Determination of 16 Amino Acids in Rice Peptides by PITCPre-column Derivatization High Performance Liquid Chromatography

HUANG Ying; HUANG Jing; YANG Xiao; CHEN Xi; HU Wuyao

doi:10.13386/j.issn1002-0306.2023020133

Volume 44 Issue 24

Dec. 2023

Turn off MathJax

Article Contents

Abstract

References

Science and Technology of Food Industry > 2023 > 44(24): 279-285. > DOI: 10.13386/j.issn1002-0306.2023020133

HUANG Ying, HUANG Jing, YANG Xiao, et al. Determination of 16 Amino Acids in Rice Peptides by PITCPre-column Derivatization High Performance Liquid Chromatography[J]. Science and Technology of Food Industry, 2023, 44(24): 279−285. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2023020133.

Citation:

PDF (1391 KB)

Determination of 16 Amino Acids in Rice Peptides by PITCPre-column Derivatization High Performance Liquid Chromatography

Anti-aging Nutrition & Health Research and Development Center of China Health Promotion Foundation,Wuhan 430000, China

More Information

Received Date: February 13, 2023
Available Online: October 15, 2023

Graphical Abstract

Abstract

Abstract

To establish a methodology for detecting the amino acid content in rice peptides, the research employed the pre-column derivative phenyl isothiocyanate (PITC) for realizing simultaneous, rapid, and effective analysis of 16 amino acids, including aspartate (Asp), glutamic acid (Glu), serine (Ser), glycine (Gly), histidine (His), arginine (Arg), threonine (Thr), alanine (Ala), proline (Pro), tyrosine (Tyr), valine (Val), methionine (Met), isoleucine (Ile), leucine (Leu), phenylalanine (Phe), and lysine (Lys). The analysis was carried out using an Athena AAA chromatographic column as the analysis column, with a flow rate of 1.0 mL/min, column temperature of 35 ℃, and UV detector wavelength of 254 nm. A gradient elution was performed with a mobile phase consisting of methanol:acetonitrile:water=20:60:20 and 50 mmol/L sodium acetate (pH6.5). The results revealed a strong linear relationship among the 16 amino acids within the range of 0.125 to 2.5 μmol/mL, with a coefficient of determination R² ≥0.9999 for 14 amino acids. The remaining 2 amino acids exhibited coefficient of determination R² were 0.9998 and 0.9994, respectively. The detection limit ranged from 0.003% to 0.018%, while the quantitative limit ranged from 0.010% to 0.059%. The spiked recovery rate ranged from 82.21% to103.59%, and the relative standard deviation (RSD) was less than 5% for all measurements. This method effectively separates the 16 amino acids in rice peptides, with glutamic acid (Glu) having the highest content ranging from 13.11%±0.39% to 21.38%±0.48%. The amino acids with the lowest content are histidine (His) or methionine (Met), ranging from 1.59%±0.05% to 2.27%±0.03% or 1.26%±0.05% to 2.32%±0.06%, respectively. The experimental method employed in this study demonstrates high accuracy, precision, sensitivity and recovery rate, thereby, indicating its suitability for analyzing the amino acid content in rice peptides.
- rice peptides,
- amino acids,
- phenyl isothiocyanate,
- pre-column derivatization,
- high-performance liquid chromatography

FullText(HTML)

References (30)

References

[1]	尹曼, 魏颖, 马勇, 等. 胶原肽和大米肽对黑色素生成的影响[J]. 食品科技, 2017, 42(10):244−247. [YIN M, WEI Y, MA Y, et al. The effects of collagen peptides and rice peptides on melanin production[J]. Food Technology, 2017, 42 (10):244−247.] YIN M, WEI Y, MA Y, et al. The effects of collagen peptides and rice peptides on melanin production[J]. Food Technology, 2017, 42 （10）: 244−247.
[2]	程云辉, 毛田米, 黄璐, 等. 大米肽中压离子交换色谱分离及其免疫活性评价[J]. 食品与机械,2018,34(3):151−155. [CHENG Y H, MAO T M, HUANG L, et al. Separation and immunological activity evaluation of rice peptides by Medium Pressure Ion Exchange Chromatography[J]. Food and Machinery,2018,34(3):151−155.] CHENG Y H, MAO T M, HUANG L, et al. Separation and immunological activity evaluation of rice peptides by Medium Pressure Ion Exchange Chromatography[J]. Food and Machinery, 2018, 34（3）: 151−155.
[3]	秦芸, 石沛霖, 刘维维, 等. 富硒大米肽体内抗氧化活性研究[J]. 食品工业科技,2017,38(17):305−309. [QIN Y, SHI P L, LIU W W, et al. Antioxidant activity of selenium-rich rice peptides in vivo[J]. Science and Technology of Food Industry,2017,38(17):305−309.] QIN Y, SHI P L, LIU W W, et al. Antioxidant activity of selenium-rich rice peptides in vivo[J]. Science and Technology of Food Industry, 2017, 38（17）: 305−309.
[4]	YANG X, WANG L, ZHANG F, et al. Effects of multi-mode S-type ultrasound pretreatment on the preparation of ACE inhibitory peptide from rice protein[J]. Food Chemistry,2020,331:1−10.
[5]	史云丽, 刘芳, 潘曼, 等. 大米生物活性肽研究进展[J]. 食品与机械,2009,25(2):167−171. [SHI Y L, LIU F, PAN M, et al. Research progress on rice bioactive peptides[J]. Food and Machinery,2009,25(2):167−171.] SHI Y L, LIU F, PAN M, et al. Research progress on rice bioactive peptides[J]. Food and Machinery, 2009, 25（2）: 167−171.
[6]	KUN L L, RUN F D, FU S C. Stability of the antioxidant peptide SeMet-Pro-Ser identified from selenized brown rice protein hydrolysates[J]. Food Chemistry,2020,319:1−7.
[7]	王申, 林利美, 周佳, 等. 大米肽的绝对分子质量与氨基酸组成集体外活性[J]. 食品科学,2013,34(13):19−23. [WANG S, LIN L M, ZHOU J, et al. Absolute molecular mass and amino acid composition of rice peptides[J]. Food Science,2013,34(13):19−23.] WANG S, LIN L M, ZHOU J, et al. Absolute molecular mass and amino acid composition of rice peptides[J]. Food Science, 2013, 34（13）: 19−23.
[8]	李志坚, 李高阳, 张群, 等. PITC柱前衍生-液相色谱法测定西瓜中L-瓜氨酸含量[J]. 食品工业科技,2013,34(20):53−55. [LI Z J, LI G Y, ZHANG Q, et al. Determination of L-citrulline in watermelon by PITC pre-column derivatization and liquid chromatography[J]. Science and Technology of Food Industry,2013,34(20):53−55.] LI Z J, LI G Y, ZHANG Q, et al. Determination of L-citrulline in watermelon by PITC pre-column derivatization and liquid chromatography[J]. Science and Technology of Food Industry, 2013, 34（20）: 53−55.
[9]	邵瑞琪, 陈双玲. 氨基酸分析仪在医学中的应用[J]. 分析仪器,2019,223(2):133−137. [SHAO R Q, CHEN S L. Application of amino acid analyzer in medicine[J]. Analytical Instrument,2019,223(2):133−137.] SHAO R Q, CHEN S L. Application of amino acid analyzer in medicine[J]. Analytical Instrument, 2019, 223（2）: 133−137.
[10]	国家卫生和计划生育委员会, 国家食品药品监督管理总局. GB 5009.124-2016 食品安全国家标准食品中氨基酸的测定[S]. 北京:中国标准出版社, 2017. [National Health and Family Planning Commission, State Food and Drug Administration. GB 5009.124-2016 National food safety standard-de termination of amino acids in food[S]. Beijing:China Standards Press, 2017.] National Health and Family Planning Commission, State Food and Drug Administration. GB 5009.124-2016 National food safety standard-de termination of amino acids in food[S]. Beijing: China Standards Press, 2017.
[11]	SHEN F, NIU X Y, YANG D T, et al. Determination of amino acids in chinese rice wine by fourier transform near-infrared spectroscopy[J]. Journal of Agriculture and Food Chemistry,2010,58(17):9809−9816. doi: 10.1021/jf1017912
[12]	ADHIKARI B, DHUNGANA S K, ALI M W, et al. Antioxidant activities, polyphenol, flavonoid, and amino acid contents in peanut shell[J]. Journal Saudi Society Agriculture Science,2019,18(4):437−442.
[13]	ZENG X Y, WANG M F, ZHU B K, et al. Simultaneous analysis of AY and amino acids in corn oligopeptides by HPLC-fluorescencedetector with OPA/FMOC-Cl pre-column derivatization[J]. International Journal of Food Agriculture and Environment,2013,11(1):86−90.
[14]	MANDRIOLI R, MERCOLINI L, RAGGI M A. Recent trends in the analysis of amino acids in fruits and derived food stuffs[J]. Analytical and Bioanalytical Chemistry,2013,405(25):7941−7956. doi: 10.1007/s00216-013-7025-8
[15]	彭真汾, 王威, 叶清华, 等. 高效液相色谱-串联质谱法定量分析橄榄果实氨基酸组分[J]. 食品科学,2018,39(24):231−238. [PENG Z F, WANG W, YE Q H, et al. Determination of amino acids in olive fruit by HPLC tandem mass spectrometry[J]. Food Science,2018,39(24):231−238.] PENG Z F, WANG W, YE Q H, et al. Determination of amino acids in olive fruit by HPLC tandem mass spectrometry[J]. Food Science, 2018, 39（24）: 231−238.
[16]	张雪艳, 郭雷, 周长明, 等. 柱前衍生高效液相色谱法测定注射用骨肽中15种氨基酸含量[J]. 中国药业,2022,31(1):74−78. [ZHANG X Y, GUO L, ZHOU C M, et al. Determination of 15 amino acids in bone peptides for injection by high performance liquid chromatography with pre-column derivations[J]. Chinese Pharmaceutical Industry,2022,31(1):74−78.] ZHANG X Y, GUO L, ZHOU C M, et al. Determination of 15 amino acids in bone peptides for injection by high performance liquid chromatography with pre-column derivations[J]. Chinese Pharmaceutical Industry, 2022, 31（1）: 74−78.
[17]	郭晓晗, 程显隆, 柳温曦, 等. 柱前衍生化-HPLC 法同时测定不同商品规格鹿茸片中15种氨基酸的含量[J]. 药物分析杂志,2020,40(10):1780−1789. [GUO X H, CHEN X L, LIU W X, et al. Simultaneous determination of 15 amino acids in different commercial antler tablets by precolumn derivatization and HPLC[J]. Journal of Pharmaceutical Analysis,2020,40(10):1780−1789.] GUO X H, CHEN X L, LIU W X, et al. Simultaneous determination of 15 amino acids in different commercial antler tablets by precolumn derivatization and HPLC[J]. Journal of Pharmaceutical Analysis, 2020, 40（10）: 1780−1789.
[18]	田洪斌, 赵可意, 孙佳玲. 氨基酸分析法测定复方骨肽注射液中多肽含量[J]. 中国药事,2011,25(8):816−820. [TIAN H B, ZHAO K Y, SUN J L. Determination of polypeptide content in Compound Guopeptide Injection by amino acid analysis[J]. Chinese Pharmaceutical Affairs,2011,25(8):816−820.] TIAN H B, ZHAO K Y, SUN J L. Determination of polypeptide content in Compound Guopeptide Injection by amino acid analysis[J]. Chinese Pharmaceutical Affairs, 2011, 25（8）: 816−820.
[19]	芮鸿飞, 张晓瑜, 刘兴泉, 等. PITC柱前衍生-反相高效液相色谱法测定黄酒种游离氨基酸和生物胺[J]. 食品科学,2016,37(8):159−163. [RUI H F, ZHANG X Y, LIU X Q, et al. Determination of free amino acids and bioamines in rice wine by PITC precolumn derivation-reversed-phase high performance liquid chromatography[J]. Food Science,2016,37(8):159−163.] RUI H F, ZHANG X Y, LIU X Q, et al. Determination of free amino acids and bioamines in rice wine by PITC precolumn derivation-reversed-phase high performance liquid chromatography[J]. Food Science, 2016, 37（8）: 159−163.
[20]	许永, 黄丽佳, 刘欣, 等. 柱前衍生-高效液相色谱法测定新鲜烟叶中游离氨基酸[J]. 化学试剂,2021,43(6):801−805. [XU Y, HUANG L J, LIU X, et al. Determination of free amino acids in tobacco leaves by high performance liquid chromatography with precolumn derivatization[J]. Chemical Reagents,2021,43(6):801−805.] XU Y, HUANG L J, LIU X, et al. Determination of free amino acids in tobacco leaves by high performance liquid chromatography with precolumn derivatization[J]. Chemical Reagents, 2021, 43（6）: 801−805.
[21]	张国华, 吴光斌, 陈昭华, 等. 异硫氰酸苯酯柱前衍生RP-HPLC法同时测定莲雾中15种游离氨基酸的含量[J]. 食品工业科技,2020,20:230−234. [ZHANG G H, WU G B, CHEN S H, et al. Simultaneous determination of 15 kinds of free amino acids in Lotus spray by RP-HPLC with precolumn derivation of phenyl isothiocyanate[J]. Science and Technology of Food Industry,2020,20:230−234.] ZHANG G H, WU G B, CHEN S H, et al. Simultaneous determination of 15 kinds of free amino acids in Lotus spray by RP-HPLC with precolumn derivation of phenyl isothiocyanate[J]. Science and Technology of Food Industry, 2020, 20: 230−234.
[22]	周敏. 应用不同柱前衍生法于HPLC测定西洋参种的氨基酸[J]. 基因组学与应用生物学,2018,37(2):524−530. [ZHOU M. Determination of amino acids of American ginseng by HPLC with different pre-column derivations[J]. Genomics and Applied Biology,2018,37(2):524−530.] ZHOU M. Determination of amino acids of American ginseng by HPLC with different pre-column derivations[J]. Genomics and Applied Biology, 2018, 37（2）: 524−530.
[23]	傅博强, 唐治玉, 王晶, 等. 转基因大米中氨基酸含量超高效液相色谱定量[J]. 中国粮油学报,2014,29(1):110−115. [FU B Q, TANG Z Y, WANG J, et al. Ultrahigh performance liquid chromatography quantification of amino acid content in genetically modified rice[J]. Chinese Journal of Cereals and Oils,2014,29(1):110−115.] FU B Q, TANG Z Y, WANG J, et al. Ultrahigh performance liquid chromatography quantification of amino acid content in genetically modified rice[J]. Chinese Journal of Cereals and Oils, 2014, 29（1）: 110−115.
[24]	周侃, 熊华, 陈升军, 等. 高蛋白含量米蛋白肽制备及产品营养成分分析[J]. 食品与生物技术学报,2009,28(1):28−33. [ZHOU K, XIONG H, CHEN S J, et al. Preparation and nutritional analysis of high protein rice protein peptide[J]. Journal of Food and Biotechnology,2009,28(1):28−33.] ZHOU K, XIONG H, CHEN S J, et al. Preparation and nutritional analysis of high protein rice protein peptide[J]. Journal of Food and Biotechnology, 2009, 28（1）: 28−33.
[25]	QIAN X, MENG W W, JU W H, et al. The role of heating time on the characteristics, functional properties and antioxidant activity of enzyme-hydrolyzed rice proteins-glucose Maillard reaction products[J]. Food Bioscience,2020,43:1−8.
[26]	THIDARAT P, MASATOSHI K, PRISANA S, et al. Characterization and bioactivities of young rice protein hydrolysates[J]. Journal of Cereal Science,2020,95:1−9.
[27]	XIN A Z, ZHONG Y Z, PEI B Y, et al. Rice peptide nanoparticle as a bifunctional food-grade Pickering stabilizer prepared by ultrasonication:Structural characteristics, antioxidant activity, and emulsifying properties[J]. Food Chemistry,2021,343:1−10.
[28]	SAPNA R, KM P, KUMAR P G. Exploration of rice protein hydrolysates and peptides with special reference to antioxidant potential:Computational derived approaches for bio-activity determination[J]. Trends in Food Science & Technology,2018,80:61−70.
[29]	ZHANG Z L, WANG Y, JIANG H, et al. Effect of dual-frequency ultrasound on the formation of lysinoalanine and structural characterization of rice dreg protein isolates[J]. Ultrasonics Sonochemistry,2020,67:105−114.
[30]	王申, 林利美, 周佳, 等. 大米肽的绝对分子质量与氨基酸组成及体外活性[J]. 食品科学,2013,34(1):19−23. [WANG S, LIN L M, ZHOU J, et al. Absolute molecular weight, amino acid composition and in vitro activity of rice peptides[J]. Food Science,2013,34(1):19−23.] WANG S, LIN L M, ZHOU J, et al. Absolute molecular weight, amino acid composition and in vitro activity of rice peptides[J]. Food Science, 2013, 34（1）: 19−23.