火龙果茎凝胶汁、多糖的初步研究
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摘要
火龙果茎中含有植物性多糖、多种矿物质元素、维生素E和甾醇等物质,能调节人体机能,提高人体免疫力,美容养颜,开发前景广阔,潜在价值高,近年来在国内外受到广泛关注,但是有关火龙果茎凝胶汁的制备和多糖的提取以及火龙果茎凝胶汁和多糖生物功效方面的研究尚未见报道。
本实验以新鲜火龙果茎为材料,首先研究火龙果茎凝胶汁的提取工艺,并针对降粘和脱色进行优化。结果表明,火龙果茎凝胶汁提取效果最佳的料液比为1:3,降粘的最佳条件是加入0.05%的果胶酶,45℃水浴浸提,浸提时间为60min;脱色的最佳条件是加入2%活性炭,60℃水浴脱色,脱色时间为30min。
其次,对火龙果茎凝胶汁进行成分分析和生物功效的检测。结果表明,火龙果茎凝胶汁中总膳食纤维(TDF-Total Dietary Fiber)含量为728.4g/kg;不溶性膳食纤维(IDF-InsolubleDietary Fiber)含量为462.9g/kg:可溶性膳食纤维(SDF-Soluble Dietary Fiber)含量为193.3g/kg:超氧化物歧化酶(SOD-Super Oxide Dlsmutase)活力为4.416U/mL;脂肪酶活力为2.16U/g;蛋白酶活力为6.46U/mL;多糖含量为130.48ug/mL;蛋白含量为1619.17ug/mL。分别采用DPPH法、邻苯三酚法和水杨酸法测定火龙果茎凝胶汁清除自由基的功效,发现其对DPPH总自由基清除率为48.55%,对超氧阴离子自由基的清除率为13.96%,对羟自由基的清除率为59.30%;以对酪氨酸酶活性的抑制法来测定其美白功效,结果显示其对酪氨酸酶的活性抑制率为21.41%;经过人体保湿实验发现其含水率高于空白对照组,失水率低于空白对照组;通过皮肤成像系统对其进行嫩肤功效的检测发现涂抹火龙果茎凝胶汁后皮肤粗糙度有所降低;进行人体抗敏实验,通过皮肤涂抹火龙果茎凝胶汁前后对致敏剂的反应,发现其能有效舒缓皮肤的过敏症状。
最后,通过正交实验来纵向和横向比较几种火龙果茎多糖的提取方法。最终确定通过微波提取的方法对火龙果茎中的植物性多糖进行提取,提取条件是微波功率200W,微波时间10min,微波温度80℃。经过冷处理法醇沉提出粗多糖(SPP),通过苯酚-硫酸法检测其总糖含量为27.85%,考马斯亮蓝染色法检测其蛋白质含量为4.61%。通过Sevage法去除粗多糖SPP中蛋白质杂质,并通过正交试验获得最佳条件:试剂配比(氯仿:正丁醇)=3:1,Sevage试剂用量(试剂/样品)=1/4,Sevage试剂取用次数3次。再经透析后冷冻干燥成粉,得到纯化后多糖。纯化后所得多糖(SPPS)为浅白色粉末,糖含量比SPP提高13.03%。
将纯化后的多糖SPPS通过高效液相色谱检测其纯度,出现单一对称的峰,证实SPPS是单一火龙果茎多糖;进行元素组成分析,碳氮比由31.67提高到101.52,证明SPP纯化为SPPS后游离蛋白质和小分子杂质基本去除;经红外图谱(IR)结构鉴定,SPPS为含有乙酰化β-D-吡喃甘露聚糖的混合物;经基质辅助激光解吸离子化飞行时间质谱(MALDI-TOF-MS)分子量测定,其范围为3900～4300。分别以自由基清除率、抑制酪氨酸酶活性和体内水分含量与散失率为指标来检测纯化后多糖SPPS在抗氧化、美白和保湿方面的生物学功效,发现其在保湿方面效果显著。
本论文有关火龙果茎凝胶汁的制备和多糖的提取以及火龙果茎凝胶汁和多糖的生物功效的初步研究为人们深入了解和应用火龙果茎提供了依据。
The stem of pitaya contains a variety of plant polysaccharides and minerals,vitamin E and sterols,which can adjust the various functions of the human body,improve human immunity.It has gerat prospects and potential high value.In recent years,it received wide attention from the country and abroad.But the research about the gel preparation and extraction of polysaccharide from the stem of pitaya,the biological efficacy of the gel and the polysaccharide have not been reported.
Fresh stem of pitaya was the material for experiment in this paper.First,through a series of orthogonal experiment to determine the optimal extraction method to get the gel from pitaya stem and optimize the process of decolourization and clarification of extracting the gelatin.The optimal technological conditions were as follows:the ratio of solid to liquid was 1:3,extraction time 60min,extraction temperature 45℃,0.05%pectinase.And the best decoloration conditions were:2%of activated carbon,decoloration temperature 60℃,decoloration time 30min.
And then,we did some analysis and detection of biological efficacy of the gel from the stem of pitaya.The results showed that in the gel the content of total dietary fiber(TDF-Total Dietary Fiber) was 728.4g/kg;the content of insoluble dietary fiber(IDF-Insoluble Dietary Fiber) was 462.9g/kg;the content of soluble dietary fiber(SDF-Soluble Dietary Fiber) was 193.3g/kg;the activity of super oxide dismutase(SOD-Super Oxide Dismutase) dynamic was 4.416U/mL;the activity of lipase was 2.16U/g;the activity of proteinase was 6.46U/mL;the content of polysaccharide in the gel was 130.48ug/mL;the content ofprotain in the gel was 1619.17ug/mL. DPPH method,pyrogallol and salicylic acid method were used to detect the efficacy of free radical scavenging of the gel from the stem of pitaya.The results showed that a total of DPPH free radical scavenging rate was 48.55%,the superoxide anion radical scavenging rate was 13.96%and the hydroxyl radical scavenging rate was 59.30%.The tyrosinase activity inhibition method was done to determine the effectiveness of the whitening.The result showed the activity of tyrosinase inhibition rate was 21.41%.Through the human body moisture,it was found that its moisture content was higher than the control group,the water loss rate was lower than the control group;imaging system through the skin of young women testing the efficacy of stem pitaya smearing was found that after the gel decreased,skin was roughness.Anti-allergy experiment was done.We found that it could effectively alleviate the situation of skin allergy by comparing the responses to allergens of the skin which used the gel before and after.
At last,polysaccharide was extracted from the stem of pitaya by hotwater,enzyme- assisted and microwave-assisted method.Finally the microwave-assisted extraction was proved to be the best one by general evaluation.The results showed that the main factors affecting the yield of polysaccharides in the process of microwave-assisted extraction were sequenced as follows:time＞microwave power＞temperature,the optimal technological conditions were as follows: extraction time 10min,extraction temperature 80℃and the microwave power 200W.The rough Stem of Pitaya Polysaccharide(SPP) was extracted from the fresh stem of pitaya.The content of total polysaccharide in SPP was 27.85%determined by phenol-vitriol method using D-glucose as standard.The content of protein in SPP was 4.61%using BSA as standard.Sevage-protocols were applied to deprotein of the SPP and the optimum condition was obtained by the experiments of orthogonal design.Then the first step purified stem of pitaya polysaccharides (SPPS) was obtained after dialyzation and freezing dryness.Using the optimal technological conditions,the yield of SPP was 1.42%.After purifying,the yield of SPPS was 0.74%.
SPPS was analysised by IR,MALDI-TOF-MS and dement analysis technique.It was approved to be a kind of polysaccharide mixture,and the molecuLar weight was between 3900 and 4300.Compared with other biological efficacy,it had great efficacy in moisture retention.
This paper will provide a technical reference for the future applications of the stems of pitaya.
本实验以新鲜火龙果茎为材料,首先研究火龙果茎凝胶汁的提取工艺,并针对降粘和脱色进行优化。结果表明,火龙果茎凝胶汁提取效果最佳的料液比为1:3,降粘的最佳条件是加入0.05%的果胶酶,45℃水浴浸提,浸提时间为60min;脱色的最佳条件是加入2%活性炭,60℃水浴脱色,脱色时间为30min。
其次,对火龙果茎凝胶汁进行成分分析和生物功效的检测。结果表明,火龙果茎凝胶汁中总膳食纤维(TDF-Total Dietary Fiber)含量为728.4g/kg;不溶性膳食纤维(IDF-InsolubleDietary Fiber)含量为462.9g/kg:可溶性膳食纤维(SDF-Soluble Dietary Fiber)含量为193.3g/kg:超氧化物歧化酶(SOD-Super Oxide Dlsmutase)活力为4.416U/mL;脂肪酶活力为2.16U/g;蛋白酶活力为6.46U/mL;多糖含量为130.48ug/mL;蛋白含量为1619.17ug/mL。分别采用DPPH法、邻苯三酚法和水杨酸法测定火龙果茎凝胶汁清除自由基的功效,发现其对DPPH总自由基清除率为48.55%,对超氧阴离子自由基的清除率为13.96%,对羟自由基的清除率为59.30%;以对酪氨酸酶活性的抑制法来测定其美白功效,结果显示其对酪氨酸酶的活性抑制率为21.41%;经过人体保湿实验发现其含水率高于空白对照组,失水率低于空白对照组;通过皮肤成像系统对其进行嫩肤功效的检测发现涂抹火龙果茎凝胶汁后皮肤粗糙度有所降低;进行人体抗敏实验,通过皮肤涂抹火龙果茎凝胶汁前后对致敏剂的反应,发现其能有效舒缓皮肤的过敏症状。
最后,通过正交实验来纵向和横向比较几种火龙果茎多糖的提取方法。最终确定通过微波提取的方法对火龙果茎中的植物性多糖进行提取,提取条件是微波功率200W,微波时间10min,微波温度80℃。经过冷处理法醇沉提出粗多糖(SPP),通过苯酚-硫酸法检测其总糖含量为27.85%,考马斯亮蓝染色法检测其蛋白质含量为4.61%。通过Sevage法去除粗多糖SPP中蛋白质杂质,并通过正交试验获得最佳条件:试剂配比(氯仿:正丁醇)=3:1,Sevage试剂用量(试剂/样品)=1/4,Sevage试剂取用次数3次。再经透析后冷冻干燥成粉,得到纯化后多糖。纯化后所得多糖(SPPS)为浅白色粉末,糖含量比SPP提高13.03%。
将纯化后的多糖SPPS通过高效液相色谱检测其纯度,出现单一对称的峰,证实SPPS是单一火龙果茎多糖;进行元素组成分析,碳氮比由31.67提高到101.52,证明SPP纯化为SPPS后游离蛋白质和小分子杂质基本去除;经红外图谱(IR)结构鉴定,SPPS为含有乙酰化β-D-吡喃甘露聚糖的混合物;经基质辅助激光解吸离子化飞行时间质谱(MALDI-TOF-MS)分子量测定,其范围为3900～4300。分别以自由基清除率、抑制酪氨酸酶活性和体内水分含量与散失率为指标来检测纯化后多糖SPPS在抗氧化、美白和保湿方面的生物学功效,发现其在保湿方面效果显著。
本论文有关火龙果茎凝胶汁的制备和多糖的提取以及火龙果茎凝胶汁和多糖的生物功效的初步研究为人们深入了解和应用火龙果茎提供了依据。
The stem of pitaya contains a variety of plant polysaccharides and minerals,vitamin E and sterols,which can adjust the various functions of the human body,improve human immunity.It has gerat prospects and potential high value.In recent years,it received wide attention from the country and abroad.But the research about the gel preparation and extraction of polysaccharide from the stem of pitaya,the biological efficacy of the gel and the polysaccharide have not been reported.
Fresh stem of pitaya was the material for experiment in this paper.First,through a series of orthogonal experiment to determine the optimal extraction method to get the gel from pitaya stem and optimize the process of decolourization and clarification of extracting the gelatin.The optimal technological conditions were as follows:the ratio of solid to liquid was 1:3,extraction time 60min,extraction temperature 45℃,0.05%pectinase.And the best decoloration conditions were:2%of activated carbon,decoloration temperature 60℃,decoloration time 30min.
And then,we did some analysis and detection of biological efficacy of the gel from the stem of pitaya.The results showed that in the gel the content of total dietary fiber(TDF-Total Dietary Fiber) was 728.4g/kg;the content of insoluble dietary fiber(IDF-Insoluble Dietary Fiber) was 462.9g/kg;the content of soluble dietary fiber(SDF-Soluble Dietary Fiber) was 193.3g/kg;the activity of super oxide dismutase(SOD-Super Oxide Dismutase) dynamic was 4.416U/mL;the activity of lipase was 2.16U/g;the activity of proteinase was 6.46U/mL;the content of polysaccharide in the gel was 130.48ug/mL;the content ofprotain in the gel was 1619.17ug/mL. DPPH method,pyrogallol and salicylic acid method were used to detect the efficacy of free radical scavenging of the gel from the stem of pitaya.The results showed that a total of DPPH free radical scavenging rate was 48.55%,the superoxide anion radical scavenging rate was 13.96%and the hydroxyl radical scavenging rate was 59.30%.The tyrosinase activity inhibition method was done to determine the effectiveness of the whitening.The result showed the activity of tyrosinase inhibition rate was 21.41%.Through the human body moisture,it was found that its moisture content was higher than the control group,the water loss rate was lower than the control group;imaging system through the skin of young women testing the efficacy of stem pitaya smearing was found that after the gel decreased,skin was roughness.Anti-allergy experiment was done.We found that it could effectively alleviate the situation of skin allergy by comparing the responses to allergens of the skin which used the gel before and after.
At last,polysaccharide was extracted from the stem of pitaya by hotwater,enzyme- assisted and microwave-assisted method.Finally the microwave-assisted extraction was proved to be the best one by general evaluation.The results showed that the main factors affecting the yield of polysaccharides in the process of microwave-assisted extraction were sequenced as follows:time＞microwave power＞temperature,the optimal technological conditions were as follows: extraction time 10min,extraction temperature 80℃and the microwave power 200W.The rough Stem of Pitaya Polysaccharide(SPP) was extracted from the fresh stem of pitaya.The content of total polysaccharide in SPP was 27.85%determined by phenol-vitriol method using D-glucose as standard.The content of protein in SPP was 4.61%using BSA as standard.Sevage-protocols were applied to deprotein of the SPP and the optimum condition was obtained by the experiments of orthogonal design.Then the first step purified stem of pitaya polysaccharides (SPPS) was obtained after dialyzation and freezing dryness.Using the optimal technological conditions,the yield of SPP was 1.42%.After purifying,the yield of SPPS was 0.74%.
SPPS was analysised by IR,MALDI-TOF-MS and dement analysis technique.It was approved to be a kind of polysaccharide mixture,and the molecuLar weight was between 3900 and 4300.Compared with other biological efficacy,it had great efficacy in moisture retention.
This paper will provide a technical reference for the future applications of the stems of pitaya.
引文
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