High-performance Liquid Chromatography-photodiode Array Detection Research Articles

Simultaneous determination of flavonoid and alkaloid compounds in Citrus herbs by high-performance liquid chromatography–photodiode array detection–electrospray mass spectrometry

The major active biological constituents in Citrus herbs are flavonoids, especially hesperidin, naringin and alkaloids, mainly synephrine, with beneficial medical effects on human health. They are used as the markers to control the quality of Citrus herbs. In this paper, a new ion pairing chromatographic method was developed to exclude the most polar solute (synephrine) from the viod volume and to maintain selectivity between the two other solutes (hesperidin and naringin). Perfluorinated carboxylic acids, which are appropriate for MS detection due to their volatility, were used as ion-pairing agents. The problems of the synephrine separation, such as band tailing and low retention, were solved successfully by using perfluorinated carboxylic acids. The effect of heptafluorobutyric acid (HFBA) was the best in the three investigated perfluorinated carboxylic acids. For the flavanone glycosides, the influence of the perfluorinated acids on retention time was rather weak. The two different kinds of the analytes were separated satisfactorily in one run using an isocratic eluent and the total analysis time takes less than 10 min. The abundance of pseudomolecular ions was recorded using selected ion monitoring (SIM) mode of m/ z 135.1, 273.1 and 303.1 for synephrine, naringin and hesperidin, respectively. The contents of hesperidin, naringin and synephrine in several Citrus herbs were simultaneously determined by the proposed method.

Fingerprint analysis of fruiting bodies of cultured Cordyceps militaris by high-performance liquid chromatography–photodiode array detection

We have developed and optimized a novel, efficient and accurate fingerprint method using high-performance liquid chromatography–photodiode array detection (HPLC–DAD) for the quality control of cultured Cordyceps militaris (L.) Link. The feasibility and advantages of the used chromatographic fingerprint were verified for the evaluation of cultured C. militaris by systematically comparing chromatograms with a professional analytical software recommended by State Food and Drug Administration (SFDA) of PR China. The results revealed that the chromatographic fingerprint combining similarity evaluation could efficiently identify and distinguish cultured C. militaris from different sources.

Regions in China identification and quality control of radix Codonopsis by chemical fingerprint: Evaluation of lobetyolin from different cultivated

By using high-performance liquid chromatography-photodiode array detection, a simple and accurate chromatographic fingerprint method was developed for the identification of Radix Codonopsis (roots of Codonopsis) from different sources. Eighteen herbs of Codonopsis at different habitats in China, including roots from Codonopsis pilosula, Codonopsis pilosula var. modesta and Codonopsis tangshen were analyzed by the fingerprint. The amount of lobetyolin was calibrated, which was found to be more consistent in roots of Codonopsis pilosula as compared to that of Codonopsis pilosula var. modesta and Codonopsis tangshen. Having the fingerprint results, hierarchical clustering analyses were performed to classify the eighteen herbs into three groups: Codonopsis pilosula, Codonopsis pilosula var. modesta and Codonopsis tangshen. This clustering analysis agrees very well with the pharmacognostic identification result, and which could be used as a tool in the quality control of Radix Codonopsis.

Fingerprinting quality control of Qianghuo by high-performance liquid chromatography-photodiode array detection

A novel, simple and accurate fingerprint method was developed using high-performance liquid chromatography-photodiode array detection (HPLC-DAD) for the quality control of Qianghuo, a Tibetan folk and Chinese herbal medicine used as a diaphoretic, an antifebrile and an anodyne. For the first time, the feasibility and advantages of employing chromatographic fingerprint were investigated and demonstrated for the evaluation of Qianghuo by systematically comparing chromatograms of aqueous extracts with the professional analytical software recommended by State Food and Drug Administration (SFDA). Our results revealed that the chromatographic fingerprint combing similarity evaluation could efficiently identify and distinguish raw herbs of Qianghuo from different sources and different species. The effects on Notopterygium forbesii Boiss (Apiaceae) chromatographic fingerprints resulted from collecting locations, harvesting time were also examined.

Development of high-performance liquid chromatographic fingerprint for the quality control of Rheum tanguticum Maxim. ex Balf.

A novel, accurate and valid fingerprint method was developed by using high-performance liquid chromatography–photodiode array detection (HPLC–DAD) to control the quality of Rheum tanguticum Maxim. ex Balf. By comprehensively comparing chromatograms of all samples, our result revealed that the chromatographic fingerprint we developed combining similarity evaluation could efficiently identify and distinguish raw materials of R. tanguticum from different sources. Besides, 21 characteristic peaks in chromatogram of R. tanguticum were unambiguously confirmed by comparison of reference compounds based on their retention time ( t R) and UV spectra.

Enrofloxacin assay validation and pharmacokinetics following a single oral dose in chickens

The pharmacokinetics of enrofloxacin (ENRO), a fluoroquinolone antimicrobial agent, was studied in male broiler chickens (Cobb) after single oral administration of 10 mg of ENRO/kg b.w. A high-performance liquid chromatography-photodiode array detector (DAD) (HPLC-DAD) method was developed and validated and used for quantitation of ENRO and its major metabolite ciprofloxacin in plasma. The HPLC analyses were carried out using a cationic-octadecyl mixed column and 0.05 mol/L phosphate buffer (pH 2.5)/acetonitrile as mobile phase. The sample preparation of plasma consisted of the precipitation of proteins followed by solid phase extraction on cationic-octadecyl mixed cartridges. The method was validated considering linear range, linearity, selectivity, sensitivity, limit of detection (LOD), limit of quantitation (LOQ), intra- and inter-day precisions and accuracy. The LOD and LOQ for both fluoroquinolones were 60 and 200 ng/mL for plasma. The plasma concentration vs. time graph was characteristic of a two-compartment open model. The maximal plasma concentration of 1.5 +/- 0.2 mg/mL was achieved at 9 +/- 2 h. The elimination half-life and the mean residence time of ENRO were 1.5 +/- 0.2 and 15.64 h, respectively. The area under the concentration-time curve was calculated as 35 +/- 4 mgxh/mL.

Combinative method using HPLC quantitative and qualitative analyses for quality consistency assessment of a herbal medicinal preparation

A selective and efficient quality consistency assessment system was developed for monitoring the manufacturing processes of a Chinese herbal preparation, Qingfu Guanjieshu (QFGJS) capsule, and for assessing its stability over time. This system is based on quantitative determination of four marker compounds, i.e., sinomenine, paeoniflorin, paeonol, and curcumin, and on qualitative fingerprinting analysis of QFGJS using high-performance liquid chromatography–photodiode array detection (HPLC–DAD) method. The separation was performed on a Phenomenex ODS column by gradient elution with acetonitrile and aqueous phase (containing 0.1% phosphoric acid, adjusted with triethylamine to pH 3.5 ± 0.2) at a flow-rate of 1.0 ml/min. In fingerprinting analysis, the chemical characteristics of four herbs present in QFGJS (excluding Radix Aconiti Lateralis Preparata) were present in the HPLC chromatographic file. In addition, quantitative determination of hypaconitine was carried out with our published HPLC method as a supplement for quality control of the Radix Aconiti Lateralis Preparata in QFGJS. The results showed that the contents of these five marker compounds and HPLC fingerprint profiles of three batches of QFGJS products collected at 3 months after production in the stability testing were relatively consistent. This well-developed method could be used for quality assessment of the complex preparations of herbal medicine.

Simultaneous analysis and identification of main bioactive constituents in extract of Zizyphus jujuba var. sapinosa (Zizyphi spinosi semen) by high-performance liquid chromatography–photodiode array detection–electrospray mass spectrometry

A high-performance liquid chromatography (HPLC) coupled with UV photodiode array detection and electrospray mass spectrometry (HPLC–DAD–ESI-MS) was developed for the simultaneous analysis and identification of main bioactive constituents in extract of zizyphi spinosi semen including three flavonoids, three alkaloids and four triterpene saponins. The separation was achieved on a C 18 column with a gradient solvent system of 0.08% aqueous formic acid solution-acetonitrile. The compounds were identified with MS-spectra and authentic standards. The contents of jujuboside A, jujuboside B, spinosin and 6‴-feruloyl spinosin were quantified by using DAD in different extract of zizyphi spinosi semen.

Flowers and Leaves of Tropaeolum majus L. as Rich Sources of Lutein

ABSTRACT: As increasing evidence supports the role of lutein and zeaxanthin in reducing the risk of cataract and macular degeneration, food sources of these carotenoids are being sought. In the present study, the lutein content of the edible flowers and leaves of Tropaeolum majus L. was determined by high-performance liquid chromatography-photodiode array detector (HPLC-PDAD), complemented by HPLC-mass spectrometry (MS) for identification. Chemical reactions were also used as identifying parameters. The yellow and brownish orange flowers had 450 ± 60 μg/g and 350 ± 50 μg/g lutein, respectively. Violaxanthin, antheraxanthin, zeaxanthin, zeinoxanthin, β-cryptoxanthin, α-carotene, and β-carotene were also detected at very low levels. The leaves had 136 ± 18 μg/g lutein, 69 ± 7 μg/g β-carotene, 74 ± 23 μg/g violaxanthin, and 48 ± 13 μg/g neoxanthin. Lutein was partly esterified in the flowers and unesterified in the leaves. The flowers of T. majus are therefore excellent food sources of lutein and the leaves good sources of both lutein and the provitamin A β-carotene.

Comparative Analyses of Bioactive Mammea Coumarins from Seven Parts of Mammea americana by HPLC-PDA with LC-MS

Mammea americana L. is tropical plant in the Clusiaceae family that bears edible fruit. Mammea coumarins are isoprenylated derivatives of the lactones of the 2-hydroxy-Z-cinnamic acids that are bioactive and have limited distribution in three Clusiaceae genera. Qualitative and quantitative analyses were performed to determine the distribution of mammea coumarins in the seed nucleus, seed coat, fruit flesh, fruit skin, leaf, stem, and root of M. americana using high-performance liquid chromatography-photodiode array detector (HPLC-PDA) and liquid chromatography-mass spectrometry (LC-MS). Ten major mammea coumarins, mammea E/BD (1), mammea E/BC (2), mammea E/BA (3), mammea E/BB (4), mammea B/BA hydroxycyclo F (5), mammea B/BD (6), mammea B/BC (7), mammea B/BA (8), mammea B/BB (9), and mammea B/BA cyclo F (10), were isolated and identified from the seed nucleus of M. americana and employed as standards. The HPLC-PDA method was validated with respect to sensitivity, linearity, recovery, accuracy, and precision. The total content (w/w %) of the 10 major mammea coumarins in M. americana was determined to be highest in the root (0.75%), followed by the leaf (0.64%), seed nucleus (0.48%), fruit skin (0.11%), stem (0.08%), seed coat (0.02%), and fruit flesh (<0.01%). The leaf and seed nucleus are rich and sustainable natural sources of mammea coumarins. Additionally, the described HPLC-PDA and LC-MS methods are sensitive and accurate and can be applied to the analysis of mammea coumarins in other samples.

Evaluation of the botanical authenticity and phytochemical profile of black cohosh products by high-performance liquid chromatography with selected ion monitoring liquid chromatography-mass spectrometry.

Black cohosh (Actaea racemosa L., syn. Cimicifuga racemosa L.) has become increasingly popular as a dietary supplement in the United States for the treatment of symptoms related to menopause, but the botanical authenticity of most products containing black cohosh has not been evaluated, nor is manufacturing highly regulated in the United States. In this study, 11 black cohosh products were analyzed for triterpene glycosides, phenolic constituents, and formononetin by high-performance liquid chromatography-photodiode array detection and a new selected ion monitoring liquid chromatography-mass spectrometry method. Three of the 11 products were found to contain the marker compound cimifugin and not cimiracemoside C, thereby indicating that these plants contain Asian Actaea instead of black cohosh. One product contained both black cohosh and an Asian Actaea species. For the products containing only black cohosh, there was significant product-to-product variability in the amounts of the selected triterpene glycosides and phenolic constituents, and as expected, no formononetin was detected.

Quantitative Determination of Triterpenes from Amphiptherygium adstringens by Liquid Chromatography and Thin-Layer Chromatography and Morphological Analysis of Cuachalalate Preparations

Amphiptherygium adstringens (Anacardiaceae/Julianaceae), local name "cuachalalate," is used in folk medicine for the treatment of cholelithiasis, fevers, fresh wounds, hypercholesterolemia, gastritis, gastric ulcers, and cancer of the gastrointestinal tract. The development of column high-performance liquid chromatography-photodiode array detector (LC-PDA) and high-performance thin-layer chromatography (HPTLC)-densitometry methods for the determination of masticadienonic acid and 3-hydroxymasticadienonic acid in cuachalalate preparations is described in this paper. Good separation of the compounds could be achieved by both methods. Either might be preparable depending on the requirements. The LC separation was performed on a Phenomenex Synergi MAX-RP 80A reversed-phase column operated at 40 degrees C with detection at 215 nm. The plant materials were extracted with methanol by sonication. The triterpenes present in the plant material and commercial extracts were separated with an acetonitrile-water reagent alcohol isocratic system. The limit of detection was 0.1-0.2 microg/mL. The relative standard deviation values for the determination of triterpenes in plant extracts were less than 1.00%. This is the first report of an analytical method developed for the quantitative analysis of triterpenes from Amphiptherygium adstringens by LC-PDA and HPTLC. The stem bark showed higher amounts of triterpenes, and low amounts in root and stem root. The microscopic description of the crude drug of cuachalalate was also provided.

Polyphenol profiles of apple juices

Focusing on 17 constituents, the polyphenol profiles of juices freshly made from various dessert (n = 4) and cider apple cultivars (n = 7) as well as commercially available apple juices (n = 24) were investigated using high-performance liquid chromatography-photodiode array detection (HPLC-DAD) and (HPLC)-electrospray ionization-tandem mass spectrometry (ESI(neg)-MS/MS) analyses. Significant differences in the total polyphenol content as well as the profiles of the apple cultivars under study were observed. For dessert apples the total polyphenol content ranged from 154 to 178 mg/L, whereas for 'old' German cider apple cultivars 261-970 mg/L were determined. Boskoop showed the highest (970 mg/L) and Granny Smith the lowest (154 mg/L) polyphenol content of the freshly prepared samples under study. Hydroxycinnamic acids, with chlorogenic acid as dominating constituent, ranged from 57 to 68 mg/L as well as from 134-593 mg/L in juices made from dessert apples and that from cider apples, respectively. Dessert apple juices showed lower contents of dihydrochalcones (10-35 mg/L) and flavan-3-ols (50-95 mg/L) compared to that of cider apples (34-171 mg/L and 70-393 mg/L, respectively). Quercetin and its derivatives were found from 0.4-4 mg/L and 0.4-27 mg/L in juices made from dessert apples and that of cider apples, respectively. Compared with freshly made juices, lower contents of polyphenols were determined in the commercial samples under study. Amounts ranging from 110-459 mg/L, dominated by chlorogenic acid with concentrations from 53-217 mg/L, were determined. Information about cultivar-typical apple polyphenol content and profile is important for bioactivity studies and, consequently, essential for the development of consumer-relevant products with particular nutritional functionalities.

Fingerprint quality control of Tianjihuang by high-performance liquid chromatography–photodiode array detection

A novel, simple and accurate fingerprint method was developed using high-performance liquid chromatography–photodiode array detection (HPLC–DAD) for the quality control of Hypericum japonicum thunb (Tianjihuang), a Chinese herbal medicine used for the treatment of several bacterial diseases, infectious hepatitis, gastrointestinal disorder, internal hemorrhage and tumors. For the first time, the feasibility and advantages of employing chromatographic fingerprint were investigated and demonstrated for the evaluation of Tianjihuang by systematically comparing chromatograms with a professional analytical software recommended by State Food and Drug Administration (SFDA). Our results revealed that the chromatographic fingerprint combining similarity evaluation could efficiently identify and distinguish raw herbs of Tianjihuang from different sources. The effects resulted from collecting locations, harvesting time and storage time on herbal chromatographic fingerprints were also examined.

Simultaneous analysis of N-nornuciferine, O-nornuciferine, nuciferine, and roemerine in leaves of Nelumbo nucifera Gaertn by high-performance liquid chromatography–photodiode array detection–electrospray mass spectrometry

A novel method has been developed for analysis of N-nornuciferine, O-nornuciferine, nuciferine, and roemerine in leaves of Nelumbo nucifera Gaertn by using high-performance liquid chromatography (HPLC)–photodiode array detection (DAD)–electrospray mass spectrometry (ESI-MS). The method was carried out by using a Shimadzu VP-ODS column with a gradient solvent system of 0.1% triethylamine aqueous solution–acetonitrile. N-Nornuciferine, O-nornuciferine, nuciferine, and roemerine were identified with authentic standard compounds and with MS-spectra. The contents of these compounds were measured by employing DAD. Linearity of around three orders in the magnitude of concentration was generally obtained and limits of detection for these compounds were in the range of 30–90 pg.

The value of plant collections in ethnopharmacology: a case study of an 85-year-old black cohosh ( Actaea racemosa L.) sample

Ethnopharmacological collections of plants used in traditional medical systems are a valuable but often underappreciated resource for scientific investigation. These collections contain many samples of plants currently employed in herbal and pharmaceutical medicine, and questions on stability and storage life can be examined using these historic collections as vouchers. A sample of black cohosh ( Actaea racemosa L.), collected in 1919 by the physician and plant explorer Henry Hurd Rusby, was recently identified in the collections of The New York Botanical Garden and analyzed for its triterpene glycosidic and phenolic constituents qualitatively and quantitatively by high-performance liquid chromatography–photodiode array detector (HPLC–PDA) and liquid chromatography–mass spectrophotometry (LC–MS). A comparison of the triterpene glycosidic and phenolic constituents of the 85-year-old plant sample with those of a modern collection of Actaea racemosa showed the similarity of the two samples, confirming the stability of the older sample, despite its curation over the years under a variety of conditions. Quantitative analyses indicated that both plant samples have similar amounts of the four major triterpene glycosides, but the total amount of the six major phenolic constituents measured in the 85-year-old plant material is lower than the amount measured in the modern plant material. Methanol extracts of the two plant materials were tested for their antioxidant activity, and both extracts showed similar antioxidant activity.

Development of a quantitative high-performance liquid chromatography–photodiode array detection measurement system for phenolic acids

A quantitative high-performance liquid chromatography–photodiode array detection method separating 16 phenolic acids was achieved. Six columns and several mobile phases were investigated. Resolution was achieved with a high-purity silica Phenomenex Luna C 18 column ( 150 mm×4.6 mm, 5 μm) and a binary gradient consisting of CH 3OH–water (with 0.1% formic acid) and flow rate set at 0.7 ml/min. Acids were detected and quantitation performed at wavelength representing the lowest energy λ max for individual acids. Extraction procedure from wine was optimized and yields ranged from 79 to 87% based on internal standard recovery. To confirm our quantitative results, identical samples were analyzed both in-house and by a collaborating laboratory. Correlation of two data sets generated linear regression equations that approached unity (0.93–0.98) and R 2 values ranging from 0.990 to 0.999.

Infiltration and adsorption of dissolved metolachlor, metolachlor oxanilic acid, and metolachlor ethanesulfonic acid by buffalograss (Buchloe dactyloides) filter strips

Vegetative filter strips (VFS) potentially reduce herbicide transport from agricultural fields by increasing herbicide mass infiltrated (Minf) and herbicide mass adsorbed (Mas) compared with bare field soil. However, there are conflicting reports in the literature concerning the contribution of Masto herbicide trapping efficiency (TE). Moreover, no study has evaluated TE among metolachlor and metolachlor metabolites in a VFS. This experiment was conducted to compare TE,Minf, andMasamong metolachlor, metolachlor oxanilic acid (OA), and metolachlor ethanesulfonic acid (ESA) in buffalograss filter strips. Runoff was applied as a point source upslope of a 1- × 3-m microwatershed at a rate of 750 L h−1. The point source was fortified with metolachlor, metolachlor OA, and metolachlor ESA, each at 0.12 μg ml−1. After moving through the plot, water samples were collected at 5-min intervals and stored at 5 C until analysis. Water samples were extracted using solid-phase extraction and analyzed by high-performance liquid chromatography–photodiode array detection. TE was significantly greater for metolachlor (25.3%) as compared with the OA (15.5%) and ESA metabolites (14.2%). The averageMinfwas 8.5% and was not significantly different among compounds. Significantly more metolachlor (17.3%) was retained as Mascompared with either metolachlor OA (7.0%) or metolachlor ESA (5.5%). Moreover,Masaccounted for 68 and 42% of the total TE for metolachlor and metolachlor metabolites, respectively. These results demonstrate that adsorption to the VFS grass, grass thatch, or soil surface (or all) is an important retention mechanism for metolachlor and metolachlor metabolites, especially under saturated conditions. Moreover, the Masdata indicate that metolachlor is preferentially retained by the VFS grass, grass thatch, or soil surface (or all) compared with the OA and ESA metabolites. Greater metolachlor retention in VFS compared with the OA and ESA metabolites may partially explain why metolachlor metabolites are frequently measured at higher concentrations than metolachlor is in surface water.

Simultaneous analysis of caffeic acid derivatives and alkamides in roots and extracts of Echinacea purpurea by high-performance liquid chromatography–photodiode array detection–electrospray mass spectrometry

High-performance liquid chromatography (HPLC) coupled with UV photodiode-array detection and electrospray ionization mass spectrometry was developed for the simultaneous analysis of caffeic acid derivatives and alkamides in the roots and extracts of Echinacea purpurea. Caffeic acid derivatives and alkamides produced very abundant peaks in the total ion current chromatogram during negative and positive cone voltage switching. Cichoric acid and the isomer pair, dodeca-2 E,4 E,8 Z,10 E/ Z-tetraenoic acid isobutylamide, were used as a standard for quantification of caffeic acid derivatives and alkamides in E. purpurea. This novel method surpasses previously published ones in product quality control and providing the HPLC chromatographic fingerprints of biological active components in E. purpurea.

Comparison of solid-phase extraction and solid-phase microextraction for carbofuran in water analyzed by high-performance liquid chromatography–photodiode-array detection

In this study a direct solid-phase microextraction (SPME) procedure has been developed for the determination of carbofuran in water. Experimental parameters such as selection of SPME coating, effect of temperature, effect of salt addition and solvent desorption were studied and optimized. Analytical parameters such as linearity, precision, detection and quantitation limits, and matrix effects for solid-phase extraction (SPE) and SPME methods were evaluated for comparison purposes with the aim of selecting the most appropriate depending on the detection capabilities required. SPE and SPME were followed by high-performance liquid chromatography with diode-array detection, using a 50×4.6 mm I.D. guard column and a 150×4.6 mm I.D. analytical column, both packed with C 18 silica. Both methods can be applied to real samples and give the same results, but SPE allows the detection of lower carbofuran concentrations (0.06 μg/L) as compared to SPME (8.9 μg/L).