大豆溶血磷脂的制备及其生物安全性分析
|
摘要
大豆溶血磷脂是原有大豆磷脂结构中失去一个脂肪酸基团的改性磷脂的统称,较大豆磷脂增强了亲水性和抗氧化性,被广泛地用于医药、食品和畜牧业。采用磷脂酶A1对大豆磷脂进行改性,以水解率和红外光谱检测其改性程度,以溶血试验和细胞毒性试验检测其生物安全性。结果表明:采用磷脂酶A1在60℃、40%大豆磷脂-水的体系中酶解大豆磷脂所制备的大豆溶血磷脂,纯化后为棕色、块状物质且无明显豆腥味;其水解率为101. 6%;大豆溶血磷脂中双键和酰胺键的吸收峰明显低于大豆磷脂;大豆溶血磷脂和大豆磷脂均有溶血现象,且二者在200μg/mL以下溶血率均低于5%,视为生物安全;大豆溶血磷脂在20μg/mL、大豆磷脂在50μg/mL以下时,Caco-2细胞的存活率均高于80%。综上,采用磷脂酶A1制备的大豆溶血磷脂,在一定质量浓度下使用,具备生物安全性。
Soybean lysophospholipids are modified phosphatides of losing a fatty acid group in the original structure of soybean phospholipids,which are more hydrophilic and antioxidative than soybean phospholipids,and are widely used in medicine,food and animal husbandry.Phospholipase A1 was used to modify the soybean phospholipids,and the degree of modification was detected by the hydrolysis rate and infrared spectrum,and the biological safety was detected by hemolytic test and cytotoxicity test.The results showed that the soybean lysophospholipids were prepared from the 40% soybean phospholipids in the water catalyzed by phospholipase A1 at 60 ℃.The purified soybean lysophospholipids were block-brown material without obvious soybean smell.The hydrolysis rate of soybean lysophospholipids was 101.6%.The results of infrared spectra showed that the double bond and amide bond were significantly decreased in soybean lysophospholipids.Meanwhile,both of soybean phospholipids and soybean lysophospholipids had hemolytic phenomena,and their hemolysis rates at 200 μg/mL were less than 5%,which could be considered as biosafety.The cell survival rates were all higher than 80% when soybean phospholipids were below 50 μg/mL and soybean lysophospholipids were below 20 μg/mL.In summary,the prepared soybean lysophospholipids were biosafe at certain mass concentration.
Soybean lysophospholipids are modified phosphatides of losing a fatty acid group in the original structure of soybean phospholipids,which are more hydrophilic and antioxidative than soybean phospholipids,and are widely used in medicine,food and animal husbandry.Phospholipase A1 was used to modify the soybean phospholipids,and the degree of modification was detected by the hydrolysis rate and infrared spectrum,and the biological safety was detected by hemolytic test and cytotoxicity test.The results showed that the soybean lysophospholipids were prepared from the 40% soybean phospholipids in the water catalyzed by phospholipase A1 at 60 ℃.The purified soybean lysophospholipids were block-brown material without obvious soybean smell.The hydrolysis rate of soybean lysophospholipids was 101.6%.The results of infrared spectra showed that the double bond and amide bond were significantly decreased in soybean lysophospholipids.Meanwhile,both of soybean phospholipids and soybean lysophospholipids had hemolytic phenomena,and their hemolysis rates at 200 μg/mL were less than 5%,which could be considered as biosafety.The cell survival rates were all higher than 80% when soybean phospholipids were below 50 μg/mL and soybean lysophospholipids were below 20 μg/mL.In summary,the prepared soybean lysophospholipids were biosafe at certain mass concentration.
引文
[1]孙浩洋,张雁,林美金,等.磷脂酶A1催化水解大豆磷脂制备溶血卵磷脂的研究[J].轻工科技,2017,33(7):13-15.
[2]侯丹熹.大豆磷脂在畜禽生产中的应用研究进展[J].饲料与畜牧,2017(11):24-27.
[3]LU S F,ZHAO N,ZHAO A Y,et al. Effect of soybean phospholipid supplementation in formulated microdiets and live food on foregut and liver histological changes of pelteobagrus fulvidraco larvae[J]. Aquaculture,2008,278:119-127.
[4]MA J Y,CHEN D Y,LI Y,et al. Zinc phthalocyaninesoybean phospholipid complex based drug carrier for switchable photoacoustic/fluorescence image,multiphase photothermal/photodynamic treatment and synergetic therapy[J]. J Control Release,2018,284:1-14.
[5]黄仪友,钟毅.大豆磷脂中磷脂酰胆碱稳定性研究[J].食品安全质量检测学报,2017,8(12):4827-4831.
[6]范勋涛.磷脂酶A1水解大豆磷脂及其产物溶血磷脂的应用研究[D].黑龙江大庆:黑龙江八一农垦大学,2009:34-36.
[7]李招,谷克仁,张海臣,等.溶血磷脂的制备、分离、检测及应用研究进展[J].中国油脂,2018,43(6):132-136,143.
[8]TONG C,LIU L,WATERS D L E,et al. The contribution of lysophospholipids to pasting and thermal properties of nonwaxy rice starch[J]. Carbohydr Polym,2015,133:187-193.
[9]KIM M J,HOSSEINDOUST A R,CHOI Y H,et al. An evaluation of metabolizable energy content of main feed ingredients for growing pigs when adding dietary lysophospholipids[J]. Livest Sci,2018,210:99-103.
[10]MELERO A,CHIARUTTINI N,KARASHIMA T,et al.Lysophospholipids facilitate COPII vesicle formation[J].Curr Biol,2018,28(12):1950-1958.
[11]宋丹军,潘家琪,李鹏旭,等.溶血磷脂酰胆碱在肝脏疾病中的研究进展[J].中国药理学通报,2014,30(12):1642-1646.
[12]毕徐堃.溶血磷脂酰胆碱与分子伴侣GRP78在心脏缺血再灌注损伤中的作用及机制的研究[D].杭州:浙江大学,2017.
[13]王恒飞,李鹏,付淑珍.低浓度溶血磷脂酸诱导海马神经元凋亡及其对c-jun磷酸化的影响[J].广东医学,2016,37(14):2091-2094.
[14]鄢黎妮,张兆辉,刘伦.溶血磷脂酸诱导原代培养的小鼠小胶质细胞的凋亡[J].卒中与神经疾病,2008,15(2):95-97,102.
[15] HIROYUKI H,RYOJI S,HEN-SIK K. Process for manufacturing vegetable lysolecithins:US5955327A[P].1999-09-21.
[16]陈斌斌.水相体系中磷脂酶A1改性大豆磷脂研究[D].郑州:河南工业大学,2010.
[17]中华人民共和国国家质量监督检验检疫总局中国国家标准化管理委员会.医用输液、输血、注射器具检验方法第2部分:生物学试验方法:GB/T 14233. 2—2005[S].北京:中国标准出版社,2005.
[18]蔡伟惠,姚成丽,居敏俐,等.丙泊酚纳米注射液中溶血磷脂的含量测定及溶血性评价[J].中国药学杂志,2013,48(3):212-215.
[2]侯丹熹.大豆磷脂在畜禽生产中的应用研究进展[J].饲料与畜牧,2017(11):24-27.
[3]LU S F,ZHAO N,ZHAO A Y,et al. Effect of soybean phospholipid supplementation in formulated microdiets and live food on foregut and liver histological changes of pelteobagrus fulvidraco larvae[J]. Aquaculture,2008,278:119-127.
[4]MA J Y,CHEN D Y,LI Y,et al. Zinc phthalocyaninesoybean phospholipid complex based drug carrier for switchable photoacoustic/fluorescence image,multiphase photothermal/photodynamic treatment and synergetic therapy[J]. J Control Release,2018,284:1-14.
[5]黄仪友,钟毅.大豆磷脂中磷脂酰胆碱稳定性研究[J].食品安全质量检测学报,2017,8(12):4827-4831.
[6]范勋涛.磷脂酶A1水解大豆磷脂及其产物溶血磷脂的应用研究[D].黑龙江大庆:黑龙江八一农垦大学,2009:34-36.
[7]李招,谷克仁,张海臣,等.溶血磷脂的制备、分离、检测及应用研究进展[J].中国油脂,2018,43(6):132-136,143.
[8]TONG C,LIU L,WATERS D L E,et al. The contribution of lysophospholipids to pasting and thermal properties of nonwaxy rice starch[J]. Carbohydr Polym,2015,133:187-193.
[9]KIM M J,HOSSEINDOUST A R,CHOI Y H,et al. An evaluation of metabolizable energy content of main feed ingredients for growing pigs when adding dietary lysophospholipids[J]. Livest Sci,2018,210:99-103.
[10]MELERO A,CHIARUTTINI N,KARASHIMA T,et al.Lysophospholipids facilitate COPII vesicle formation[J].Curr Biol,2018,28(12):1950-1958.
[11]宋丹军,潘家琪,李鹏旭,等.溶血磷脂酰胆碱在肝脏疾病中的研究进展[J].中国药理学通报,2014,30(12):1642-1646.
[12]毕徐堃.溶血磷脂酰胆碱与分子伴侣GRP78在心脏缺血再灌注损伤中的作用及机制的研究[D].杭州:浙江大学,2017.
[13]王恒飞,李鹏,付淑珍.低浓度溶血磷脂酸诱导海马神经元凋亡及其对c-jun磷酸化的影响[J].广东医学,2016,37(14):2091-2094.
[14]鄢黎妮,张兆辉,刘伦.溶血磷脂酸诱导原代培养的小鼠小胶质细胞的凋亡[J].卒中与神经疾病,2008,15(2):95-97,102.
[15] HIROYUKI H,RYOJI S,HEN-SIK K. Process for manufacturing vegetable lysolecithins:US5955327A[P].1999-09-21.
[16]陈斌斌.水相体系中磷脂酶A1改性大豆磷脂研究[D].郑州:河南工业大学,2010.
[17]中华人民共和国国家质量监督检验检疫总局中国国家标准化管理委员会.医用输液、输血、注射器具检验方法第2部分:生物学试验方法:GB/T 14233. 2—2005[S].北京:中国标准出版社,2005.
[18]蔡伟惠,姚成丽,居敏俐,等.丙泊酚纳米注射液中溶血磷脂的含量测定及溶血性评价[J].中国药学杂志,2013,48(3):212-215.