Mobile Phase Of Acetonitrile-water Research Articles

Simultaneous Analysis of Thirteen Compounds in Yeokwisan Using High-Performance Liquid Chromatography-Photodiode Array Detection and Ultra-Performance Liquid Chromatography-Tandem Mass Spectrometry and Their Antioxidant Effects.

Yeokwisan (YWS) is an herbal medicine prescription consisting of six oriental herbal medicines, developed to treat reflux esophagitis. We focused on developing an analytical method capable of simultaneously quantifying 13 compounds in YWS samples using high-performance liquid chromatography-photodiode array detection (HPLC-PDA) and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and exploring their antioxidant effects. All compounds examined in both analytical systems were chromatographically separated on a SunFireTM C18 (4.6 × 250 mm, 5 μm) column and an Acquity UPLC BEH C18 (2.1 × 100 mm, 1.7 μm) column using gradient elution of a water-acetonitrile mobile phase. Antioxidant effects were evaluated based on radical scavenging activity (DPPH and ABTS tests) and ferrous ion chelating activity. In two analytical methods, the coefficient of determination of the regression equation was ≥0.9965, the recovery range was 81.11-108.21% (relative standard deviation (RSD) ≤ 9.33%), and the precision was RSD ≤ 11.10%. Application of the optimized analysis conditions gave quantitative analysis results for YWS samples of 0.02-100.36 mg/g. Evaluation of the antioxidant effects revealed that baicalein and baicalin exhibit significant antioxidant activity, suggesting that they play an important role in the antioxidant effects of YWS.

Chiral Hydroxy Metabolite of Mebendazole: Analytical and Semi-Preparative High-Performance Liquid Chromatography Resolution and Chiroptical Properties.

Mebendazole (MBZ) is a benzimidazole carbamate anthelmintic used worldwide for the treatment and prevention of parasitic disorders in animals and humans. A large number of in vivo and in vitro studies have demonstrated that MBZ also has anticancer activity in multiple types of cancers. After oral administration, the phenylketone moiety of MBZ is rapidly reduced to the hydroxyl group to form the chiral hydroxy metabolite (MBZ-OH). To the best of our knowledge, there is no information in the literature on the stereochemical course of transformation and the anthelmintic and antitumor activity of individual enantiomers of MBZ-OH. In the present study, we describe in detail the direct HPLC resolution of MBZ-OH on a 100 mm × 4.6 mm Chirapak IG-3 column packed with 3 μm silica particles containing amylose (3-chloro-5-methylphenylcarbamate) as a selector. At 25 °C and using pure methanol as the mobile phase, the enantioseparation and resolution factors were 2.38 and 6.13, respectively. These conditions were scaled up at a semi-preparative scale using a 250 mm × 10 mm Chiralpak IG column to isolate multi-milligram amounts of both enantiomeric forms of the chiral metabolite. The chiroptical properties of the collected enantiomers were determined and, through a theoretical study, were related to the more stable conformations of MBZ-OH. The first and second eluted enantiomers were dextrorotatory and levorotatory, respectively, in dimethylformamide solution. Finally, by recording the retention factors of the enantiomers as the water content in the water-acetonitrile mobile phases was progressively varied, U-shaped retention maps were generated, indicating a dual and competitive hydrophilic interaction liquid chromatography and reversed-phase liquid chromatography retention mechanism on the Chirapak IG-3 chiral stationary phase.

Determination of the hydrogen-bond basicity descriptor by reversed-phase liquid chromatography

Except for alkanes, most organic compounds are hydrogen-bond bases. The B° descriptor of the solvation parameter model provides a convenient measure of the effective (or summation) hydrogen-bond basicity of organic compounds. A fast and convenient method to assign the B° descriptor is required to support studies of hydrogen-bonding in separation systems. A two-column system with acetonitrile-water mobile phase compositions and the measurement of up to eleven isocratic retention factors is proposed for this purpose. Several reversed-phase column chemistries and mobile phases were evaluated with the two-column system consisting of a pentafluorophenylpropylsiloxane-bonded and octadecylsiloxane-bonded silica columns recommended for this purpose. To assess the accuracy of the method values for B° were taken from the Wayne State University (WSU) compound descriptor database, which were assigned using conventional multi-technique methods and large datasets. The two-column systems provided an unbiased assignment of B° with an average deviation of 0.008 and an average absolute deviation of 0.021 compared with the target value for 55 varied compounds. The two-column system is unsuitable for assigning the other descriptors used in the solvation parameter model and results in erroneous assignments of B° for nitrogen-containing compounds capable of electrostatic interactions on silica-based reversed-phase columns.

An improved ultra-high performance liquid chromatography-tandem mass spectrometry method determining hispidulin and homoplantaginin in rat plasma and associated pharmacokinetic studies

AbstractFlavonoids are the most abundant components in Salvia plebeia, with significant pharmacological antioxidant and hepatoprotective properties. Hispidulin and homoplantaginin are the main flavonoid components in S. Plebeia. In this study, we established an ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to determine hispidulin and homoplantaginin in rat plasma samples, which were precipitated using acetonitrile-methanol (9:1, v/v). We used a UPLC HSS T3 (100 mm × 2.1 mm, 1.7 μm diameter) chromatographic column, an acetonitrile-water (containing 0.1% formic acid) mobile phase, and a gradient elution flow rate of 0.4 mL min−1 in an elution time of 4 min. We used electrospray ionization (ESI) detection in positive ion mode, and multiple reaction monitoring mode (MRM) for quantitative analysis: m/z 301 → 286 for hispidulin, m/z 463 → 301 for homoplantaginin, and m/z 465 → 303 for internal standard (IS). In pharmacokinetic studies, 24 rats were orally administered hispidulin and homoplantaginin (5 mg kg−1) and received sublingual intravenous injections (1 mg kg−1) at two different doses, four groups with six rats/group. Differences in hispidulin and homoplantaginin pharmacokinetics in rat plasma were evaluated. The calibration curve showed good linearity in the 0.5–1,000 ng mL−1 range, with r > 0.99. Precision, accuracy, recovery, matrix effects, and stability results all met standard biological sample detection requirements. Our pharmacokinetic studies showed hispidulin bioavailability was much higher than homoplantaginin at 17.8% and 0.1%, respectively.

DEVELOPMENT AND VALIDATION OF A HPLC METHOD FOR THE QUANTIFICATION OF BENZOYL PEROXIDE IN FLOUR

An isocratic high performance liquid chromatographic method has been developed and validated for the estimation of benzoyl peroxide in flours.The benzoyl peroxide was extracted with acetonitrile and analysed by Waters HPLC instrument on a C18 column using water-acetonitrile mobile phase with a flow rate of 1.5 ml/min and UV detection at 235 nm.The linearity was found in the range of 5-50 mg/l.The method is simple,reliable, specific,accurate and precise. KEY WORDS: Benzoyl peroxide, Acetonitrile, HPLC, Method Development,Method Validation,.

Development of a UPLC-ESI-MS/MS method for the determination of triamcinolone acetonide in human plasma and evaluation of its bioequivalence after a single intramuscular injection in healthy volunteers.

Introduction: Triamcinolone acetonide (TA) is commonly used in the treatment of various inflammatory conditions. To ensure its efficacy and safety, it is important to accurately determine its concentration in human plasma and evaluate its bioequivalence. In this study, an efficient ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) method was developed for the quantification of TA in human plasma after a single intramuscular injection. The internal standard used in this method was cortisone acetate (CA). Methods: TA and CA were extracted from plasma using ethyl acetate and N-hexane (4:1, v/v), separated on a C18 reverse-phase column with a mobile phase of acetonitrile-water containing 1% formic acid (55:45, v/v), and analyzed by UPLC-ESI-MS/MS. Multiple-reaction monitoring was performed using the transitions m/z 435.4→397.3 for TA and m/z 403.4→163.1 for CA. Results: The developed UPLC-ESI-MS/MS method demonstrated linearity over a concentration range of 0.53-21.20ng/mL, with a lower limit of quantification of 0.53ng/mL. The intra- and inter-run precision values ranged from 3.007% to 9.960% and 3.528% to 11.26%, respectively. The intra- and inter-run accuracy ranges were -1.962% to -6.577% and -3.371% to 0.348%, respectively. The matrix effect, extraction recovery, and stability of TA all met the acceptance criteria recommended by the National Medical Products Administration (NMPA) for bioassays. In healthy volunteers who received a single intramuscular injection of 80mg of either the test or reference formulation of TA, various pharmacokinetic parameters were determined. C max was found to be 8.616 ± 1.232 and 8.285 ± 1.218ng/mL for the test and reference formulations, respectively. T max was approximately 1.833 ± 0.243 and 1.861 ± 0.230h. The t 1/2 was calculated to be 181.249 ± 78.585 and 201.782 ± 83.551h. AUC 0-720 was 835.642 ± 297.209 and 830.684 ± 331.168ngh/mL, AUC 0-∞ was 991.859 ± 355.939 and 1018.665 ± 420.769ngh/mL for the test and reference formulations, respectively. The average relative bioavailability of TA, determined using AUC 0-720, was 105.4 ± 26.9%. Bioequivalence was evaluated through variance analysis and a double unilateral test, and the 90% confidence intervals of AUC 0-720, C max, and AUC 0-∞ were 92.8%-113.4%, 99.1%-109.1%, and 89.7%-110.9%, respectively (all p > 0.05). Discussion: These results met the bioequivalence criteria set by the NMPA, indicating that the developed UPLC-ESI-MS/MS method accurately determined TA concentrations in the plasma of healthy Chinese volunteers and that the test and reference formulations exhibited bioequivalence in these individuals.

Preparation of polyvinyl pyrrolidone stationary phase and its application in separation and analysis of flavonoids

AbstractPolyvinylpyrrolidone (PVP) is known as a polymer with special adsorption properties for polyphenolic compounds. In this paper, chemical‐bonded stationary phase (PVP@Silica) was prepared and applied to the separation of flavonoids. System constants determined by linear solvation energy relationships with methanol–water mixtures as the mobile show that the column can provide stronger hydrogen bonding and π–π interaction than C18 column. Under reversed‐phase conditions, traditional flavonoids have much longer retention on PVP@Silica than on C18 column with methanol–water mobile phase due to multimodal retention mechanism, including hydrogen bonding and π–π interaction. In addition, a U‐shaped retention curve was observed in acetonitrile–water mobile phase because of the enhanced hydrogen bonding under the high proportion of acetonitrile. The selectivity to polyhydroxy structure gives the stationary phase unique separation ability for flavonoids. The separation orthogonality was further investigated by a sample set containing 33 flavonoids with different substitution structures. The (RP‐PVP@Silica)‐(RP‐C18) system exhibited 60% orthogonality metric (OM) for these flavonoids with methanol–water mobile phase. A high OM of 63.5% was achieved in (RP‐PVP@Silica)‐(HILIC‐PVP@Silica) system. Finally, PVP@Silica was applied to the purification of total flavonoids in Ginkgo biloba extract (GBE). The offline (RP‐PVP@Silica)‐(RP‐C18) two‐dimensional separation system was used for the analysis of flavonoids in GBE.

Quantitative Analysis of Eight Compounds in Traditional Korean Medicine, Gongjindan Using HPLC, UPLC–MS/MS, and GC–MS/MS Systems

Gongjindan (GJD) is a traditional Korean medicine consisting of four herbal medicines and two animal-derived medicines, and is taken as a tonic in Republic of Korea. In this study, the goal was to develop and validate a simultaneous analytical method to quantify eight compounds in commercially available GJD samples using high-performance liquid chromatography (HPLC), ultra-performance liquid chromatography with tandem mass spectrometry (UPLC–MS/MS), and gas chromatography with tandem mass spectrometry (GC–MS/MS) systems. In HPLC and UPLC–MS/MS, seven components (gallic acid, 5-(hydroxymethyl)furfural, morroniside, loganin, nodakenin, decursin, and decursinol angelate) were separated and quantified using a distilled water–acetonitrile mobile phase system on a Capcell Pak UG80 C18 column and an Acquity UPLC BEH C18 column, respectively. Muscone was quantified using GC–MS/MS. The developed assays were validated by evaluating the linearity, limit of detection, limit of quantitation, recovery, and precision. In the regression equations of all compounds, the coefficient of determination was ≥0.9917, showing good linearity. The recovery was 93.70–108.17%, and the relative standard deviation values in the precision test were all <1.50%. Using the developed analysis methods, GJD samples were determined to contain the eight target compounds in concentrations from non-detected to 10.75 mg/g. The analytical assays developed and validated in this study can be used to obtain data for the quality control of commercially available GJDs and for the further expansion of efficacy and clinical studies.

Relationships of Enantioselective Retention of Chiral Oxazolopyrroloquinolones on a Stationary Phase with Grafted Ristocetin А Antibiotic from Aqueous Methanol Solutions

The mechanisms of retention and separation of the enantiomers of chiral oxazolopyrroloquinolones on a stationary phase with graftеd macrocyclic antibiotic ristocetin A under conditions of high-performance liquid chromatography with aqueous methanol mobile phases have been studied. The retention factor was found to decrease monotonically when the CH3OH concentration in the mobile phase increased to 90 vol %; then it slightly increased as the composition of the mobile phase approached pure methanol. The thermodynamics of adsorption of oxazolopyrroloquinolones in different regions of this dependence was studied. A mathematical model has been proposed; it describes well the experimental data and allows for analyte solvation in the mobile phase and competitive adsorption (with mobile phase components) on a chiral selector. The results were compared with previously obtained data for water–acetonitrile mobile phases. The reasons for the observed differences were discussed.

Simultaneous determination of six triterpenoid saponins in beagle dog plasma by UPLC-MS/MS and its application to a pharmacokinetic study after oral administration of the extract of the Eleutherococcus senticosus (Rupr. & Maxim.) Maxim. leaves

AbstractA rapid and simple ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) method was developed and validated for simultaneous determination of six analytes from the Eleutherococcus senticosus (Rupr. & Maxim.) Maxim. leaves (ESL) in beagle dog plasma for the first time, including 3-O-α-l-rhamnopyranosyl-(1→2)-α-l-arabinopyranoside-29-hydroxy oleanolic acid, 3-O-β-d-glucopyranosyl-(1→2)-α-l-arabinopyranoside-29-hydroxy oleanolic acid, 3-O-β-d-glucopyranosyl-(1→2)-α-l-arabinopyranosyl-30-norlean-12,20 (29) –dien-28-olic acid, ciwujianoside E, guaianin N, and eleutheroside K. The chromatographic separation was performed using an ACQUITY UPLC BEH C18 column (2.1 × 100 mm, 1.7 μm) using a gradient elution way with a mobile phase of acetonitrile-water containing 0.1% formic acid. Analytes were detected on a triple-quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source with multiple reaction monitoring (MRM) mode. Calibration curves were all linear (r ≥ 0.9933) over the concentration range. The mean extraction recoveries and matrix effect of analytes and I.S. were ranged from 80.26% to 98.32% and from 91.27% to 111.67%, respectively. The intra-day and inter-day precision were ranged from 2.20% to 14.81%, and the accuracy range was 1.60–14.60%. The analytical method was successfully applied for the pharmacokinetic characteristics of the six analytes in beagle plasma after oral administration of ESL extracts. The T1/2 of six analytes was more than 3.09 ± 0.78 h.

High performance liquid chromatographic Assessment of berberine in Methanol Extract of diverse parts of Tinospora cordifolia

In the current work, a straight forward, affordable, reliable, repeatable, selective, and accurate high performance liquid chromatography (HPLC) approach for berberine measurement in both a 60% methanolic extract of Tinospora cordifolia and a commercial formulation was developed and validated. The berberine peak, which has a retention time of 9.52±1.03 min, is crisp and well-defined in the acetonitrile-water (25:75%, v/v) mobile phase. Berberine is estimated using HPLC at a wavelength of 266 nm and a flow rate of 1.0 ml/min. The calibration curve’s results from linear regression analysis demonstrates a strong linear association with a correlation coefficient of 0.997 in the concentration range of 0.5 to 50 g/ml. y=29716x- 4417.4 is the equation for the linear regression. The thresholds for quantification and detection are 0.18 and 0.55 g/ml, respectively. According to ICH criteria, the proposed technique has been verified for accuracy, precision, repeatability, and robustness. For the purpose of estimating berberine in methanolic extract and formulation, the proposed approach is used, which has a high degree of precision and accuracy. The approach is exact, repeatable, selective, and accurate for estimating berberine, according to statistical analysis.

Quality evaluation of Ginseng Radix et Rhizoma based on UPLC characteristic chromatogram and quantitative analysis of multi-components by single marker (QAMS)

Based on UPLC characteristic chromatogram and quantitative analysis of multi-components by single marker(QAMS), the content of seven types of ginsenosides in Ginseng Radix et Rhizoma was simultaneously determined, and the quality of Ginseng Radix et Rhizoma was evaluated by the principal component analysis(PCA). The chromatographic separation was performed on the Acquity UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 μm) with the mobile phase of acetonitrile-water for gradient elution at the flow rate of 0.3 mL·min~(-1), the column temperature of 30 ℃, the detection wavelength of 203 nm, and the injection volume of 2 μL. The UPLC chromatogram was established with 19 batches of Ginseng Radix et Rhizoma samples from three producing areas by Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine(version 2012). Thirteen characteristic peaks were determined and seven components were identified. SPSS 26.0 was used to conduct PCA on the characteristic peak areas. With the peak of ginsenoside Rb_1 as reference peak S, ginsenoside Rb_1 showed good durability of relative correction factor as compared with other ginsenosides. The QAMS method for the determination of seven ginsenosides in Ginseng Radix et Rhizoma was established. There was no significant difference in results between the QAMS method and the external standard method. As revealed by the results of PCA and the determination of the total content of seven ginsenosides, the four batches of Ginseng Radix et Rhizoma numbered S19, S18, S1, and S2 were of superior quality. The characteristic chromatogram and QAMS method for the determination of seven ginsenosides in this study were convenient and accurate, which greatly shortened the analysis time and improved the analysis efficiency. The findings of this study are expected to provide a basis for the overall quality evaluation of Ginseng Radix et Rhizoma.

In Vitro and In Vivo Genotoxicity Assessments and Phytochemical Analysis of the Traditional Herbal Prescription Siryung-Tang.

Siryung-tang (SRT) is a traditional herbal prescription containing Oryeong-san and Soshiho-tang that is used to treat digestive system diseases. We performed safety evaluations of SRT based on genotoxicity and developed an assay for quality control using high-performance liquid chromatography with a photodiode array detector. Genotoxicity was evaluated based on bacterial reverse mutation (Salmonella typhimurium TA1535, TA98, TA100, and TA1537, and Escherichia coli WP2 uvrA), chromosomal aberration (Chinese hamster lung cells), and micronucleus (mouse) tests. Quality control analysis was conducted using a SunFire C18 column and gradient elution with a distilled water–acetonitrile mobile phase system containing 0.1% (v/v) formic acid for 12 markers (5-(hydroxy-methyl)furfural, 3,4-dihydroxybenzaldehyde, liquiritin apioside, liquiritin, coumarin, baicalin, wogonoside, cinnamaldehyde, baicalein, glycyrrhizin, wogonin, and atractylenolide III). SRT showed no genotoxicity in three tests. Ames tests showed that SRT at 313–5000 μg/plate did not significantly increase the number of revertant colonies with or without metabolic activation among five bacterial strains. Moreover, in vivo micronucleus testing showed that SRT did not increase the frequency of bone marrow micronuclei. The number of chromosomal aberrations associated with SRT was similar to that observed in the negative controls. The 12 markers were detected at 0.04–16.86 mg/g in a freeze-dried SRT sample and completely eluted within 45 min. The extraction recovery was 95.39–104.319% and the relative standard deviation value of the precision was ≤2.09%. Our study will be used as basic data for the safety and standardization of SRT.

Development of a Simple, Underivatized Method for Rapid Determination of Free Amino Acids in Honey Using Dilute-and-Shoot Strategy and Liquid Chromatography-Tandem Mass Spectrometry.

A simple, fast and reliable analytical method was developed for 20 free amino acids (FAAs) determination in honey samples through a dilute-and-shoot strategy and hydrophilic interaction liquid chromatography tandem mass spectrometry. Compared with previous reports, direct dilution by water has significantly reduced the matrix effect and facilitated full extraction of FAAs. Further, a 5 min determination method was established with an acetonitrile–water mobile phase system with 0.1% formic acid addition. The established method was validated and demonstrated several advantages including short detection time, wide linear range over 3–4 orders of magnitude, high sensitivity down to 0.1 ng/mL and negligible matrix effect. Twenty FAAs were determined in 10 honey samples from different botanical origins by this method, and 19 FAAs were found. This general applicable method was also promising for fast determination of FAAs in other practical samples.

Adsorption, separation, and purification of cyclosporine A using reversed-phase liquid chromatography

环孢素A(cyclosporine A, CsA)是由11个氨基酸组成的中性环状多肽,是临床拮抗器官和组织移植后排异反应的首选药物。高效液相色谱法广泛应用于CsA的分离分析,开展CsA色谱行为的研究是使用制备高效液相色谱纯化CsA的关键。该文首先在C18固定相上比较了CsA在甲醇-水和乙腈-水两种流动相体系中的保留行为,结果表明其保留时间对有机相比例变化比较敏感。控制甲醇比例在84%~88%,或者乙腈比例在75%~85%, CsA的保留因子(k)在3~7范围内。考察了上样量对CsA峰形的影响。随着上样量增加,在两种流动相体系中,CsA的峰形都由对称开始变得拖尾,保留时间前移。因此在进行CsA纯化时,需要特别注意前杂的去除情况。然后采用吸附等温线描述CsA的保留行为,当流动相中CsA的质量浓度较低时,有机相比例对溶质在固定相上的吸附量影响并不明显。随着溶质的质量浓度增加至0.5 g/L以上,有机相比例降低有助于提高CsA在固定相上的吸附量。和甲醇-水体系相比,在乙腈-水体系中固定相对溶质有更大的吸附容量。用模型对CsA的等温吸附曲线拟合,在甲醇-水体系中符合Langmuir模型,在乙腈-水体系中为Moreau模型。由模型参数可知在两种体系下,CsA在C18固定相上均为单层吸附,区别在于乙腈-水体系中CsA会产生较大的分子间作用力。最后,实验采用0~60 min 65%~75%乙腈、60~80 min 75%乙腈的条件开展了环孢素A纯化的探索实验,可将CsA的杂质控制在0.2%以下。本研究结果对采用制备高效液相色谱纯化CsA具有指导意义。

Quantitation of Ciprofloxacin by RP-HPLC from Active and Dosage Formulations

A simple, sensitive isocratic reversed-phase HPLC method for detecting and quantifying ciprofloxacin in pharmaceutical dosage forms has been developed and validated according to ICH guidelines. In the method, the separation was achieved on a C18 reversed-phase column with a mobile phase acetonitrile-water at a ratio of (80:20). The pH was adjusted to 2.7 using 85% of phosphoric acid with ultraviolet detection at 275.0 nm. A flow rate of 1.7 mL/minute was used at room temperature. The method'

DEVELOPMENT OF A RAPID AND EFFICIENT METHOD FOR QUANTITATIVE DETERMINATION OF N-NITROSODIMETHYLAMINE IMPURITY BY HIGH-PERFORMANCE LIQUID CHROMATOGRAPHY IN METFORMIN HYDROCHLORIDE, LOSARTAN POTASSIUM, VALSARTAN AND RANITIDINE MEDICINAL RAW MATERIALS AND ITS PRODUCTS

In this study, chromatographic conditions were developed for the separation and quantification of N-nitrosodimethylamine (NDMA) by HPLC. Our method would be useful for the rapid screening and quantification of NDMA impurity in Metformin hydrochloride, Losartan potassium, Valsartan and Ranitidine drugs substance and its products. The chromatographic separation was achieved on a Phenomenex C18 column (250 mm, 5 µm, 4.6 mm) by multistep gradient elution, using a water-acetonitrile mobile phase containing 0.1% formic acid, which provided better c hromatographic separation. Column temperature was maintained at 30℃; mobile phase flow rate: 1 mL/min; wavelength: 254 nm; and injection volume of 20 μL.

Volume and composition of semi-adsorbed stationary phases in hydrophilic interaction liquid chromatography. Comparison of water adsorption in common stationary phases and eluents

Pycnometric and homologous series retention methods are used to determine the volume and mean composition of the water-rich layers partially adsorbed on the surface of several hydrophilic interaction liquid chromatography (HILIC) column fillings with acetonitrile-water and methanol-water as eluents. The findings obtained in this work confirm earlier studies using direct methods for measuring the stationary phase water content performed by Jandera's and Irgum's research groups. Water is preferentially adsorbed on the surface of the HILIC bonded phase in hydroorganic eluents containing more than 40% acetonitrile or 70% methanol, and a gradient of several water-rich transition layers between the polar bonded phase and the poorly polar bulk mobile phase is formed. These layers of reduced mobility act as HILIC stationary phases, retaining polar solutes. The volume of these layers and concentration of adsorbed water is much larger for acetonitrile-water than for methanol-water mobile phases.In hydroorganic eluents with less than 20-30% acetonitrile or 40% methanol the amount of preferentially adsorbed water is very small, and the observed retention behavior is close to the one in reversed-phase liquid chromatography (RPLC). In eluents with intermediate acetonitrile-water or methanol-water compositions a mixed HILIC-RPLC behavior is presented.Comparison of several HILIC columns shows that the highest water enrichment in the HILIC retention region for acetonitrile-water mobile phases is observed for zwitterionic and aminopropyl bonded phases, followed in minor grade for diol and polyvinyl alcohol functionalizations. Pentafluorophenyl bonded phase, usually considered a HILIC column, does not show significant water adsorption, nor HILIC retention.

Molecular identification and alkaloids content of endangered Chinese medicinal plant Coptis chinensis var. brevisepala

According to the records of Chinese materia medica,Coptis chinensis var. brevisepala is an authentic Chinese medicinal plant highly recommended by ancient physicians since its rhizome is like a string of beads and has a good medicinal value. However,its medicinal components and values remain to be studied as it is endangered because of overexploitation. Therefore,this study aims to quantitatively determine its effective components based on UPLC-QTOF-MS,and to compare the contents of isoquinoline alkaloids in C.chinensis var. brevisepala with those in other Coptis species. Meanwhile,molecular methods accurately identified 12 batches of C. chinensis var. brevisepala,9 batches of C. chinensis,4 batches of C. deltoidea,and 1 batch of C. teeta. Gradient elution was performed with Waters CORTECS C18 column( 4. 6 mm× 150 mm,2. 7 μm) and the mobile phase acetonitrile-water with 0. 4% formic acid. Mass spectrometry was conducted in ESI positive mode. The quantitative results showed that 8 main alkaloids had a good linear relationship within the concentration range( R~2>0. 996),with the recovery rate of 95. 18%-105. 0% and the RSD of 0. 28%-3. 7%. Compared with that of other Coptis species,the rhizome of C. chinensis var. brevisepala had the highest contents of berberine and columbamine. The total content of the 8 alkaloids in C. chinensis var. brevisepala was similar to that in C. chinensis but higher than that of the other two species. PCA was performed to compare the alkaloids among the 4 species. Besides,the 8 alkaloids were evaluated in different parts of C. chinensis var. brevisepala. The results indicate that this method is reliable and efficient and can provide a reference for the quality research.

Simultaneous Analysis to Evaluate the Quality of Insamyangpye–Tang Using High-Performance Liquid Chromatography–Photo Diode Array Detection

Insamyangpye–tang (ISYPT) is a traditional medicinal formula comprised of 13 herbs and has been used in East Asia to treat lung-related diseases. However, to our knowledge, no method of analysis for its quality control has been reported. In this study, a method of analysis for quality control of ISYPT was developed using high-performance liquid chromatography. Chromatographic separation, analysis, and assay verification were performed with a distilled water–acetonitrile mobile phase system, both containing 0.1% (v/v) trifluoroacetic acid, and a Gemini C18 analytical column (4.6 mm × 250 mm, 5 μm) using authentic standards for eight marker compounds. These marker constituents were detected simultaneously at 0.09–5.95 mg/g. The analysis method developed can be used for basic quality control of ISYPT.