A Sensitive Reversed-Phase High-Performance Liquid Chromatography Method for the Quantitative Determination of Milk Xanthine Oxidase Activity

A new reversed-phase high performance liquid chromatography method was developed to quantitate the activity of xanthine oxidase involved in milk fat globule membrane with xanthine as the substrate and the separation of product (uric acid). The increment of uric acid in the reaction system was used to calculate the total activity of XO. The optimized assay conditions, linearity of detection, recovery of uric acid and chromatogram were developed in text, indicating this method is simple, rapid and efficient. It is an alternative potential method for the determination of the activity of XO in milk.

INTERSECTION POINT RULE OF THE RETENTION VALUE AND NORMAL BOILING POINT OF THE HOMOLOGUES IN REVERSED-PHASE HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

In this paper, a linear relationship between the logarithm of capacity factor k and normal boiling point to of the homologues has been derived, based on the basic retention equation of liquid chromatography according to statistical thermodyoamics proposed by professor Ln Peizhang and others, This equation has been verified by a large number of experimental data, all the strsight lines of lnk- of bumologues for different mobile phass coaiposltion cross each other at the same point, So the intereection point equation van proposed, wbich was used to prodict the retention valu, the result was satisfactory.

INTERSECTION POINT RULE OF THE RETENTION VALUE AND MOBILE PHASE COMPOSITION OF THE HOMOLOGUES IN REVERSED-PHASE HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

Based on the intersection point rule of the retention value and normal boiling point of homologues in reversed-phase high-performance liquid chromatography(RPLC), the intersection point rule of the retention value of homologues and mobile phase composition has been derived, and was testified by a lot of experimental data from the literature. With this newly proposed equation, we can use the retention value of the compound in one mobile phase composition to predict its retention value in any other mobile phase composition. For fourteen groups of homologues in five mobile phase compositions on five Kinds of columns, the overall average absolute error of 721 data sets is 2.8%.

Quantitative analysis by reversed-phase high-performance liquid chromatography and retinal neuroprotection after topical administration of moxonidine

AIM:To determine moxonidine in aqueous humor and iris-ciliary body by reversed-phase high performance liquid chromatography(RP-HPLC),and to evaluate the retinal neuroprotective effect after topical administration with moxonidine in a high intraocular pressure(IOP)model.METHODS:The eyes of albino rabbits were administered topically and ipsilaterally with 0.2%moxonidine.A RPHPLC method was employed for the identification and quantification of moxonidine between 2 and 480 min,which presented in the aqueous humor and iris-ciliary body.Flash electroretinography(F-ERG)amplitude and superoxide dismutase(SOD)level were measured between day 1 and day 15 after topical administration with moxonidine in a rabbit model of high IOP.Histological and ultrastructural observation underwent to analyze the changes of retinal morphology,the inner retinal layers(IRL)thickness,and retinal ganglion cell(RGC)counting.RESULTS:Moxonidine was detectable between 2 and 480 min after administration,and the peak concentration developed both in the two tissues at 30 min,0.51μg/m Lin aqueous humor and 1.03μg/g in iris-ciliary body.In comparison to control,F-ERG b-wave amplitude in moxonidine eyes were significantly differences between day 3 and day 15(P<0.01)in the high IOP model;SOD levels were significantly higher at all time-points(P<0.01)with a maximum level of 20.29 U/mgprot at day 15;and RGCs were significantly higher(P<0.05).CONCLUSION:Moxonidine is a viable neuroprotective agent with application to high IOP model.All layers of retina,including RGC layer,retinal nerve fiber layer and INL,are more preserved after moxonidine administration.SOD plays a neuroprotective role in ocular hypertension-mediated RGC death.

A liquid chromatography with tandem mass spectrometry method for quantitating total and unbound ceritinib in patient plasma and brain tumor

A rapid, sensitive, and robust reversed-phase liquid chromatography with tandem mass spectrometry method was developed and validated for the determination of total and unbound ceritinib, a secondgeneration ALK inhibitor, in patient plasma and brain tumor tissue samples. Sample preparation involved simple protein precipitation with acetonitrile. Chromatographic separation was achieved on a Waters ACQUITY UPLC BEH C_(18) column using a 4-min gradient elution consisting of mobile phase A(0.1% formic acid in water) and mobile phase B(0.1% formic acid in acetonitrile), at a flow rate of 0.4 m L/min. Ceritinib and the internal standard([^(13)C_6]ceritinib) were monitored using multiple reaction monitoring mode under positive electrospray ionization. The lower limit of quantitation(LLOQ) was 1 n M of ceritinib in plasma. The calibration curve was linear over ceritinib concentration range of 1–2000 n M in plasma. The intra-and interday precision and accuracy were within the generally accepted criteria for bioanalytical method( o15%).The method was successfully applied to assess ceritinib brain tumor penetration, as assessed by the unbound drug brain concentration to unbound drug plasma concentration ratio, in patients with brain tumors.

Comparison of liquid-liquid extraction-thin layer chromatography with solid-phase extraction-high-performance thin layer chromatography in detection of urinary morphine

Liquid-liquid extraction-thin layer chromatography （LLE-TLC） has been a common and routine combined method for detection of drugs in biological materials. Solid-phase extraction （SPE） is gradually replacing the tra- ditional LLE method. High performance thin layer chromatography （HPTLC） has several advantages over TLC. The present work studied the higher efficiency of a new SPE-HPTLC method over that of a routine LLE-TLC method, in extraction and detection of urinary morphine. Fifty-eight urine samples, primarily identified as mor- phine-positive samples by a strip test, ＇were re-screened by LLE-TLC and SPE-HPTLC. The results of LLE-TLC and SPE-HPTLC were then compared with each other. The results showed that the SPE-HPTLC detected 74% of total samples as morphine-positive samples whereas the LLE-TLC detected 48% of the same samples. We further discussed the effect of codeine abuse on TLC analysis of urinary morphine. Regarding the importance of morphine detection in urine, the present combined SPE-HPTLC method is suggested as a replacement method for detection of urinary morphine by many reference laboratories.

Ion-pair Reversed-phase×Low-pH Reversed-phase Two-dimensional Liquid Chromatography for In-depth Proteomic Profiling

High-resolution liquid chromatography separation is essential to in-depth proteomic profiling of complex biological samples.Herein,we established an ion-pair reversed-phase×reversed-phase two-dimensional liquid chromatography(IPRP×RP 2DLC)strategy for comprehensive proteomic analysis.Both RPLC separation dimensions were performed at low pH,with trifluoroacetic acid(TFA)and formic acid(FA)as mobile phase addictive,respectively.As the good separation resolution offered by ion-pairing effect of TFA,the fractionation efficiency was greatly improved with 74.0%peptides identified in just one fraction.Comparing with conventional high pH RP fractionation,the overall separation rate of IPRP was about 1.6 times that of high-pH RP,which increased the number of identified peptides by 21%.Further,2169 proteins and 8540 peptides were confidently identified from crude serum sample by our IPRP×RP 2DLC strategy,exhibiting great potential in clinical proteomics in the future.

建立HPLC-RID测定枸杞子中枸杞多糖含量研究

目的通过高效液相色谱-示差折光检测法(HPLC-RlD)摸索枸杞子中枸杞多糖(枸杞多糖以葡萄糖计)的提取方式和含量测定方法,达到检测方法准确可靠操作简便的目的。方法本法采取加水回流提取方式,利用高效液相色谱仪和示差折光检测器进行检测,色谱柱为LC026(Prevail Carbohyrate ES为填充剂250 mm×4.6 mm,5μm),流动相为乙腈-水(70∶30),检测器温度40℃,流速1.0 ml/min,柱温40℃。结果枸杞子在0.5~4.0 mg/ml线性范围内呈良好线性关系,R^(2)=0.9999,平均回收率为104.81%,RSD=1.10%。结论该方法操作简便,灵敏度高,重复性好,准确可靠,可用于测定药材枸杞子中枸杞多糖的含量,并有效控制枸杞子质量。

Preparation and Evaluation of Silicon Quantum Dots-Bonded Silica Stationary Phase for Reversed-Phase Chromatography

In this paper,silicon quantum dots(SiQDs)with green fluorescence are synthesized by solvothermal reaction of 3-(2,3-epoxypropoxy)propyltrimethoxysilane(GPTMS)and ethylenediaminetetraacetic acid(EDTA),and then SiQDs are bonded to the surface of silica to obtain a new nano-on-micro stationary phase(SiO_(2)-SiQDs)for reversed-phase chromatography.The successful preparation of SiO_(2)-SiQDs stationary phase is demonstrated by a variety of characterizations,such as transmission electron microscopy,laser confocal microscopy,elemental analysis and Fourier infrared spectroscopy.In addition,the chromatographic performance of the prepared stationary phase is evaluated and it shows good separation performance for non-polar substances such as alkylbenzene,aniline and polycyclic aromatic hydrocarbons in reversed-phase liquid chromatography.It is also verified that the stationary phase has good methyl selectivity and shape selectivity.More interestingly,the separation of prednisolone and hydrocortisone isomers can also be achieved at a low ratio of organic solvents,indicating that this new stationary phase has a good application prospect in isomer separation.

Predicting a, c indices in reversed-phase high-performance liquid chromatography (RP-HPLC) from Kovats index in gas chromatography (GC)

KOVATS index is the most precise index system reflecting the interaction between the molecules of solutes and stationary phase in gas chromatography at present. Large quantity of Kovats in dex data have been published. It is a good way to use Kováts index in gas chromatography to predict the retention value in liquid chromatography, which is significant in theory and apphcation.

PEAK IDENTIFICATION FROM INTERACTION INDEX IN REVERSED-PHASE HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY

The interaction index c which was derived from the fundamental retention equationlogk’ =a + cC_B in reversed--phase high--performancc liquid chromatography (RP-HPLC) quan-titatively describes the difference in the interaction between solute--strong. solvent andsolute--weak solvent; it has shown to be a constant for a specific solute even when columnsystems with different C18 packings are used. The theoretical basis for peak identificationby using interaction index has been proposed, which was based on the a,c values on stan-dard C18 column by utilizing linear a-a plots on column pairs and the linear relationship be-tween parameters a and c for the structural related compounds. Through the establishmentof parameters a,c data based on the standard C18 column for a certain type of compounds,the retention of thesc compounds on various C18 columns can be predicted. Typical exam-ples have been given to verify the correctness of this method.

高效液相色谱法测定食用植物油中黄曲霉毒素B_(1)前处理方法的改进

改进GB 5009.22—2016《食品安全国家标准食品中黄曲霉毒素B族和G族的测定》中高效液相色谱法测定食用植物油中的黄曲霉毒素B_(1)的方法,将甲醇-水溶液(体积比70∶30)用量提高至25 mL,选择均质提取4 min作为前处理提取方式,加入5.0 g氯化钠消除乳化现象,取消氮吹过程,净化洗脱完后直接定容。该优化方法提取效率高,较好地减少了被检测含量的损失,净化过程较原有方法更简单、方便,回收率和精密度更满意,适用于植物油脂黄曲霉毒素B_(1)的检测分析。

SPE-HPLC双检测器法同时测定地表水中的苯并[a]芘和阿特拉津含量及不确定度评定

采用全自动固相萃取(SPE)-高效液相色谱(HPLC)双检测器法同时定量分析地表水中的苯并[a]芘(BaP)和阿特拉津(AT)含量,建立了BaP和AT含量不确定度的分析方法。从样品前处理过程、样品重复测定、标准溶液配制和标准曲线拟合等方面评定了不确定度分量的贡献。评定结果显示:在影响检测结果的4个分量中,HPLC测定地表水中BaP含量时,标准曲线拟合和样品前处理过程引入的不确定度对结果影响较大,当BaP质量浓度为2.01 ng/L时,其扩展不确定度(U)为0.24 ng/L(k=2);测定AT含量时,标准溶液配制和样品重复测定引入的不确定度对结果影响较大,当AT质量浓度为50.09 ng/L时,U=4.72 ng/L(k=2)。

高效液相色谱法在中药复方成分分析中的应用

随着越来越多学者对中药复方制剂有效成分的深入挖掘,高效液相色谱法(HPLC)作为一种高效、准确、灵敏度高的分析手段受到越来越广泛的应用。目前,高效液相色谱法多与其他分析手段如质谱、红外光谱、荧光检测器等技术联用便于对中药复杂成分的分析。

基于不同内参物的一测多评法测定黄刺玫中5种成分的含量

建立基于不同内参物的一测多评法测定黄刺玫中矢车菊素-3-O-葡萄糖苷、芦丁、异槲皮苷、木犀草素和槲皮素5种成分的含量。分别以芦丁、异槲皮苷和木犀草素为内参物,计算另外4种成分的相对校正因子建立一测多评法,计算12批黄刺玫样品中5种成分的含量,并比较外标法和一测多评法测定结果之间的差异。结果表明外标法和一测多评法测定12批黄刺玫样品中5种成分的含量相对标准偏差均小于3.0%,表明两种方法的测定结果无明显差异。所建立的一测多评法同时测定黄刺玫中5种成分的含量与外标法结果无明显差异,可用于黄刺玫的质量控制和评价。

采用HPLC技术快速检测发酵大豆制品中异黄酮的含量

本文介绍了利用高效液相色谱(HPLC)技术快速检测发酵大豆制品中异黄酮含量的方法。在Lichrosper100RP-18(4.6mm×25cm×5μm)色谱柱上,以含有0.1%(v/v)冰醋酸的乙腈和水溶液为流动相进行梯度洗脱,检测波长为254nm,平均回收率达到99%以上。分析结果表明此分析方法灵敏、准确、重现性好。在上述条件下检测发酵大豆制品中异黄酮含量,结果表明发酵大豆制品中异黄酮主要以甙元形式存在。

川芎伍用丹参煎剂对川芎嗪药物动力学的影响

三维高效液相色谱（３Ｄ－ＨＰＬＣ）测定川芎煎剂及其灌胃大鼠后血清中均含有多种组分，其中一种用ＨＰＬＣ提纯后经紫外、红外光谱，质谱和核磁共振谱鉴定为川芎嗪（ＴＭＰ）。用ＨＰＬＣ法测定了大鼠灌胃川芎煎剂（Ｉ组，６只，３０ｇ／ｋｇ）和川芎丹参煎剂（Ⅱ组，６只，４０ｇ／ｋｇ，川芎：丹参＝３：１）后血清ＴＭＰ浓度，并研究了两组的药代动力学（药动学）。ＴＭＰ药动学参数中的一级吸收速度常数（Ｋａ）、与血药一时曲线下面积（ＡＵＣ）等值表明：Ⅱ组明显低于Ｉ组（Ｐ＜０．０５～０．０１），说明伍用丹参后引起ＴＭＰ吸收减慢和生物利用度降低。

二氧化硅-氧化石墨烯复合物固相萃取-高效液相色谱法检测植物油中黄曲霉毒素B_1、B_2

以二氧化硅-氧化石墨烯复合物为固相萃取材料,建立了植物油中黄曲霉毒素B1、B2的高效液相色谱（HPLC）检测方法。优化的条件为：复合材料的最佳用量为0.15 g,最佳萃取时间20 min,洗脱溶剂为乙腈,洗脱次数为2次。结果表明,在优化条件下,建立的二氧化硅-氧化石墨烯复合物固相萃取-高效液相色谱法对黄曲霉毒素B1、B2的检出限分别为0.17和0.05μg/L。将本方法应用于植物油实际样品的检测中,加标回收率在81.4%~105.3%之间,相对标准偏差为1.3%~8.6%。

分子印迹固相萃取-高效液相色谱法测定饲料中莱克多巴胺

以莱克多巴胺为模板分子,丙烯酰胺为功能单体,乙二醇二甲基丙烯酸酯为交联剂,合成了对莱克多巴胺具有高选择性的分子印迹聚合物。考察了甲醇、乙腈、丙酮和氯仿-甲醇与三乙胺构成致孔剂合成的聚合物性能及其形貌特征。通过正交试验优化的聚合反应配方为:1.0 mmol莱克多巴胺,4.0 mmol丙烯酰胺,20.0 mmol乙二醇二甲基丙烯酸酯,6.0 mL乙腈-三乙胺(30∶1,v/v),50.0 mg偶氮二异丁腈。建立的基于分子印迹固相萃取-高效液相色谱测定饲料试样中莱克多巴胺的方法,在0.50～100 mg/L质量浓度范围内有良好的线性关系(r=0.999 4);饲料试样中1.0、10及100 mg/kg 3个添加水平的莱克多巴胺平均回收率大于80%;批内、批间测定的相对标准偏差小于10%;检出限(信噪比为3)达到0.1 mg/kg。该方法灵敏、可靠,用于饲料等复杂基质中莱克多巴胺检测的效果优于相关标准分析方法。

高效液相色谱法测定银杏叶提取物中槲皮素的含量

采用反相高效液相色谱法分离银杏叶提取物中黄酮类化合物并测定其中槲皮素含量。ＹＷＧＣ１８（０４６ｃｍ×１５ｃｍ）色谱柱，流动相甲醇水磷酸（５０∶４９８∶０２），检测波长３６０ｎｍ。回收率在９５３％～１０３２％之间，ＲＳＤ为１８６％。