21世纪，人类面临着更为严峻的人口剧增、资源匾乏及环境污染的压力和挑战，开辟新资源、走可持续发展道路是我国农业产业化发展的必由之路。昆虫作为地球上最大的未被充分利用的资源宝库而倍受关注。昆虫蛋白质资源的开发及其产业化，必将进入以高科技、高收益为特点的多层次、综合利用阶段。昆虫富含蛋白质、不饱和脂肪酸、微量元素，是人类的高级营养源。
黄粉虫的蛋白质中含有18种氨基酸，且包含人体所必需的8种氨基酸，含量占氨基酸总量的40%，其配比接近于人体氨基酸需要量的模式，还含有多种矿物质、维生素和微量元素，被称为“动物营养宝库”。国外有研究表明黄粉虫蛋白具有提高人体免疫力、抗疲劳、延缓衰老、降低血脂和促进胆固醇代谢功能。本文主要对超临界CO2萃取技术提取黄粉虫油制备黄粉虫脱脂粉的最佳工艺条件进行了研究，以得到的黄粉虫蛋白粉为原料，使用碱提法和酶提法对黄粉虫进行蛋白的提取研究，并对这两种提取工艺进行比较研究。
首先对黄粉虫进行脱脂制备粗蛋白粉，采用超临界CO2萃取技术对黄粉虫进行脱脂处理，以黄粉虫油提取率为指标，采用三因素三水平正交试验设计确定超临界CO2萃取黄粉虫油的最佳工艺条件为：萃取温度45℃，压力25MPa，时间180min。黄粉虫油提取率为95.99%。
碱法提取具有提取率高，成本低的特点，采用单因素试验和正交试验结果表明，碱提法的最佳条件为：碱液浓度为1.0%mol/L，在70℃，液固比为12:1的条件提取黄粉虫蛋白90min，在该提取工艺条件下，黄粉虫蛋白提取率可达到70.11%，纯度为91.20%。
The 21st century, mankind is faced with a more severe population growth, lack of important resources and environmental pollution pressures and challenges, and open up new resources, the path of sustainable development is the development of agricultural industrialization in China the only way. Insects as the largest on earth has not been fully utilized the resources of the treasure-house and has drawn greater attention. Insect protein resource development and industrialization, will enter the high-tech, high-yield characterized by multi-level, comprehensive utilization stage. Insect-rich protein, unsaturated fatty acids, trace elements, is a senior human nutrition source.
Tenebrio molitor protein contains 18 kinds of amino acids, and essential to the human body contains eight kinds of amino acids, amino acid content accounted for 40% of the total, the ratio closer to the model of the human amino acid requirements, but also contains a variety of minerals, vitamins and trace elements, known as the treasure house of animal nutrition. Tenebrio study abroad to improve the human immune system protein, anti-fatigue, anti-aging, reducing blood lipids and promote cholesterol metabolism. In this paper, supercritical CO2 extraction technology of oil extraction Preparation of Tenebrio molitor Tenebrio molitor powder skim the optimum conditions were studied in order to get the Tenebrio molitor protein powder as raw material, the use of base formulation and enzyme formulations for proteins of Tenebrio molitor Extraction of the extraction process of these two comparative studies.
Tenebrio first preparation of crude protein to Defatted powder, supercritical CO2 extraction technology to skim handling of Tenebrio molitor, Tenebrio molitor in the oil extraction rate as an index, a three-factor three-level orthogonal experimental design to determine Tenebrio supercritical CO2 extraction of oil The optimum conditions are as follows: extraction temperature 45 ℃, the pressure of 25MPa, the time 180min. Tenebrio oil extraction rate of 95.99%.
Alkaline extraction with high extraction rate, the characteristics of low-cost, single-factor experiments and orthogonal test results show that the best conditions for alkali formulation as follows: alkali concentration of 1.0% mol / L, at 70 ℃, liquid-solid ratio 12:1 Tenebrio protein extraction conditions for 90min, in the extraction conditions, Tenebrio molitor protein extraction rate can reach 70.11%, 91.20% purity.酶法提取具有反应条件温和和底物特异性好等特点。不同的蛋白酶提取黄粉虫蛋白有不同的作用方式，以蛋白提取率和蛋白提取液的品质为指标，比较各种蛋白酶提取黄粉虫蛋白的效果，认为复合蛋白酶和复合风味蛋白酶提取效率高和最终蛋白提取液的品质佳。为提高原料利用率和黄粉虫蛋白提取率，选择这两种酶为提取用酶。采用两种酶单独提取、分步作提取及复合作提取的结果进行比较，以蛋白提取率为指标，水解度为参考指标，采用正交实验和单因素实验，确定黄粉虫蛋白酶法提取的最佳工艺为复合蛋白酶和复合风味蛋白酶双酶复合进行酶法提取：加酶量6%复合蛋白酶与复合风味蛋白酶加量比1:3、液固比9:1、提取时间8小时，pH6. 5，温度为55℃。在该最佳工艺条件下提取黄粉虫蛋白，提取率可以达到64. 83%，纯度为92.17%。
黄粉虫体内含有各种色素和多酚类物质，其中多酚类物质在反应中易产生美拉德反应，在酶提过程中或多或少会产生苦味肽，而且为了保持其在最适pH值范围内，需加入酸、碱，产生了大量的盐，而且得到的黄粉虫蛋白提取液颜色较深，黄粉虫蛋白需精制处理。通过添加3%活性碳进行脱色、脱苦，可获得色清、味好的蛋白提取液。通过阴阳离子交换树脂可除去91.20%的盐分，最后通过喷雾干燥得到浅黄色的蛋白质粉末。
碱提法获得的蛋白产品的安全毒理学评价结果表明:碱提法所得蛋白产品是基本安全无毒的(其小鼠LD50大于120g/Kg)，因此，可以作为人和其他动物的高蛋白资源。
总之，本论文通过两种蛋白质提取方法的比较研究，初步确立了黄粉虫中蛋白质的最佳提取方法和工艺，为以后开发黄粉虫蛋白质资源提供了理论基础。
关键词：黄粉虫；蛋白质；超临界萃取；碱法；酶法；提取工艺；氨基酸
Enzymatic extraction with mild reaction conditions and substrate specificity of the characteristics of a good. Tenebrio different protease extract the role of protein have different approaches to protein extraction rate and the quality of protein extract as an index to compare the protease protein of Tenebrio molitor extract the effect of protease and that the complex compound extracted flavourzyme efficient and the final protein good quality extract. To enhance the utilization of raw materials and Tenebrio molitor protein extraction rate, selection of these two enzymes for the enzyme extraction. Using two separate enzyme extraction, step-by-step for the extraction and re-extract the results of cooperation in comparison to the rate of protein extraction as an index for the reference degree of hydrolysis, the use of single factor experiments and orthogonal experiment to determine the Tenebrio protease extracted the most good process for the composite and complex protease flavourzyme to conduct two-enzyme complex extraction: 6% the amount of enzyme protease complex and composite plus flavourzyme than 1:3, liquid-solid ratio 9:1, extraction time 8 hours, pH6. 5 temperature of 55 ℃. Optimum conditions in the extraction of Tenebrio molitor protein, extraction rate can reach 64.83%, 92.17% purity.
Tenebrio body contains a variety of pigments and polyphenols, of which polyphenols easily in response to Maillard reaction, in the course of enzyme to produce more or less bitter peptide, and in order to maintain its optimum pH value range, the need to join the acid, alkali, resulting in a lot of salt, and the protein extract of Tenebrio molitor darker color, to be refined to deal with Tenebrio molitor protein. 3% by adding activated carbon for decolorization, debittering available color clear extract protein taste good. Through the cation-anion exchange resin can remove 91.20% of the salt, and finally through the spray-drying the protein to be light yellow powder.
Alkali formulation obtained toxicology protein product safety evaluation results show that: protein products derived from the reference base is the basic security of the non-toxic (LD50 in mice is greater than its 120g/Kg), therefore, can be used as high-protein and other animals resources.
In short, this paper two methods of protein extraction studies, the initial establishment of the Tenebrio molitor protein extraction methods and the best technology, the development of Tenebrio molitor for the future of protein resources to provide a theoretical basis.
Keywords: Tenebrio molitor; protein; supercritical fluid extraction; alkali; Enzyme; extraction process; amino acid
21世纪，人类面临着更为严峻的人口剧增、资源匾乏及环境污染的压力和挑战，开辟新资源、走可持续发展道路是我国农业产业化发展的必由之路。昆虫作为地球上最大的未被充分利用的资源宝库而倍受关注。昆虫蛋白质资源的开发及其产业化，必将进入以高科技、高收益为特点的多层次、综合利用阶段。昆虫富含蛋白质、不饱和脂肪酸、微量元素，是人类的高级营养源。
黄粉虫的蛋白质中含有18种氨基酸，且包含人体所必需的8种氨基酸，含量占氨基酸总量的40%，其配比接近于人体氨基酸需要量的模式，还含有多种矿物质、维生素和微量元素，被称为“动物营养宝库”。国外有研究表明黄粉虫蛋白具有提高人体免疫力、抗疲劳、延缓衰老、降低血脂和促进胆固醇代谢功能。本文主要对超临界CO2萃取技术提取黄粉虫油制备黄粉虫脱脂粉的最佳工艺条件进行了研究，以得到的黄粉虫蛋白粉为原料，使用碱提法和酶提法对黄粉虫进行蛋白的提取研究，并对这两种提取工艺进行比较研究。
首先对黄粉虫进行脱脂制备粗蛋白粉，采用超临界CO2萃取技术对黄粉虫进行脱脂处理，以黄粉虫油提取率为指标，采用三因素三水平正交试验设计确定超临界CO2萃取黄粉虫油的最佳工艺条件为：萃取温度45℃，压力25MPa，时间180min。黄粉虫油提取率为95.99%。
碱法提取具有提取率高，成本低的特点，采用单因素试验和正交试验结果表明，碱提法的最佳条件为：碱液浓度为1.0%mol/L，在70℃，液固比为12:1的条件提取黄粉虫蛋白90min，在该提取工艺条件下，黄粉虫蛋白提取率可达到70.11%，纯度为91.20%。
酶法提取具有反应条件温和和底物特异性好等特点。不同的蛋白酶提取黄粉虫蛋白有不同的作用方式，以蛋白提取率和蛋白提取液的品质为指标，比较各种蛋白酶提取黄粉虫蛋白的效果，认为复合蛋白酶和复合风味蛋白酶提取效率高和最终蛋白提取液的品质佳。为提高原料利用率和黄粉虫蛋白提取率，选择这两种酶为提取用酶。采用两种酶单独提取、分步作提取及复合作提取的结果进行比较，以蛋白提取率为指标，水解度为参考指标，采用正交实验和单因素实验，确定黄粉虫蛋白酶法提取的最佳工艺为复合蛋白酶和复合风味蛋白酶双酶复合进行酶法提取：加酶量6%复合蛋白酶与复合风味蛋白酶加量比1:3、液固比9:1、提取时间8小时，pH6. 5，温度为55℃。在该最佳工艺条件下提取黄粉虫蛋白，提取率可以达到64. 83%，纯度为92.17%。
黄粉虫体内含有各种色素和多酚类物质，其中多酚类物质在反应中易产生美拉德反应，在酶提过程中或多或少会产生苦味肽，而且为了保持其在最适pH值范围内，需加入酸、碱，产生了大量的盐，而且得到的黄粉虫蛋白提取液颜色较深，黄粉虫蛋白需精制处理。通过添加3%活性碳进行脱色、脱苦，可获得色清、味好的蛋白提取液。通过阴阳离子交换树脂可除去91.20%的盐分，最后通过喷雾干燥得到浅黄色的蛋白质粉末。
碱提法获得的蛋白产品的安全毒理学评价结果表明:碱提法所得蛋白产品是基本安全无毒的(其小鼠LD50大于120g/Kg)，因此，可以作为人和其他动物的高蛋白资源。
总之，本论文通过两种蛋白质提取方法的比较研究，初步确立了黄粉虫中蛋白质的最佳提取方法和工艺，为以后开发黄粉虫蛋白质资源提供了理论基础。
关键词：黄粉虫；蛋白质；超临界萃取；碱法；酶法；提取工艺；氨基酸
The 21st century, mankind is faced with a more severe population growth, lack of important resources and environmental pollution pressures and challenges, and open up new resources, the path of sustainable development is the development of agricultural industrialization in China the only way. Insects as the largest on earth has not been fully utilized the resources of the treasure-house and has drawn greater attention. Insect protein resource development and industrialization, will enter the high-tech, high-yield characterized by multi-level, comprehensive utilization stage. Insect-rich protein, unsaturated fatty acids, trace elements, is a senior human nutrition source.
Tenebrio molitor protein contains 18 kinds of amino acids, and essential to the human body contains eight kinds of amino acids, amino acid content accounted for 40% of the total, the ratio closer to the model of the human amino acid requirements, but also contains a variety of minerals, vitamins and trace elements, known as the treasure house of animal nutrition. Tenebrio study abroad to improve the human immune system protein, anti-fatigue, anti-aging, reducing blood lipids and promote cholesterol metabolism. In this paper, supercritical CO2 extraction technology of oil extraction Preparation of Tenebrio molitor Tenebrio molitor powder skim the optimum conditions were studied in order to get the Tenebrio molitor protein powder as raw material, the use of base formulation and enzyme formulations for proteins of Tenebrio molitor Extraction of the extraction process of these two comparative studies.
Tenebrio first preparation of crude protein to Defatted powder, supercritical CO2 extraction technology to skim handling of Tenebrio molitor, Tenebrio molitor in the oil extraction rate as an index, a three-factor three-level orthogonal experimental design to determine Tenebrio supercritical CO2 extraction of oil The optimum conditions are as follows: extraction temperature 45 ℃, the pressure of 25MPa, the time 180min. Tenebrio oil extraction rate of 95.99%.
Alkaline extraction with high extraction rate, the characteristics of low-cost, single-factor experiments and orthogonal test results show that the best conditions for alkali formulation as follows: alkali concentration of 1.0% mol / L, at 70 ℃, liquid-solid ratio 12:1 Tenebrio protein extraction conditions for 90min, in the extraction conditions, Tenebrio molitor protein extraction rate can reach 70.11%, 91.20% purity.
Enzymatic extraction with mild reaction conditions and substrate specificity of the characteristics of a good. Tenebrio different protease extract the role of protein have different approaches to protein extraction rate and the quality of protein extract as an index to compare the protease protein of Tenebrio molitor extract the effect of protease and that the complex compound extracted flavourzyme efficient and the final protein good quality extract. To enhance the utilization of raw materials and Tenebrio molitor protein extraction rate, selection of these two enzymes for the enzyme extraction. Using two separate enzyme extraction, step-by-step for the extraction and re-extract the results of cooperation in comparison to the rate of protein extraction as an index for the reference degree of hydrolysis, the use of single factor experiments and orthogonal experiment to determine the Tenebrio protease extracted the most good process for the composite and complex protease flavourzyme to conduct two-enzyme complex extraction: 6% the amount of enzyme protease complex and composite plus flavourzyme than 1:3, liquid-solid ratio 9:1, extraction time 8 hours, pH6. 5 temperature of 55 ℃. Optimum conditions in the extraction of Tenebrio molitor protein, extraction rate can reach 64.83%, 92.17% purity.
Tenebrio body contains a variety of pigments and polyphenols, of which polyphenols easily in response to Maillard reaction, in the course of enzyme to produce more or less bitter peptide, and in order to maintain its optimum pH value range, the need to join the acid, alkali, resulting in a lot of salt, and the protein extract of Tenebrio molitor darker color, to be refined to deal with Tenebrio molitor protein. 3% by adding activated carbon for decolorization, debittering available color clear extract protein taste good. Through the cation-anion exchange resin can remove 91.20% of the salt, and finally through the spray-drying the protein to be light yellow powder.
Alkali formulation obtained toxicology protein product safety evaluation results show that: protein products derived from the reference base is the basic security of the non-toxic (LD50 in mice is greater than its 120g/Kg), therefore, can be used as high-protein and other animals resources.
In short, this paper two methods of protein extraction studies, the initial establishment of the Tenebrio molitor protein extraction methods and the best technology, the development of Tenebrio molitor for the future of protein resources to provide a theoretical basis.
Tenebrio molitor; protein; supercritical fluid extraction; alkali; Enzyme; extraction process; amino acid
The 21st century, mankind is faced with a more severe population growth, lack of important resources and environmental pollution pressures and challenges, and open up new resources, the path of sustainable development is the development of agricultural industrialization in China the only way. Insects as the largest on earth has not been fully utilized the resources of the treasure-house and has drawn greater attention. Insect protein resource development and industrialization, will enter the high-tech, high-yield characterized by multi-level, comprehensive utilization stage. Insect-rich protein, unsaturated fatty acids, trace elements, is a senior human nutrition source.
Tenebrio molitor protein contains 18 kinds of amino acids, and essential to the human body contains eight kinds of amino acids, amino acid content accounted for 40% of the total, the ratio closer to the model of the human amino acid requirements, but also contains a variety of minerals, vitamins and trace elements, known as the treasure house of animal nutrition. Tenebrio study abroad to improve the human immune system protein, anti-fatigue, anti-aging, reducing blood lipids and promote cholesterol metabolism. In this paper, supercritical CO2 extraction technology of oil extraction Preparation of Tenebrio molitor Tenebrio molitor powder skim the optimum conditions were studied in order to get the Tenebrio molitor protein powder as raw material, the use of base formulation and enzyme formulations for proteins of Tenebrio molitor Extraction of the extraction process of these two comparative studies.
Tenebrio first preparation of crude protein to Defatted powder, supercritical CO2 extraction technology to skim handling of Tenebrio molitor, Tenebrio molitor in the oil extraction rate as an index, a three-factor three-level orthogonal experimental design to determine Tenebrio supercritical CO2 extraction of oil The optimum conditions are as follows: extraction temperature 45 ℃, the pressure of 25MPa, the time 180min. Tenebrio oil extraction rate of 95.99%.
Alkaline extraction with high extraction rate, the characteristics of low-cost, single-factor experiments and orthogonal test results show that the best conditions for alkali formulation as follows: alkali concentration of 1.0% mol / L, at 70 ℃, liquid-solid ratio 12:1 Tenebrio protein extraction conditions for 90min, in the extraction conditions, Tenebrio molitor protein extraction rate can reach 70.11%, 91.20% purity.
Enzymatic extraction with mild reaction conditions and substrate specificity of the characteristics of a good. Tenebrio different protease extract the role of protein have different approaches to protein extraction rate and the quality of protein extract as an index to compare the protease protein of Tenebrio molitor extract the effect of protease and that the complex compound extracted flavourzyme efficient and the final protein good quality extract. To enhance the utilization of raw materials and Tenebrio molitor protein extraction rate, selection of these two enzymes for the enzyme extraction. Using two separate enzyme extraction, step-by-step for the extraction and re-extract the results of cooperation in comparison to the rate of protein extraction as an index for the reference degree of hydrolysis, the use of single factor experiments and orthogonal experiment to determine the Tenebrio protease extracted the most good process for the composite and complex protease flavourzyme to conduct two-enzyme complex extraction: 6% the amount of enzyme protease complex and composite plus flavourzyme than 1:3, liquid-solid ratio 9:1, extraction time 8 hours, pH6. 5 temperature of 55 ℃. Optimum conditions in the extraction of Tenebrio molitor protein, extraction rate can reach 64.83%, 92.17% purity.
Tenebrio body contains a variety of pigments and polyphenols, of which polyphenols easily in response to Maillard reaction, in the course of enzyme to produce more or less bitter peptide, and in order to maintain its optimum pH value range, the need to join the acid, alkali, resulting in a lot of salt, and the protein extract of Tenebrio molitor darker color, to be refined to deal with Tenebrio molitor protein. 3% by adding activated carbon for decolorization, debittering available color clear extract protein taste good. Through the cation-anion exchange resin can remove 91.20% of the salt, and finally through the spray-drying the protein to be light yellow powder.
Alkali formulation obtained toxicology protein product safety evaluation results show that: protein products derived from the reference base is the basic security of the non-toxic (LD50 in mice is greater than its 120g/Kg), therefore, can be used as high-protein and other animals resources.
In short, this paper two methods of protein extraction studies, the initial establishment of the Tenebrio molitor protein extraction methods and the best technology, the development of Tenebrio molitor for the future of protein resources to provide a theoretical basis.
Keywords: Tenebrio molitor; protein; supercritical fluid extraction; alkali; Enzyme; extraction process; amino acid
The 21st century, the humanity is facing the more stern population sharp increase, the resources plaque deficiency and the environmental pollution pressure and the challenge, opens the new resources, walks the sustainable development path is our country agriculture industrial production development way that must be taken.But the insect takes on the Earth biggest time not to pay attention by the full use resources treasure house.The insect protein resources development and the industrial production, will certainly to enter take the high tech, the high income as the characteristic multi-level, the comprehensive utilization stage.The insect contains the protein, the unsaturated fatty acid, the trace element richly, is humanity's high-level nutrition source
Tenebrio molitor protein contains 18 kinds of amino acids, and essential to the human body contains eight kinds of amino acids, amino acid content accounted for 40% of the total, the ratio closer to the model of the human amino acid requirements, but also contains a variety of minerals, vitamins and trace elements, known as the treasure house of animal nutrition. Tenebrio study abroad to improve the human immune system protein, anti-fatigue, anti-aging, reducing blood lipids and promote cholesterol metabolism. In this paper, supercritical CO2 extraction technology of oil extraction Preparation of Tenebrio molitor Tenebrio molitor powder skim the optimum conditions were studied in order to get the Tenebrio molitor protein powder as raw material, the use of base formulation and enzyme formulations for proteins of Tenebrio molitor Extraction of the extraction process of these two comparative studies.
Tenebrio first preparation of crude protein to Defatted powder, supercritical CO2 extraction technology to skim handling of Tenebrio molitor, Tenebrio molitor in the oil extraction rate as an index, a three-factor three-level orthogonal experimental design to determine Tenebrio supercritical CO2 extraction of oil The optimum conditions are as follows: extraction temperature 45 ℃, the pressure of 25MPa, the time 180min. Tenebrio oil extraction rate of 95.99%.
The alkaline process extraction has the extraction rate to be high, the cost low characteristic, uses the single factor experiment and the orthogonal test result indicated, the alkali formulation optimum condition is: The lye density is 1.0%mol/L, in 70℃;, the fluid solid ratio is 12:1 condition extraction flour weevil protein 90min, under this extraction technological conditions, the flour weevil protein extraction rate may achieve 70.11%, the purity is 91.20%.
Enzymatic extraction with mild reaction conditions and substrate specificity of the characteristics of a good. Tenebrio different protease extract the role of protein have different approaches to protein extraction rate and the quality of protein extract as an index to compare the protease protein of Tenebrio molitor extract the effect of protease and that the complex compound extracted flavourzyme efficient and the final protein good quality extract. To enhance the utilization of raw materials and Tenebrio molitor protein extraction rate, selection of these two enzymes for the enzyme extraction. Using two separate enzyme extraction, step-by-step for the extraction and re-extract the results of cooperation in comparison to the rate of protein extraction as an index for the reference degree of hydrolysis, the use of single factor experiments and orthogonal experiment to determine the Tenebrio protease extracted the most good process for the composite and complex protease flavourzyme to conduct two-enzyme complex extraction: 6% the amount of enzyme protease complex and composite plus flavourzyme than 1:3, liquid-solid ratio 9:1, extraction time 8 hours, pH6. 5 temperature of 55 ℃. Optimum conditions in the extraction of Tenebrio molitor protein, extraction rate can reach 64.83%, 92.17% purity.
Tenebrio body contains a variety of pigments and polyphenols, of which polyphenols easily in response to Maillard reaction, in the course of enzyme to produce more or less bitter peptide, and in order to maintain its optimum pH value range, the need to join the acid, alkali, resulting in a lot of salt, and the protein extract of Tenebrio molitor darker color, to be refined to deal with Tenebrio molitor protein. 3% by adding activated carbon for decolorization, debittering available color clear extract protein taste good. Through the cation-anion exchange resin can remove 91.20% of the salt, and finally through the spray-drying the protein to be light yellow powder.
Alkali formulation obtained toxicology protein product safety evaluation results show that: protein products derived from the reference base is the basic security of the non-toxic (LD50 in mice is greater than its 120g/Kg), therefore, can be used as high-protein and other animals resources.
In short, this paper two methods of protein extraction studies, the initial establishment of the Tenebrio molitor protein extraction methods and the best technology, the development of Tenebrio molitor for the future of protein resources to provide a theoretical basis.
Tenebrio molitor; protein; supercritical fluid extraction; alkali; Enzyme; extraction process; amino acid
