Quality evaluation of Galli Gigerii Endothelium Corneum based on HPLC fingerprints and content determination of nucleosides / 中国中药杂志

Galli Gigerii Endothelium Corneum(GGEC), the dried gizzard membrane of Gallus gallus domesticus is a Chinese medicinal material commonly used for digestion. However, due to the particularity of texture and composition, its active ingre-dients have not been clarified so far, and there is also a lack of quality evaluation indicators. In this study, UPLC-Q-TOF-MS was used to analyze the chemical components from the water extract of GGEC, and ten nucleosides were identified for the first time. HPLC fingerprints of the water extracts of GGEC were established and the content of seven nucleosides was determined. The fingerprint similarities of 40 batches of GGEC samples ranged from 0.765 to 0.959, indicating that there were great differences among the GGEC products processed with different methods. In addition, SPSS 22.0 and SIMCA 14.1 were used for hierarchical cluster analysis(HCA) and principal component analysis(PCA) on the 19 common peaks of the HPLC fingerprints of GGEC, and the 40 batches of samples were divided into three categories: raw GGEC, fried GGEC and vinegar-processed GGEC. Eight differential components in GGEC were marked by orthogonal partial least squares discrimination analysis(OPLS-DA), two of which were adenine and thymine. The results of content determination showed that the total content of the seven nucleosides in raw GGEC, fried GGEC and vinegar-processed GGEC were 182.5-416.8, 205.3-368.7, and 194.2-283.0 μg·g~(-1), respectively. There were significant differences in the content of hypoxanthine, thymine and thymidine among the GGEC products processed with different methods(P<0.05), which were graded in the order of fried GGEC>vinegar-processed GGEC>raw GGEC. This suggested that the content of hypoxanthine, thymine and thymidine tended to increase during the frying process, and the variation range might be related to the degree of heat exposure. The established methods in this study were simple and reproducible, and could be used for qualitative and quantitative analysis of GGEC and its processed pro-ducts. This study also provided reference for the establishment of quality standards of GGEC with chemical components as control index.

Isolation, characterization, and validation of RP-HPLC method for the quantification of quercetin in Huberantha senjiana leaf extract

Huberantha senjiana (Annonaceae) – an endemic tree of Pakkamalai Reserve Forest Gingee Hills, Tamil Nadu, Indiawas subjected to preliminary phytochemical tests and high-performance liquid chromatography (HPLC) fingerprintchromatogram was established for use in future research. Subsequently, the HPLC-DAD-ESI (+) MS analysis toidentify the common biomarkers revealed the presence of Quercetin, a polyphenolic flavonol in the ethyl acetateextract of the leaves. An isocratic reversed-phase (RP)-HPLC method for the quantification of quercetin was developedwith phenomenex, RP C18 column using mobile phase [(H2O (0.1% formic acid, A); (MeOH: ACN, (40:15v/v), B),40:60 (v/v)] with a flow rate of 0.8 ml/min, detection max at 254 nm with a retention time of 7.58 minutes and wasvalidated according to the International Conference on Harmonization Q2B guidelines. Linearity was achieved withthe concentration range of 5.0–17.5 μg/ml with a R2 value of 0.996. Sensitivity was demonstrated with the limit ofdetection of 2.28 µg/ml and the limit of quantification of 6.92 µg/ml. The robustness was estimated from purposefulchanges in the composition of the mobile phase, wavelength, and flow rate and the limits were Not More Than (NMT)4%. The foliar concentration of quercetin was found to be 0.0055% w/w. The structural confirmation was done by theisolation and characterization by the divergent spectral analysis. This is the first published report on the identification,quantification, and isolation of quercetin in the leaves of H. senjiana species

Comparation of HPLC fingerprints of Lagotis integra and Lagotis brevituba between different origins / 中草药

Objective: To establish HPLC fingerprints of Lagotis integra and Lagotis brevituba which were contained in National Drug Standard and compared to Lagotis ramalana, Lagotis alutacea and Lagotis brachystachya by the established method. Methods: The similarity of HPLC fingerprints of L. integra and L. brevituba was low, so the HPLC fingerprint methods were established for both. The HPLC analysis was performed on Waters X-Bridge C18 (250 mm × 4.6 mm, 5 μm), using acetonitrile and 0.2% formic acid solution as the mobile phase at a flow rate of 1.0 mL/min, the detection wave-length was 328 nm and column temperature was 35 ℃ (L. integra) and 25 ℃ (L. brevituba). Results: There were 14 common peaks in the fingerprint of L. integra, and plantamajoside, hemiphroside B, 10-O-trans-p-methoxycinnamoyl-catalpol and 10-O-[(E)-3,4-dimethoxycinna moyl]-catalpol were standardized. There were 13 common peaks in the fingerprint of L. brevituba, and echinacoside, plantamajoside and acteoside were standardized. The similarity of HPLC fingerprint was more than 0.9 between L. integra and L. alutacea, but the others had low similarity with them. The similarity of HPLC fingerprint was more than 0.9 between L. brevituba from different batches and L. ramalana, while the others had low similarity with them. Conclusion: The established method could effectively identify L. brevituba, L. integra and L. alutacea were advised to be recorded in Medical Standards of the Ministry of Health. Lagotis ramalana could be used as a new base for "Honglian" (origin: Lagotis brevituba).

Vitellaria paradoxa nutshells from seven sub-Saharan countries as potential herbal medicines for treating diabetes based on chemical compositions, HPLC fingerprints and bioactivity evaluation / 中国天然药物

The aim of the study was to determine the feasibility of the Vitellaria paradoxa nutshell as a new medicinal resource for treating diabetes. A total of forty-one compounds were identified by HPLC-DAD-Q-TOF-MS and phytochemical methods in V. paradoxa nutshell methanol extract. Based on HPLC fingerprints, four characteristic constituents were quantified and the origin of twenty-eight V. paradoxa nutshells from seven sub-Saharan countries was compared, which were classified into three groups with chemometric method. Twenty-eight samples contained high total phenolic content, and exhibited moderate-higher antioxidant activity and strong α-glucosidase inhibitory activity. Furthermore, all fractions and isolated compounds were evaluated for their antioxidant and α-glucosidase inhibitory activities, and α-glucosidase inhibitory action mechanism of four characteristic constituents including protocatechuic acid, 3, 5, 7-trihydroxycoumarin, (2R, 3R)-(+)-taxifolin and quercetin was investigated via molecular docking method, which were all stabilized by hydrogen bonds with α-glucosidase. The study provided an effective approach to waste utilization of V. paradoxa nutshell, which would help to resolve waste environmental pollution and provide a basis for developing potential herbal resource for treating diabetes.

Analysis and comparison of HPLC fingerprints of wanai and Artemisiae argyi from thirty-one growing areas by multistatistical / 中成药

AIM To analyze and compare HPLC fingerprints of wanai and Artemisiae argyi Levi.et Vant from thirty-one growing areas by multistatistical.METHODS The analysis of 80％ methanol extract of A.argyi was developed on a 30 ℃ thermostatic Agilent TC-C18 column (4.6 mm × 250 mm,5 μm),with the mobile phase comprising of acetonitrile-water (containing 0.2％ methanoic acid) flowing at 1 mL/min in a gradient elution manner,and the detection wavelength was set at 330 nm.RESULTS There were eighteen,twenty-five common peaks in the fingerprints of thirty-one batches of A.argyi,fifteen batches of wanai,respectively,with the similarities all more than 0.900.The similarities of thirty-one batches of samples from different growing area were good and together as a category except samples from Dengzhou city,Luohe city and Anhui province.Fifteen batches of wanai samples got together with Qiai among them.The cumulative contribution rate of the four principal components from A.argyi was 86.049％.Twelve batches of wanai samples had higher scores than Qiai.CONCLUSION This stable and reliable method can be used for the quality control of A.argyi.

Establishment of HPLC fingerprints of Kangfuxin Liquid and determintion of six constituents / 中成药

AIM To establish the HPLC fingerprints of Kangfuxin Liquid (extract of Periplaneta americana L.) and to determine the contents of six constituents.METHODS The analysis of this drug was performed on a TOSOH TSK-GEL ODS column (250 mm × 4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-water (containing 0.07％ acetic acid) flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelength was set at 280 nm.RESULTS There were twenty-four common peaks in the fingerprints of ten batches of samples (Ⅰ-Ⅹ) with the similarities of 0.932-0.993 (except for sample Ⅰ).Uracil,hypoxanthine,xanthine,inosine,protocatechuic acid and Cyclo (Gly-Tyr) showed good linear relationships within the ranges of 3.460-173.0,3.960-198.0,3.596-179.8,1.338-66.9,3.672-183.6 and 3.552-177.6 μg/mL,whose average recoveries (RSDS) were99.8％ (2.65％),98.0％ (2.55％),99.7％ (1.59％),100.7％ (2.80％),102.0％ (2.09％) and 99.6％ (1.88％),respectively.CONCLUSION This accurate,stable and simple method can be used for the quality control of Kangfuxin Liquid.

Establishment of HPLC fingerprints of Geranium strictipes and pattern recognition / 中成药

AIM To establish HPLC fingerprints of Geranium strictipes R.Kunthand to perform pattern recognition.METHODS The analysis of 70％ methanol extract of G.strictipes was performed on a 25℃ thermostatic ZORBAX SB-C18 column(250 mm× 4.6 mm,5 μm),with the mobile phase comprising of acetonitrile-water in a gradient elution manner,and the detection wavelength was set at 254 nm.The results were analyzed by cluster analysis and principal component analysis.RESULTS There were thirteen common peaks in the HPLC fingerprints of twenty-one batches of samples with the similarities of more than 0.894.Twenty-one batches of samples were divided into two categories.The cumulative contribution rate of the first principal component was 31.067％.CONCLUSION This accurate,stable and reproducible method can be used for the quality control of G.strictipes.

Comparison of chemical quality characteristics between radial striations and non-radial striations in tuberous root of Rehmannia glutinosa / 中国中药杂志

An HPLC method was established to determine the contents of catalpol, acteoside, rehmaionoside A, rehmaionoside D, leonuride in three part of Rehmanni glutinosa in Beijing No.1 variety R. glutinosa during the growth period, This method, in combination with its HPLC fingerprint was used to evaluate its overall quality characteristics.The results showed that：① the content of main components of R. glutinosa varied in different growth stages ;② there was a great difference of the content of main components between theradial striations and the non-radial striations; ③ the two sections almost have the same content distribution of catalpol, acteoside and rehmaionoside D; ④the content of rehmaionoside A in non-radial striations was higher than that in radial striations,while the content of leonuride in radial striations was higher than that in non-radial striations.; ⑤the HPLC fingerprint of radial striations, non-radial striations and whole root tuber were basically identical, except for the big difference in the content of chemical components. The result of clustering displayed that the radial striations, non-radial striations, and whole root were divided into two groups. In conclusion, there was a significant difference in the quality characteristics of radial striations and non-radial striations of R. glutinosa. This research provides a reference for quality evaluation and geoherbalism of R. glutinosa.

Establishment of HPLC fingerprints of Zhihuang Tongfeng Decoction and determination of four constituents / 中成药

AIM To establish HPLC fingerprints of Zhihuang Tongfeng Decoction (Phellodendri chinensis Cortex,Anemarrhenae Rhizoma,Plantaginis Semen,etc.) and to determine the contents of four constituents.METHODS The analysis of aqueous extract of this drug was performed on a 30 ℃ thermostatic Wondasil C18 column (4.6 mm ×250 mm,5 μm),with the mobile phase comprising of acetonitrile-0.5％ acetic acid in a gradient elution manner,and the detection wavelength was set at 245 nm.RESULTS There were sixteen common peaks in the HPLC fingerprints of ten batches of samples with the similarities of more than 0.9.Berberine hydrochloride,mangiferin,geniposidic acid and salvianolic acid B showed good linear relationships within the ranges of 0.636 2-3.575 μg (R2 =0.9999),0.338-1.425 μg (R2 =0.9990),0.6452-2.7188 μg (R2 =1) and0.938 8-5.275 μg (R2 =0.999 3),whose average recoveries were 100.56％,98.35％,100.25％ and 102.11％ with the RSDs of 2.41％,1.17％,1.24％ and 2.02％,respectively.CONCLUSION This accurate,reliable and specific method can be used for the quality control of Zhihuang Tongfeng Decoction.

Establishment of HPLC fingerprints of Qingfeiyucuo Pills and determination of four constituents / 中成药

AIM To establish the HPLC fingerprints of Qingfeiyucuo Pills (an agent for the management of acne,containing Scutellariae Radix,Eriobotryae Folium,Paeoniae Radix Rubra,etc.) and to determine the contents of four constituents.METHODS The analysis of methanol extract of Qingfeiyucuo Pills was carried out on a 35 ℃ thermostatic Agilent HC-C1s column (4.6 mm ×250 mm,5 μm),with the mobile phase comprising of acetonitrile-0.4％ phosphoric acid flowing at 0.8 mL/min in a gradient elution manner,and the detection wavelength was set at 230 nm.RESULTS Eleven common peaks with the similarities of more than 0.9 were observed in the HPLC fingerprints of ten batches of samples.Paeoniflorin,hesperidin,baicalin and salvianolic acid B showed good linear relationships within the ranges of 0.08-0.25 μg,0.07-0.22 μg,0.2-0.60 μg and 0.06-0.18 μg,whose average recoveries were 98.7％,96.9％,97.4％ and 98.2％ with the RSDs of 0.98％,1.09％,0.82％ and 1.66％,respectively.CONCLUSION This sensitive and specific method can be used for the quality control of Qingfeiyucuo Pills.

Establishment of HPLC fingerprints of Cajanus cajan leaves and determination of two constituents / 中成药

AIM To establish the HPLC fingerprints of Cajanus cajan (L.) Millsp.leaves and to determine the contents of orientoside and luteolin.METHODS The analysis of 65％ methanol extract from C.cajan leaves was performed on a 25 ℃ thermostatic Agilent Zorbax SB-C18 column (4.6 mm ×250 mm,5 μm),with the mobile phase comprising of methanol-1％ acetic acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelength was set at 260 nm.RESULTS There were twenty-one common peaks in ten batches of samples (S1-S10),whose similarities were more than 0.950,except for that of S3 (0.516).Orientoside and luteolin showed good linear relationships within the ranges of 0.089 5-3.960 μg and 0.015 5-0.408 μg,whose average recoveries were 99.43％ (RSD =1.32％) and 98.50％ (RSD =0.82％),respectively.The contents of two constituents in the samples from three growing areas (Guangdong,Yunnan and Hainan) showed obvious differences.CONCLUSION This simple,accurate and reproducible method can be used for the quality control of C.cajan leaves.

Establishment of HPLC fingerprints of Zhihuang Tongfeng Decoction and determination of four constituents / 中成药

AIM To establish HPLC fingerprints of Zhihuang Tongfeng Decoction (Phellodendri chinensis Cortex,Anemarrhenae Rhizoma,Plantaginis Semen,etc.) and to determine the contents of four constituents.METHODS The analysis of aqueous extract of this drug was performed on a 30 ℃ thermostatic Wondasil C18 column (4.6 mm ×250 mm,5 μm),with the mobile phase comprising of acetonitrile-0.5％ acetic acid in a gradient elution manner,and the detection wavelength was set at 245 nm.RESULTS There were sixteen common peaks in the HPLC fingerprints of ten batches of samples with the similarities of more than 0.9.Berberine hydrochloride,mangiferin,geniposidic acid and salvianolic acid B showed good linear relationships within the ranges of 0.636 2-3.575 μg (R2 =0.9999),0.338-1.425 μg (R2 =0.9990),0.6452-2.7188 μg (R2 =1) and0.938 8-5.275 μg (R2 =0.999 3),whose average recoveries were 100.56％,98.35％,100.25％ and 102.11％ with the RSDs of 2.41％,1.17％,1.24％ and 2.02％,respectively.CONCLUSION This accurate,reliable and specific method can be used for the quality control of Zhihuang Tongfeng Decoction.

Establishement of HPLC fingerprints of flavonoids in Ziziphi spinosae Semen and determination of two constituents / 中成药

AIM To establish the HPLC fingerprints of flavonoids in Ziziphi spinosae Semen and to determine the contents of two constituents.METHODS With spinosin as a reference peak,the HPLC fingerprints of ten batches of samples were established.The analysis of methanol extract of flavonoids in Ziziphi spinosae Semen was performed on a 30 ℃ thermostatic Agilent TC-C18column (4.6 mm × 250 mm,5 μm),with the mobile phase comprising of acetonitrile-0.1％ acetic acid flowing at 1 mL/min in a gradient elution manner,and the detection wavelength was set at 280 nm.RESULTS There were ten common peaks in the HPLC fingerprints,eight of which (vicenin-Ⅱ,glucosylvitexin,isospinosin,spinosin,6'-pyridyloylspinosin,6'-p-hydroxybenzoylspinosin,6'-feruloylspinosin and 6'-p-coumaroylspinosin) were identified.Spinosin and 6'-feruloylspinosin showed good linear relationships within the ranges of 15.00-40.00 μg and 5.00-14.00 μg,whose average recoveries (RSDs)were 100.5％ (1.6％) and 100.4％ (1.6％),respectively.CONCLUSION This accurate,simple and reliable method can be used for the quality control of flavonoids in Ziziphi spinosae Semen.

Optimization of high pressure machine decocting process for Dachengqi Tang using HPLC fingerprints combined with the Box-Behnken experimental design / 药物分析学报

Using Dachengqi Tang (DCQT) as a model, high performance liquid chromatography (HPLC) fingerprints were applied to optimize machine extracting process with the Box–Behnken experimental design. HPLC fingerprints were carried out to investigate the chemical ingredients of DCQT; synthetic weighing method based on analytic hierarchy process (AHP) and criteria importance through intercriteria correlation (CRITIC) was performed to calculate synthetic scores of fingerprints; using the mark ingredients contents and synthetic scores as indicators, the Box–Behnken design was carried out to optimize the process parameters of machine decocting process under high pressure for DCQT. Results of optimal process showed that the herb materials were soaked for 45 min and extracted with 9 folds volume of water in the decocting machine under the temperature of 140 1C till the pressure arrived at 0.25 MPa;then hot decoction was excreted to soak Dahuang and Mangxiao for 5 min. Finally, obtained solutions were mixed, filtrated and packed. It concluded that HPLC fingerprints combined with the Box–Behnken experimental design could be used to optimize extracting process of traditional Chinese medicine (TCM).

Quality Control of Taraxacum Mongolicum Based on Comprehensive Score Method / 中国药师

Objective: To study the effects of different origins, collection and processing methods on the quality of Taraxacum mongolicum. Methods:The HPLC fingerprints of Taraxacum mongolicum were established. Totally 11 batches of Taraxacum mongoli-cum were analyzed by similarity evaluation and cluster analysis. Results:According to the results of HPLC, 11 batches of Taraxacum mongolicum had good baseline separation and showed 5 common peaks. Based on the established HPLC method, the quality of different batches of Taraxacum mongolicum showed difference according to the results of similarity evaluation and cluster analysis. The quality of batch 121231-1 was the best. Conclusion:The origin, collection and processing method show notable influence on the quality of Ta-raxacum mongolicum, and the comprehensive score method can be applied in the quality evaluation of Chinese herbs.

Selection of Guanxin Suhe Pill reference preparation by internal standard quantified fingerprints method / 中草药

Objective: To establish the Guanxin Suhe Pill (GXSHP)-HPLC internal standard quantified fingerprints method (ISQFM) for selecting the reference preparation of GXSHP by ISQFM and providing the quality control method of reference preparation. Methods: Agilent Poroshell 120 SB C18 column (150 mm × 4.6 mm, 2.7 μm), 0.2% ethylic acid-water, and 0.2% ethylic acid-methanol were used as the mobile phases that were eluted with liner program. The flow rate was 0.5 mL/min, the UV absorbance was monitored at 275 nm by injecting 5 μL of the sample solution each time, and the column temperature was maintained at (25.00 ± 0.15)°C. The ISQFM was used to identify the quality and select the reference preparation. Results: The 36 common peaks were marked by choosing IS as the referential peaks and the quality of 12 batches of GXSHPs was identified by the ISQFM. The identification results were that the quality of five batches (S3, S5-S7, and S9) were best, four batches (S4, S8, S10, and S11) were better, two batches (S1 and S2) were good, and one batch (S12) was lower than good, according to the results of this method, S3, S5-S7, and S9 could be used as the reference preparation of GXSHPs. Conclusion: This method could reduce the use of reference substances and improve the accuracy of the measured results. The reference preparation of GXSHPs, which is selected by this method, could be used for the quality control of large quantities of GXSHPs with the great significance in practical production.

Identification of Scutellaria amoena CH Wright and Scutellaria baicalensis Georgi with HPLC Finger-prints / 中国药房

OBJECTIVE:To identify different origin of Scutellaria amoena CH Wright and Scutellaria baicalensis Georgi indexed in China Pharmacopeia by HPLC fingerprints.METHODS:RP-HPLC method was adopted in which the samples were separated on Zorbax SB-C18(250mm?4.6mm,5?m)column at a column temperature of 30℃.The gradient elution was performed on the mobile phase which consisted of acetonitrile(contairing 1.0% methanoic acid)-water(containing 1.0% methanoic acid)at a flow rate of 1.0mL?min-1.The detection wavelength was set at 274nm.RESULTS:The fingerprints of Scutellaria amoena CH Wright that from different places were of great resemblance,so were the fingerprints between Scutellaria baicalensis Georgi that from different places,but the fingerprints between Scutellaria amoena CH Wright and Scutellaria baicalensis Georgi were of significant difference,with resemblance of only 0.859.CONCLUSION:The RP-HPLC fingerprint serves as basis for the further study of the quality criteria of Scutellaria amoena CH Wright and Scutellaria baicalensis Georgi.

Evaluation on in vitro release of gastric floating sustained-release preparation of Tripterygium wilfordii by HPLC fingerprints / 中草药

Objective To investigate the drug in vitro release behavior of gastric floating sustained-release preparation of Tripterygium wilfordii and establish their quality evaluating methods.Methods HPLC was Employed to gain the fingerprints of releasing medium of preparations.Triptolide was selected as marker to calculate the linear equation concerning peak area then the relative drug release and f2 simila-rity factor value were acquired.Results Various components in gastric floating sustained-release tablets and pellets of T.wilfordii could not release synchronously while sustained-releasing but all of them in the gastric floating sustained-release capsules of T.wilfordii prepared by using multiparticulate site-controlled release technology could release synchronously while sustained-releasing.Conclusion The quality and in vitro release behavior of gastric floating sustained-release preparation of T.wilfordii could be evaluated more scientifically and comprehensively by using HPLC fingerprints method.

Quality evaluation of Semen Cassiae by both indicated component determination and HPLC fingerprint / 中草药

Objective To evaluate the quality of Semen Cassiae from different habitats objectively. Methods To determine the content of chryso-phanic according to ChP and establish HPLC fingerprints with the gradient elution solvent composed of acetonitrile and 1% HAC. A C18 column (250 mm?4.6 mm, 5 ?m) was used, flow rate: 1 mL/min, detecting wavelength: 254 nm, and the column temperature: room temperature. The clustering analysis was carried out by SAS software according to the content of chrysophanic and similarity of HPLC fingerprints obtained by the software of similarity analysis. Results The established HPLC fingerprint has desirable precision, reproducibility, and stability. The content of chrysophanic and HPLC fingerprints of Semen Cassiae from various habitats are different, which differs from the habitats. The content range of chrysophanic in Semen Cassiae is 0.037%-0.170% and the similarity is 0.864-0.962. Conclusion The method indicates the difference of the chemical component in Semen Cassiae from various habitats and can be used as a quality control method for Semen Cassiae.

Qualitative and quantitative assessment on the HPLC fingerprint of Ginkgo biloba L.extract by the involution similarity / 中成药

AIM:To establish the assessing method of traditional Chinese medicine chromatogrphic fingerprints by the involution similarity and quantitative involution similarity and its application. METHODS: The fingerprint of Ginkgo biloba L.extract(GBE) was evaluated by the involution similarity and quantitative involution similarity,and the contrast studies were implemented by other appraisal targets,eg.the included angle cosine S_F,the correlation coefficient r and the indexes with quantitative characteristic,such as the apparently quantitative similarity R%、the average mass percent M%,etc.. RESULTS: The involution similarity could reflect the similarity change in each sample from different bathes and be of quantitative function.The quantitative involution similarity and quantitative weighted similarity could perfectly reflect the changes in contents of the consituents of GBE. CONCLUSION: The involution similarity and quantitative involution similarity can qualitatively and quantitatively assess the similarity between sample and the referential fingerprint.They are certainly the novel method of evaluating the traditional Chinese medicine chromatographic fingerprints.