HPLC法同时测定复合氨基酸多维胶囊中L-茶氨酸和维生素B_6的含量

目的建立同时测定复合氨基酸多维胶囊中L-茶氨酸和维生素B6含量的HPLC法。方法采用Phmemex LunaC18柱,以0.005 mol/l的十二烷基硫酸钠溶液(pH3.0)-乙腈(66:34)为流动相,流速1.0 ml/min,检测波长205 nm。结果专属性良好,L-茶氨酸、维生素B6、空白辅料与其他功效成分以及其酸、碱、氧化破坏产物无干扰。L-茶氨酸在64.19～96.29μg/ml的范围内线性关系良好,γ=0.9994,回收率为101.1%,中间精密度、重复性良好。维生素B6在8.10～12.15μg/ml范围内呈良好线性,γ=0.9993,回收率为100.1%,中间精密度、重复性良好。结论该反向高效液相色谱法能有效分离检测本胶囊中L-茶氨酸和维生素B6的含量。     

农产品中含磷氨基酸类除草剂的HPLC测定

对含磷氨基酸类除草剂的残留测定,已往用HPLC法、GC法,且每种农药分别用不同方法,操作较繁琐。本文报道了用HPLC法同时测定草甘膦(glyphosate,GLY)、草丙膦(glufosinate,GLU)、bi-lanafos(BIA)的灵敏分析方法。用于西红柿、柑桔、黄瓜、苹果中三种农药的测定,对添加2mg/kg标准品的回收率为69.7%～126.8%,检出限为0.02～0.04mg/kg。HPLC测定条件　色谱柱:Nucleosil100-SB(内径4.6mm×250mm);流动相:1mol/L磷酸-乙腈-水(用5mol/LKOH调至pH3.1)(1∶4∶5);流速:0.7ml/min;柱温:50℃;荧光测定波长:λex=255nm.λem=310nm;进样量:10…     

反相高效液相色谱荧光法测定大鼠血清和脑组织中茶氨酸含量

目的建立一种反相高效液相色谱荧光检测法,测定大鼠血清和脑组织中茶氨酸含量。方法用ODS-SP C18色谱柱(4.6 mm×250 mm,粒径5μm),以磷酸二氢钾缓冲液(0.1 mol/L,pH6.0):四氢呋喃:甲醇体积比=62:3:35为流动相;高丝氨酸为内标;邻苯二甲醛柱前衍生;流速0.85 ml/min;荧光检测器激发波长340 nm、发射波长455 nm检测大鼠血清、脑组织中茶氨酸含量。结果茶氨酸浓度在1～20μmol/L范围内线性关系良好,相关系数为0.9997;浓度为1、10、20μmol/L时日内RSD(%)分别为3.51、1.58、0.75,日间RSD(%)分别为4.54,3.25,2.87;茶氨酸平均回收率范围为91.8%～99.1%;最低检测限为0.02μmol/L。SD大鼠给予200 mg/kg.bw茶氨酸灌胃后,血清中峰值出现在0.5～1.0 h之间,脑组织中浓度5 h达到峰值。结论该反相高效液相色谱荧光法灵敏度高,重复性好,能有效分离检测大鼠血清、脑组织中茶氨酸含量。     

高效液相色谱法测定青霉素V钾分散片的高分子杂质

目的:建立一种高效液相色谱法测定青霉素V钾分散片的高分子杂质的方法.方法:在SephadexG-10(10mm×300mm)色谱柱上;以0.1mol/L磷酸盐缓冲液(pH7.0)(取21.85gNa2HPO4·12H2O和6.08gNaH2PO4·2H2O,加水溶解并稀释至1000ml,用磷酸调pH至7.0)为流动相A;以蒸馏水为流动相B;检测波长268nm;流量为1.0ml/min.结果:青霉素V钾分散片的高分子杂质均小于2.0%.结论:本方法准确、简便,适用于本品的质量控制.     

化妆品中马来酸氯苯那敏与咖啡因的测定

目的建立同时测定化妆品中马来酸氯苯那敏和咖啡因含量的HPLC方法。方法采用Sheshido SPOLAR C18(4.6 mm×250 mm,5μm)色谱柱,以1%三乙胺溶液(磷酸调至p H 2.8)-甲醇-乙腈(85∶3∶12)为流动相,检测波长为262 nm,进样量10μL,流速为1.0 m L/min,柱温为30℃。结果方法线性浓度范围马来酸氯苯那敏为0~103.2μg/m L,咖啡因为0~262.5μg/m L,相关系数均大于0.999;最低检出限马来酸氯苯那敏为8.7 mg/kg,咖啡因为2.5 mg/kg;高、中、低3个浓度水平的加标回收率为96%~103%;测定值的相对标准偏差(n=6)小于5%。结论 HPLC法准确、专属性强、灵敏度高,适用于同时测定化妆品中马来酸氯苯那敏和咖啡因。     

HPLC法测定盐酸多西环素片中盐酸多西环素的含量

[目的]建立HPLC测定盐酸多西环素片的含量.[方法]采用ODS柱(250mm×4.6mm,5 um),以0.05 mol/L草酸铵溶液:二甲基甲酰胺:0.2 mol/L磷酸氢二铵溶液(65:30:5) (用氨试液调pH 8.3)为流动相,检测波长为280 nm. [结果]峰面积对浓度的回归方程为A=21 489 C-20 287,盐酸多西环素在20-120 ug/ml浓度范围内,呈良好的线性关系(r=0.999 9),平均回收率为100.84%,RSD为0.80%(n=5). [结论]该法简便、准确,适用于盐酸多西环素片剂的含量的测定.     

反相高效液相色谱法测定大鼠胰腺组织中头孢噻肟钠的含量

[目的]建立反相高效液相色谱法测定大鼠胰腺组织中头孢噻肟钠含量. [方法]以茶碱为内标,采用YMC-C18柱,以0.015 mol/L 磷酸二氢钾缓冲液-甲醇溶液 (pH 调4.70) (82:18) 为流动相,流速 0.8 ml/min,紫外检测波长为254 nm. [结果]标准曲线线性范围为:16～0.5(μg/ml);头孢噻肟检测限为0.25μg/ml;提取回收率为:76.70%～85.24%;方法回收率为 106.21%～107.79%;日内RSD在5.74%～10.76%之间,日间 RSD 在5.53%～9.14%之间. [结论]该方法揉作简单,结果准确,重现性好,用于大鼠胰腺组织中头孢噻肟钠含量测定得到满意结果.     

HPLC法测定祛风止痛浓缩丸中没食子酸的含量

目的建立祛风止痛浓缩丸中没食子酸的含量测定方法。方法采用HPLC法测定其中没食子酸的含量。以十八烷基硅烷键合硅胶为填充剂;甲醇-0.025mol/L磷酸(用三乙胺调pH2.7)(5:95)为流动相;检测波长为266nm。结果在选定的色谱条件下,供试液中没食子酸与相邻组分离度良好,阴性液没有干扰,方法的稳定性、重现性、精密度、回收率试验(RSD%=1.8%,n=6)均符合相关规定,三批样品中每丸含没食子酸分别为4.21mg、4.45mg和3.96mg。结论采用HPLC法可以作为祛风止痛浓缩丸中没食子酸的含量测定方法。     

高效液相色谱法同时测定抗疲劳保健食品中的3种违禁成分

目的 建立抗疲劳类保健食品中违禁成分咖啡因、西地那非和他达那非的高效液相色谱法。方法 样品中的待测组分经甲醇超声提取、过滤后,采用Welch C18柱(4.6 mm×250 mm,5 μm)为分离柱,以三乙胺溶液(取三乙胺7 ml,加水至1 L,并用磷酸调节pH值至4.0)(A)-乙腈(B)为流动相,梯度洗脱流速为1.0 ml/min;检测波长为280 nm;柱温为40 ℃,标准曲线法定量。结果 3种化合物分离度良好,线性相关系数均大于0.999 9,回收率为93.2 %～103.7 %,检出限为3.0～4.4 μg/g,定量限为8.9～13.2 μg/g。结论 本法简便、准确,可用于抗疲劳类保健食品中非法添加化学药物的检测。     

蔬菜中百草枯的高效液相色谱测定法

目的建立蔬菜中百草枯残留量的高效液相色谱-紫外检测器测定方法。方法选用AgilentTC-C18色谱柱(4.6×250mm,5μm),检测波长257nm,以10mmol/L庚烷磺酸钠(用磷酸调pH=3.0)-甲醇(50+50)作为流动相,柱流量1.0ml/min,采用高效液相色谱法检测蔬菜中百草枯含量。结果该方法的线性范围为0.00～10.0μg/ml,R2=0.9994,RSD为0.75%～3.65%,回收率为96.1%～104.0%,检出限为0.03μg/ml。结论该方法简单、快速、准确度高,适用于蔬菜中百草枯残留量的测定。     

茶叶及茶饮料中儿茶素和咖啡因的多组分HPLC分析方法

目的　建立茶叶及茶饮料中儿茶素和咖啡因多组分HPLC测定方法。方法　以水 -乙醇 (3∶7)为提取剂 ,超声提取 ,提取液离心后 ,采用C1 8柱分离 ,HPLC- UV检测。以 0 . 1%H3PO4 的甲醇溶液为流动相A ,0 .1%H3PO4 的水溶液为流动相B ,进行梯度洗脱 ,流速为 1ml min,检测波长为 2 10nm。结果　在给定的浓度范围内 ,各儿茶素组分和咖啡因的峰面积与其相应浓度呈现良好的线性相关 ,r>0. 999。添加已知儿茶素含量的茶粉回收试验结果显示 ,各组分儿茶素及咖啡因的平均回收率为 6 1. 7%～ 117. 4 % ,相对标准差 <10 %。结论　本方法分离度好 ,灵敏度高 ,准确快速 ,为研究儿茶素在机体内的吸收利用提供了可靠手段。     

An improved HPLC method for simultaneous determination of phenolic compounds,purine alkaloids and theanine in Camellia species

A simple and fast HPLC analysis method for phenolic compounds, purine alkaloids and theanine in various Camellia species was developed. Using an amide-C16 column, catechins, gallic acid, caffeine, theobromine, theacrine, theophyline and theanine could be rapidly separated within 45 min with a gradient elution system. Excellent linearity was observed for all 14 compounds in the range studied, with correlation coefficients between 0.9994 and 0.9999. Limit of detection and limit of quantification of the 14 compounds varied from 0.0001 to 0.072 ng/μL and 0.0004 to 0.24 ng/μL, respectively. Four kinds of Camellia species were analyzed using this method. In traditional cultivated tea trees, Camellia sinensis and Camellia assamica, the main purine alkaloid was caffeine and main phenolic compound was (−)-epigallocatechin gallate (EGCG). The main purine alkaloids in Camellia ptilophylla and C. assamica var. kucha were theobromine and theacrine; their contents were 4.001±0.1184% dry weight and 2.116±0.0270% dry weight, respectively. The main phenolic compounds in C. ptilophylla and C. assamica var. kucha were (−)-gallocatechin gallate (GCG) and EGCG respectively. The content of theanine in the four Camellia species samples ranged from 0.136±0.0026% to 1.485±0.0491% dry weight.     

绿茶中阻断亚硝胺合成的有效成分的分离鉴定

茶叶用沸水(1:10)提取10 min,过滤并冷冻干燥(Ⅰ);另将茶水用氯仿、乙酸乙酯先后提取浓缩,并与残水一起分别冷冻干燥,得组分Ⅱ、Ⅲ、Ⅳ。用Sephadex LH20对组分Ⅱ分离,以2 ml/min流速用氯仿:甲醇:石油醚(1:2:1)洗脱,监测280 nm处紫外吸收以收集馏分。对馏分F_(7～10)浓缩、纯化与重结晶,经UV和MS分析并与茶儿茶素标准相对照,F_7为(一)-表儿茶素(EC)和(一)-儿茶素(C), F_8为(一)-表没食子儿茶素(EGC), F_9为(一)-表儿茶素没食子酸酯ECG,F_10为(一)-表没食子儿茶素没食子酸酯(EGCG);HPLC测定其纯度,F_7为17％C和83％EC,F_8为99％以上EGC,F_9为92.5％ECG,F_10为93.6％EGCG。茶叶中阻断N-亚硝化的主要有效成分为茶多酚(组分Ⅲ),尤为茶儿茶素混合物;茶儿茶素的阻断能力与其与NO_2~-摩尔比及本身的抗氧化能力有关。     

Acute effects of theanine,caffeine and theanine–caffeine combination on attention

Objective: l-theanine is a constituent of tea which is claimed to enhance cognitive functions. We aimed to determine whether theanine and theanine–caffeine combination have acute positive effects on cognitive and neurophysiological measures of attention, compared to caffeine (a positive control) and a placebo in healthy individuals.

Design: In a placebo-controlled, five-way crossover trial in 20 healthy male volunteers, we compared the effects of l-theanine (200?mg), caffeine (160?mg), their combination, black tea (one cup) and a placebo (distilled water) on cognitive (simple [SVRT] and recognition visual reaction time [RVRT]) and neurophysiological (event-related potentials [ERPs]) measures of attention. We also recorded visual (VEPs) and motor evoked potentials (MEPs) to examine any effects of treatments on peripheral visual and motor conduction, respectively.

Results: Mean RVRT was significantly improved by theanine (P?=?0.019), caffeine (P?=?0.043), and theanine–caffeine combination (P?=?0.001), but not by tea (P?=?0.429) or placebo (P?=?0.822). VEP or MEP latencies or SVRT did not show significant inter-treatment differences. Theanine (P?=?0.001) and caffeine (P?=?0.001) elicited significantly larger mean peak-to-peak N2-P300 ERP amplitudes than the placebo, whereas theanine–caffeine combination elicited a significantly larger mean N2-P300 amplitude than placebo (P?P?=?0.029) or caffeine (P?=?0.005). No significant theanine?×?caffeine interaction was observed for RVRT or N2-P300 amplitude.

Discussion: A dose of theanine equivalent of eight cups of back tea improves cognitive and neurophysiological measures of selective attention, to a degree that is comparable with that of caffeine. Theanine and caffeine seem to have additive effects on attention in high doses.     

EGCG和红茶多酚对HepG_2细胞脂质代谢相关基因表达的影响

目的:探讨表没食子儿茶素没食子酸酯(EGCG)和红茶多酚对人肝细胞癌HepG2细胞脂质代谢相关基因表达的影响。方法:HepG2细胞经EGCG(5μmol/L),红茶多酚(5μg/ml)或0.1%二甲亚枫(对照组)处理8h后,抽提RNA,并杂交至人14kcDNA芯片进行基因表达谱分析。应用实时RT-PCR技术对芯片结果进行验证。结果:HepG2细胞经EGCG和红茶多酚处理后表达差异的脂质代谢相关基因数分别为13条和29条,其中表达差异一致的基因有6条。实时RT-PCR结果与芯片结果一致。结论:EGCG和红茶多酚影响脂质代谢的机制是复杂的,此次实验发现的新靶标为今后茶多酚降脂作用研究提供了新的依据。     

EGCG乙酰化衍生物对肝癌细胞细胞毒增敏作用

目的探讨表没食子儿茶素没食子酸酯(epigallocatechin gallate,EGCG)乙酰化衍生物(AcEGCG)是否对人肝癌细胞SMMC-7721有细胞毒增敏作用。方法用四甲基偶氮噻唑蓝(MTT)法检测EGCG和AcEGCG分别联合柔红霉素(DNR)对人肝癌细胞SMMC-7721的毒性作用,实验设35、100、160μmol/L EGCG及AcEGCG组、35、100、160μmol/L EGCG及AcEGCG分别联合柔红霉素DNR(0.9μmol/L)组,金正均法计算q值,判断两药相互作用。结果 35、100μmol/L AcEGCG联合DNR对人肝癌细胞SMMC-7721的细胞毒增敏作用较相同剂量的EGCG强(67.0%对48.1%,78.9%对65.0%),差异具有统计学意义(P<0.05),160μmol/L AcEGCG联合DNR对人肝癌细胞SMMC-7721的细胞毒增敏作用比EGCG稍弱(P>0.05)。结论在一定剂量范围内,AcEGCG联合DNR对人肝癌细胞SMMC-7721的细胞毒增敏作用较EGCG强。     

表没食子儿茶素没食子酸酯对丙烯酰胺致小脑颗粒神经元氧化损伤的防护作用研究

目的探讨表没食子儿茶素没食子酸酯(EGCG)对丙烯酰胺(ACR)致小脑颗粒神经元(CGNs)损伤的保护作用。方法采用体外细胞培养的方法,建立ACR损伤CGNs的体外模型。经不同浓度EGCG(5、10、25、50、100 mol/L)预处理24h后,加入ACR(5 mmol/L)染毒,24 h后用四甲基偶氮唑盐(MTT)比色法检测细胞存活率,倒置显微镜下观察细胞形态学变化,测定细胞内超氧化物歧化酶(SOD)活性及丙二醛(MDA)的含量,Hochest 33342染色观察凋亡时细胞核形态的变化,末端脱氧核苷酸转移酶介导的原位缺口末端标记(TUNEL)法检测细胞凋亡指数。结果同ACR损伤组比较,终浓度为10、25、50 mol/L的EGCG能减轻ACR引起的CGNs损伤,可明显提高细胞的存活率(P<0.05),SOD活性增高,MDA含量降低(P<0.05),Hoechst33342染色和TUNEL荧光标记显示,细胞核固缩、致密浓染现象较ACR损伤模型组改善明显,凋亡指数下降(P<0.05),且呈现剂量依赖趋势,而终浓度为5、100 mol/L EGCG组较正常组无明显变化。结论 EGCG在一定剂量范围内对ACR氧化损伤的CGNs有防护作用。[营养学报,2012,34(4):362-367]     

The Effects of Epigallocatechin-3-Gallate on Thermogenesis and Fat Oxidation in Obese Men: A Pilot Study

Objectives: The development of obesity is characterized by an increase in adipose tissue mass and by concomitant and profound changes in almost all organ functions leading to diseases such as hypertension, diabetes mellitus and coronary heart disease. Recent data from human studies indicate that the consumption of green tea and green tea extracts may help reduce body weight, mainly body fat, by increasing postprandial thermogenesis and fat oxidation. However, human studies investigating the metabolic effects of the most predominant tea catechin, EGCG, alone are absent.

Methods: In a randomized double blind, placebo-controlled, cross-over pilot study, six overweight men were given 300 mg EGCG/d for 2d. Fasting and postprandial changes in energy expenditure (EE) and substrate oxidation were assessed.

Results: Resting EE did not differ significantly between EGCG and placebo treatments, although during the first postprandial monitoring phase, respiratory quotient (RQ) values were significantly lower with EGCG compared to the placebo.

Conclusions: These findings suggest that EGCG alone has the potential to increase fat oxidation in men and may thereby contribute to the anti-obesity effects of green tea. However, more studies with a greater sample size and a broader range of age and BMI are needed to define the optimum dose.     

液相色谱-串联质谱法同时测定饮料中的咖啡因、茶碱和可可碱

目的建立市售饮料中咖啡因、茶碱和可可碱的液相色谱-串联质谱法(LC-MS/MS)。方法将饮料用水稀释100倍后,采用三氯甲烷∶异丙醇(9∶1,V/V)进行提取,提取液经Allure PFP Propyl柱(100 mm×2.1 mm,5μm)分离,以5 mmol/L乙酸铵溶液(含5%甲醇、0.05%的甲酸)-甲醇为流动相进行梯度洗脱,采用电喷雾正离子化(ESI+)、多反应监测模式同时测定咖啡因、茶碱和可可碱。结果咖啡因、茶碱和可可碱在10 ng/ml^1000 ng/ml范围内方法的线性相关系数(r)均>0.9970,检出限为2 ng/ml^5 ng/ml,加标回收率均>60.68%,精密度均<14.41%。应用于实际市售饮料中,均检出咖啡因成分,部分饮品检出茶碱、可可碱成分。结论该方法简便、灵敏,可用于监测市售饮料中咖啡因、茶碱、可可碱的含量。