Analysis of Amino Acids in Houttuynia Cordata Thunb. by High Performance Liquid Chromatography-Diode Array Detector (HPLC-DAD)

[Objectives] The biologically active compounds of Houttuynia cordata Thunb. as a medicinal and edible plant are well documented. However, amino acid as an important indicator of the value of H. cordata is little reported. [Methods] Amino acids of the H. cordata were analysed by pre-column derivatization with 2,4-dinitrochlorobenzne(CDNB) coupled to HPLC/DAD. The optimized conditions of the amino acids derivatized by CDNB were followed: 110 ℃ of hydrolysis temperature, 1∶12 of the equivalent proportion between amino acid and CDNB, 80 min of derivatization reaction time and 80 ℃ of derivatization reaction temperature. The method was used to determine the amino acids of H. cordata samples from 11 different geographical sites in Guizhou. [Results] The results suggested that the contents of 17 amino acids were quite variable at different H. cordata samples. [Conclusions] This study will provide a basis for the quality evaluation of H. cordata nutrition.

Rapid Determination of Amino Acids in Ginseng by High Performance Liquid Chromatography and Chemometric Resolution

Ginseng is one of the most important traditional Chinese medicines and functional foods.A method for the fast determination of amino acids in ginseng samples using high performance liquid chromatography（HPLC） was developed,in which strong isocratic elution was employed for simplifying the separation and speeding up the analysis.All amino acids were eluted within 3 min with the chromatogram composed of overlapped peaks from the interferences.Then,non-negative immune algorithm（NNIA） was adopted to resolve the chromatographic signals of the components from the chromatogram measured.The results show that the signals of the amino acids can be correctly extracted by NNIA and the signal extracted can be used for the quantitative analysis.The method was validated via determining six amino acids of four different samples of ginseng.The recoveries of the spiked samples are in a range of 96.6％-106.3％.

Metabolic profile analysis of free amino acids in experimental autoimmune uveoretinitis rat plasma

AIM: To determine the differences of amino acid(AA) levels in experimental autoimmune uveoretinitis(EAU). METHODS: AA analysis of the plasma samples in EAU rats induced by interphotoreceptor retinoid-binding protein emulsion were performed with high performance liquid chromatography(HPLC) and phenylisothiocyanate(PITC) pre-column derivation methods were performed. Using partial least squares discriminant analysis(PLS-DA), the potential biomarkers were identified in EAU rat plasma, and the metabolic pathways related to EAU were further analyzed. RESULTS: The method results showed that linear(r≥0.9957), intra-day reproducible [relative standard deviation(RSD)=0.04%-1.33%], inter-day reproducible(RSD=0.06%-2.07%), repeatability(RSD=0.03%-0.89%), stability(RSD=0.05%-2.48%) and recovery(RSD=1.98%-4.39%), with detection limits of 0.853-11.4 ng/mL. The metabolic profile in EAU rats was different from that in the control groups five AAs concentrations were increased and nine AAs were reduced. Moreover, five metabolic pathways were related to the development of EAU. CONCLUSION: The developed method is a simple, rapid and convenient for determination of AAs in EAU rat plasma, and these findings will provide a comprehensiveinsight on the metabolic profiling of the pathological changes in EAU.

Effect of borneol, moschus, storax, and acorus tatarinowii on expression levels of four amino acid neurotransmitters in the rat corpus striatum

The present study collected cerebrospinal fluid samples from the corpus striatum in rats treated with borneol, moschus, storax, and acorus tatarinowii using brain microdialysis technology. Levels of excitatory neurotransmitters aspartic acid and glutamate, as well as inhibitory neurotransmitters glycine and ^-aminobutyric acid, were measured in samples using reversed-phase high-performance liquid chromatography, phosphate gradient elution, and fluorescence detection. Results showed that concentrations of all four amino acid neurotransmitters significantly increased in the corpus striatum following treatment with borneol or moschus, but effects due to borneol were more significant than moschus. Acorus tatarinowii treatment increased ^-aminobutyric acid expression, but decreased glutamate concentrations. Storax increased aspartic acid concentrations and decreased glycine expression. Results demonstrated that borneol and moschus exhibited significant effects on con amino acid neurotransmitter expression; storax exhibited excitatory effects and acorus tatarinowii resulted in inhibitory effects.

Determination of 18 Kinds of Amino Acids in Fresh Tea Leaves by HPLC Coupled with Pre-column Derivatization

A rapid and accurate quantitative method of high performance liquid chromatography( HPLC) with fluorescence detector has been developed for the analysis of 18 kinds of amino acids in fresh tea leaves. The samples were minced and mixed,and extracted with ultra pure water at 90℃ for 20 min. The 6-aminoquinolyl N-hydroxy-succinimidyl carbamate( AQC) was used as pre-column derivatization reagent. Gradient HPLC separation was performed on a C_(18) column( Symmetry C_(18),3. 9 mm × 15 cm,4 μm). Good linearity between concentrations and peak areas was achieved in the concentration range of 5. 0-250 μmol/L for 18 kinds of amino acids. The method was validated by the analysis of five replicates. The 18 kinds of amino acid standards were spiked in fresh tea leaf samples and the average recovery rate was 86. 25%-109. 05% with relative standard deviations( n = 5) ranging from 6. 03% to 10. 56%. The limit of detection( LOD) for the analytes was0. 05-1. 27 μmol/L. The method was successfully applied to the analysis of the 18 kinds of amino acids in fresh tea leaves from east Dongting and west Dongting mountains in Suzhou. The results indicate that the method is simple,rapid,precise and reliable.

高效液相色谱-紫外检测法同时测定洗面奶中4种N-月桂酰氨基酸表面活性剂

N-月桂酰氨基酸(NLAAs)表面活性剂与传统表面活性剂相比在安全性上具有明显的优势,应用领域日益增多。但由于NLAAs的紫外吸收较弱,采用高效液相色谱-紫外检测进行含量测定存在一定困难,一般使用示差折光指数检测器、蒸发光散射检测器等通用型检测器来进行检测。为解决这一难题,本研究通过优化流动相的组成,选择合适的色谱柱和检测波长,建立了高效液相色谱-紫外检测同时测定月桂酰谷氨酸(LG)、月桂酰甘氨酸(LC)、月桂酰丙氨酸(LA)、月桂酰肌氨酸(LS)4种NLAAs表面活性剂的方法。样品在乙腈-0.10%H_(3)PO_(4)水溶液(60∶40,v/v)中超声提取10 min,滤膜过滤后用高效液相色谱仪测定,以乙腈-0.10%H_(3)PO_(4)水溶液为流动相进行梯度洗脱,经Agilent Eclipse Plus C_(18)色谱柱(150 mm×4.6 mm,5μm)分离,在205 nm波长处进行检测。结果显示:4种NLAAs在2.0~800.0 mg/L范围内呈良好的线性关系,相关系数(r)均≥0.9995,检出限(LOD)为0.17~0.49 mg/L,定量限(LOQ)为0.57~1.63 mg/L。在0.60、4.50、15.00、24.00 mg/g 4个加标水平下,NLAAs的加标回收率在94.3%~107.4%范围内。采用该方法对5个洗面奶样品中的NLAAs进行含量测定,发现所有样品都含有一种或多种NLAAs化合物,NLAAs的总含量在64.58~97.01 mg/g范围内。该方法前处理简单,测试快速,方法精密度、准确性、稳定性良好,适于洗面奶中NLAAs的含量测定,也可为该类型表面活性剂原料纯度、合成产率检测及产品的质量控制提供有效的技术参考。

柱前衍生-高效液相色谱法测定巴戟天中氨基酸含量及营养价值评价

建立柱前衍生-高效液相色谱法测定巴戟天中氨基酸含量,采用氨基酸比值系数法评价其营养价值。采用盐酸水解法制得巴戟天氨基酸供试品溶液,以异硫氰酸苯酯和三乙胺为柱前衍生化试剂,采用高效液相色谱法,使用Ultimate Amino Acid色谱柱(4.6 mm×250 mm,5μm)以0.1 mol/L无水乙酸钠-乙腈为流动相,梯度洗脱,柱温40℃,检测波长为254 nm,测定氨基酸含量;通过氨基酸比值、氨基酸比值系数、氨基酸比值系数分等评价其营养价值,并采用系统聚类和主成分分析对其进行分类分析。结果表明,15种氨基酸线性关系良好,相关系数r≥0.9996。16份巴戟天氨基酸总含量为17.87~102.15 mg/g,均检测到15种氨基酸,其中包含6种必需氨基酸,占总氨基酸含量的14.15%~32.06%;8种药用氨基酸,占总氨基酸含量的45.20%~56.31%。限制氨基酸为苏氨酸和亮氨酸,氨基酸比值系数分为33.87~61.31。不同巴戟天样品氨基酸组成相关性显著;主成分分析提取2个主成分,累计方差贡献率为97.085%,可将16批巴戟天分为三类。该实验测定方法简单有效,可同时分离测定15种氨基酸,精密度与重复性良好,有较高的药用价值,可为巴戟天的品质分析和资源开发利用提供参考。

柱前衍生-高效液相色谱法测定化妆品中16种氨基酸的含量

提出了2,4-二硝基氟苯(DNFB)柱前衍生-高效液相色谱法测定化妆品中甘氨酸、亮氨酸、组氨酸、缬氨酸、甲硫氨酸、脯氨酸、半胱氨酸、苏氨酸、谷氨酸、赖氨酸、酪氨酸、精氨酸、异亮氨酸、色氨酸、苯丙氨酸、丙氨酸等16种氨基酸含量的方法。取化妆品样品5 g置于50 mL离心管中,加入5 mL水,超声10 min,加热煮沸15 min,冷却后加入15 mL 35 g·L^(−1)磺基水杨酸溶液,以转速10500 r·min^(−1)离心20 min。取全部上清液,用35 g·L^(−1)磺基水杨酸溶液定容至25 mL。取1 mL样品溶液,加入0.5 mol·L^(−1)碳酸氢钠溶液(pH 9.0)1 mL和1%(体积分数)DNFB衍生溶液1 mL,混匀,于60℃加热10 min,冷却后用0.01 mol·L^(−1)磷酸二氢钾溶液(pH 7.0)定容至5 mL,所得溶液中16种氨基酸经X Bridge C_(18)色谱柱分离,以不同体积比的含1%(体积分数)N,N-二甲基甲酰胺的0.05 mol·L^(−1)乙酸钠缓冲液(pH 6.5)-体积比1∶1的乙腈-水混合溶液为流动相进行梯度洗脱,在360 nm处检测。结果表明:16种氨基酸标准曲线的线性范围均为0.004~40.0 mg·L^(−1),检出限(3S/N)为0.1~0.2 mg·kg^(−1);对空白样品进行3个浓度水平的加标回收试验,回收率为89.0%~105%,测定值的相对标准偏差(n=7)均不大于10%。方法用于11种市售化妆品分析,检出的氨基酸成分多为组氨酸、甘氨酸、甲硫氨酸、谷氨酸。

湖南石门县地方品种鸡肉氨基酸组成分析及蛋白质营养价值评价

目的探究湖南省石门当地7个品种鸡肉中的基本营养成分及氨基酸组成差异,并对其进行蛋白质营养价值评价。方法采用相关国家标准测定7个品种鸡肉中水分、粗灰分、粗脂肪、粗蛋白含量,并利用高效液相色谱仪对鸡肉氨基酸含量进行测定,基于联合国粮食及农业组织和世界卫生组织(Foodand Agriculture Organization/World Health Organization,FAO/WHO)标准模式谱对不同品种鸡胸肉和腿肉蛋白质分别进行营养学评价。结果7个品种鸡胸肉、腿肉中水分含量分别为71.20~74.97 g/100 g、72.65~76.22 g/100 g,粗灰分含量分别为0.90~1.38 g/100 g、0.91~1.28 g/100 g,粗脂肪含量分别为0.15~0.64 g/100 g、2.30~7.74 g/100 g,粗蛋白质含量分别为22.29~24.61 g/100 g、19.00~21.80 g/100 g,不同品种、部位间存在差异。7个品种鸡肉均检测出17种氨基酸,均以谷氨酸含量为最高;胸肉氨基酸含量高于腿肉(除壶瓶山鸡),其中青脚鸡胸肉和壶瓶山鸡腿肉的总氨基酸含量最高。7个品种鸡肉必需氨基酸与总氨基酸的比值与FAO/WHO推荐的理想蛋白质模式较为接近;其中苏氨酸、异亮氨酸、亮氨酸、苯丙氨酸+酪氨酸、赖氨酸占总氨基酸的质量分数均明显高于FAO/WHO推荐的模式谱标准值;除麻鸡胸肉、青脚鸡腿肉外,其余鸡种胸肌和腿肌氨基酸比值系数分均能达70以上。结论7个品种鸡肉中蛋白质与FAO/WHO规定的人体蛋白需求相类似,蛋白质营养价值较高,本研究结果以期为树立石门当地肉鸡口碑、进一步选育和开发利用新鸡种提供科学依据。

市售鱼鳔氨基酸及蛋白质成分分析

目的 分析16批不同基原的市售鱼鳔中氨基酸及蛋白质成分。方法 建立一种6-氨基喹啉基-N-羟基琥珀酰亚胺基甲酸酯柱前衍生、高效液相色谱法,测定鱼鳔中17种氨基酸含量,并对不同基原鱼鳔中氨基酸类成分进行分析;采用十二烷基硫酸钠聚丙烯酰胺凝胶电泳分析鱼鳔中的蛋白质分子量分布,并采用蛋白质组学法鉴定鱼鳔中的蛋白质。结果 17种氨基酸在一定浓度范围内线性关系良好(R2≥0.998 0),平均加样回收率85.62%~109.60%,精密度、稳定性、重复性RSD均小于3.5%。16批鱼鳔氨基酸组成与含量存在差异,氨基酸总含量介于468.31~620.05 mg/g;从16批鱼鳔中共鉴定688种蛋白质,包括Ⅰ型在内的胶原蛋白多达11种,此外,鱼鳔中还含有较多分子质量52~95 kDa的低丰度蛋白。结论 该研究可为鱼鳔的质量评价和开发利用提供参考。

基于超高效液相色谱-三重四极杆串联质谱法的黑蒜加工过程中特征成分变化规律

基于超高效液相色谱-三重四极杆串联质谱技术建立了黑蒜中10种风味前体物质和21种游离氨基酸同时测定的检测方法。将该方法应用于黑蒜加工过程中特征成分变化规律的研究,结果表明,在温度75℃、相对湿度85%条件下,黑蒜中的特征成分含量在加工过程中发生了较大改变。其中,γ-氨基丁酸、S-烯丙基-L-半胱氨酸、异蒜氨酸、谷氨酰胺、甲基蒜氨酸、蒜氨酸、色氨酸和γ-L-谷氨酰-S-烯丙基-L-半胱氨酸变化最为显著,是黑蒜加工过程中小分子代谢物变化的化学标志物。该方法灵敏度高、准确性好,可满足检测需求。

用于检测羧酸的基于高效毛细管电泳的微流控芯片

设计了一种可以进行高效毛细管电泳分析的微流控芯片,微流控芯片具有一个十字架结构的沟道,沟道横截面的尺寸为100μm×50μm。利用5-氨基甲基荧光素作为荧光探针,使用乙腈溶液与磷酸盐缓冲液作为流动相进行高效液相色谱分析,等量洗脱确认了丙酸到十八烷酸(C3~C18)的衍生物色谱图,并通过高效毛细管电泳分析将丙酸(C3)、己酸(C6)、癸酸(C10)进行分离,成功分离C3和C10,C3和C10的分离度为2.2,分离效果良好。此外,针对高效毛细管电泳分析出现的不稳定电渗现象而导致实验数据不稳定的情况,加入阳离子聚合物六甲溴铵和阴离子聚合物硫酸葡聚糖钠盐形成连续的多个离子聚合物涂层,使得微流控芯片对羧酸检测的稳定性大大提高。

柱前衍生-HPLC法测定天龙泉糯米调味酒中18种氨基酸

为满足企业对糯米调味酒精细化生产的品控需求,该研究利用反相色谱柱建立了测定天龙泉糯米调味酒中18种氨基酸的柱前衍生-高效液相色谱(HPLC)方法。通过优化洗脱梯度,使待测糯米调味酒的18种氨基酸色谱峰实现有效分离,确定最优色谱条件,进而建立了定量方法并对其进行方法学考察。结果表明,优化后的色谱条件为以异硫氰酸苯酯作为衍生试剂,乙腈∶水(4∶1,V/V)为流动相A,0.019 mol/L乙酸钠溶液(pH=6.2)为流动相B,梯度洗脱,柱温28℃,流速1 mL/min,检测波长245 nm。在各氨基酸标准曲线的线性范围内相关系数(R^(2))均≥0.998 7,平均加标回收率为92.38%~104.86%,相对标准偏差(RSD)≤4.45%,重复性、精密度试验结果RSD均<5%。该方法操作简单,准确高效,可定量测定天龙泉糯米调味酒及相关产品中各氨基酸的含量。

磁性分散固相微萃取/UHPLC-Q-Orbitrap HRMS测定运动营养食品中27种氨基酸

基于磁性分散固相微萃取净化,建立了测定运动营养食品中27种氨基酸含量的超高效液相色谱-四极杆-静电场轨道阱高分辨质谱法。通过优化液相色谱条件、质谱条件和样品前处理过程,在20 min内实现了对27种目标物的测定。样品经涡旋振荡,超声提取,磁性氧化石墨烯分散固相微萃取净化,采用Thermo Accucore HILIC色谱柱分离,以0.1%甲酸水溶液(含5.0 mmol/L甲酸铵)和0.1%甲酸乙腈(含5.0 mmol/L甲酸铵)为流动相进行梯度洗脱,静电场轨道阱高分辨质谱检测,外标法定量。结果表明,27种目标化合物在一定质量浓度范围内线性良好,相关系数(r^(2))均大于0.99,方法的定量下限为0.10~0.25 mg/kg,平均回收率为70.0%~93.0%,日内相对标准偏差(RSD,n=6)为1.6%~10%,日间RSD(n=5)为1.4%~5.2%。该方法高效灵敏,准确可靠,适用于运动营养食品中27种氨基酸的测定。

柱前衍生-高效液相色谱法测定特殊配方食品中的牛磺酸

目的建立实心核颗粒技术-柱前衍生-高效液相色谱法测定氨基酸配方及蛋白水解配方等特殊配方食品中牛磺酸的分析方法。方法样品加水溶解提取后沉淀蛋白,上清液与足量丹磺酰氯衍生反应;采用实心核颗粒C18色谱柱分离,流动相为乙腈-乙酸钠溶液,梯度洗脱,高效液相色谱荧光检测器测定,外标法定量。结果牛磺酸峰型良好,与杂质基线分离,在0~40.0μg/mL范围内线性良好(r^(2)>0.999),方法检出限和定量限分别为0.04mg/100g、0.10mg/100g;在低、中、高3个浓度水平下的平均回收率范围为90.5%~105.4%,相对标准偏差为0.5%~8.1%。结论该方法准确度高、重复性和稳定性好,适用于氨基酸配方及蛋白水解配方等特殊配方食品的检测,解决了国标方法检测氨基酸配方及深度水解蛋白乳粉等特殊配方食品中牛磺酸方法的不足。

OPA柱前衍生高效液相色谱法同时测定黄豆酱中氨基酸和生物胺的含量

本文采用OPA衍生黄豆酱中的组氨酸、酪氨酸、组胺和酪胺,再通过高效液相色谱进行分离测定。结果表明,该方法的相对标准偏差在0.04%~1.16%,回收率在93.4%~101.2%,检出限为6~9 mg·kg^(-1)。该方法能够满足黄豆酱组氨酸、酪氨酸、组胺和酪胺的检测需求。

HPLC法测定青稞中γ-氨基丁酸含量的研究

建立了一种测定青稞中γ-氨基丁酸含量的方法,青稞样品中的γ-氨基丁酸与邻苯二甲醛进行衍生反应,采用高效液相色谱法测定,通过色谱柱分离,用外标法定量,最后得出样品中γ-氨基丁酸的含量,γ-氨基丁酸含量在0.1~1.0 mg/m L范围内线性关系良好,平均回收率为98%~103%,RSD为2.0%,此方法可以准确快速的检测青稞中γ-氨基丁酸的含量。

72份武汉茶叶氨基酸检测与高茶氨酸品种筛选

本文对来自湖北的72份茶叶原样进行检测分析,拟选育出高茶氨酸品系。实验参考《植物中游离氨基酸的测定》(GB/T 30987—2020)中的方法,结合高效液相色谱仪对茶叶中各种游离氨基酸的含量进行检测分析,对其中茶氨酸含量与游离氨基酸含量进行综合分析评估,并筛选出优质高茶氨酸品种作为推广种植质资源。实验通过对茶氨酸总含量、茶氨酸在游离氨基酸中的占比,以及茶叶品种差异进行分析,为高茶氨酸茶的实现提供可参考思路。本文为选育高茶氨酸品系提供一种方案,旨在加强高茶氨酸品种的开发,提升茶叶产品质量和丰富度,便于茶叶产品推广,增强中国茶叶的影响力、竞争力。

超高效液相色谱法测定细胞培养液中游离氨基酸浓度

目的探讨基于超高效液相色谱(ultra performance liquid chromatography,UPLC)法,对细胞培养液中游离氨基酸浓度的测定方法。方法采用UPLC技术,结合优化的样品处理方法,对细胞培养液中游离氨基酸进行高效、精准的测定。结果该UPLC法能够快速、可靠地测定细胞培养液中的游离氨基酸浓度,为生物制药工业中细胞培养监控与优化提供了有效手段。结论对氨基酸浓度的准确测定,不仅有助于更好地了解细胞培养液中关键营养成分的动态变化,还为生产过程中的工艺优化提供了重要数据支持。

异硫氰酸苯酯柱前衍生化反相高效液相色谱法同时测定18种氨基酸

建立了一种利用反相高效液相色谱同时测定 18种氨基酸的方法。以正亮氨酸为内标物 ,异硫氰酸苯酯为柱前衍生剂 ,用C18色谱柱在柱温 38℃下采用二元梯度洗脱 ,于 2 5 4nm波长处检测。氨基酸质量浓度在 3 5mg/L～ 5 5 6mg/L时 ,其峰面积与内标物峰面积的比值和氨基酸的质量浓度的线性相关系数 ,除胱氨酸 (0 96 2 )外均大于 0 99;18种氨基酸的加标回收率在 96 0 %～ 10 2 .4 %。信噪比为 2时 ,亮氨酸最低检测限为 0 5mg/L。应用该方法对小牛血去蛋白注射液中的游离氨基酸含量进行了测定 。