化合成相应乙酯的酯化率随所选酸底物碳原子数的增加而增加,但都对乳酸乙酯的生成有一定的抑制作用,原因可能与乳酸的化学结构有关。不同霉菌酯化酶对同一脂肪酸的酯化作用效果不同,其中根霉酯化酶对乙酸、乳酸、庚酸具有较高的酯化率;红曲酯化酶对辛酸有较高的酯化率;功能性大曲对丁酸、戊酸、己酸有较高的酯化率。混合酸与乙醇在不同霉菌酯化酶的作用下效果不同,其中根霉酯化酶、红曲酯化酶和功能性大曲都对己酸有较高的酯化率。
2.霉菌酯化酶水相己酸乙酯合成技术的研究:三种霉菌酯化酶的适宜催化温度范围为35～40℃,适宜催化己酸浓度为1mL/100mL;根霉酯化酶和功能性大曲合成己酸乙酯适宜的乙醇浓度为10mL/100mL,红曲酯化酶对乙醇有较强耐受性,其适宜乙醇浓度为12mL/100mL;红曲酯化酶和功能性大曲的适宜酯化酶用量是7g/100mL,而根霉酯化酶适宜用量为9g/100mL。通过正交实验优化功能性大曲在水相中合成己酸乙酯条件为:己酸浓度为1.5mL/100mL,乙醇浓度为10mL/100mL,功能性大曲用量9g/100mL,催化温度为40℃。
3.黄水酯化液制备技术的研究:浓香型白酒生产过程中的副产物黄水和酒尾中含有大量的酯类前体物质,以黄水、酒尾为主要原料,采用酯化酶酶法制备酯化液,优化得出的酯化液酶法制备参数为:黄水、水与酒尾的比例为2:2:1,乙醇浓度10mL/100mL,己酸添加量1.5mL/100mL,功能性大曲用量7g/100mL,在温度30℃的条件下,酯化7d,得到的酯化液中香味物质含量高,比例适宜,风味协调,经处理可应用于提高浓香型白酒主体香味物质的含量,提高白酒优质率。
Esterifying enzyme is a kind of enzyme which can make precursor substance form flavor components.Esterifying enzyme of Rhizopus sp, esterifying enzyme of Monascus sp and functional daqu are mainly used in liquor-making industry to increase the main fragrance of luzhou-flavour liquor, but there are many practical problems, such as low enzyme activity, unstable effect and unwide usage and so on. In order to definite the action rules and improve application of mold esterifying enzymes, this text studied the substrate specifity of mold esterifying enzymes, the technique of synthesis of ethyl hexanoate in aqueous medium and the preparation technique of esterifying liquid . Primarily study results as follows:
1. Study on the Substrate Specifity of Mold Esterifying Enzymes: The esterification rate of acetic acid, butyric acid, valeric acid, hexanoic acid, heptanoic acid and octanoic acid increased with the increase of acid carbon number by the three different esterifying enzymes. The three esterifying enzymes inhibited the formation of ethyl lactate.The reason might be related to the chemical structure of lactic acid.There were different synthesis effects of different enzymes. The esterification rates of acetic acid, lactic acid and heptanoic acid were higher by esterifying enzyme of Rhizopus sp.The esterification rate of octanoic acid was higher by esterifying enzyme of Monascus sp.The esterification rates of butyric acid, valeric acid and hexanoic acid were higher by functional daqu. But ethyl hexanoate was highly synthesized by all different enzymes in the reaction system of four-mixed acid and ethanol
2. Research on the Technique of Synthesis of Ethyl Hexanoate in Aqueous Medium:The conditions of synthesis of ethyl hexanoate in aqueous medium were different by different kinds of esterifying enzymes.The appropiate conditions for three esterifying enzymes were : the temperature 35~40℃; the concentration of hexanoic acid 1mL/100mL; the concentration of ethanol 10~12mL/100mL;the dosage of esterifying enzyme7~9g/100mL.The optimum conditions for the synthesis of ethyl hexanoate by mixed esterifying enzyme were:the concentration of ethanol 10mL/100mL,the concentration of hexanoic acid 1.5mL/100mL,the dosage of functional daqu 9g/100mL and temperature 40℃。
3. Research on the Preparation Technique of Esterifying Liquid:The preparation technique of esterifying liquid was studied by using yellow water, ending liquor and functional daqu as main materals.With the aid of single factor experiments and orthogonal experiment, the optimal preparation technique of esterifying liquid was the concentration of ethanol 10mL/100mL, the addition of hexanoic acid 1.5mL/100mL,the dosage of functional daqu 7g/100mL and temperature 30℃.
引文
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