Simultaneous determination of seven flavonoids in Epimedium by liquid chromatography-tandem mass spectrometry method

In this paper, a sensitive and specific liquid chromatography-electrospray ionisation-mass spectrometry （LC-ESI-MS） method has been developed and validated for the identification and determination of seven flavonoids, namely epimedin A, epimedin B, epimedin C, icariin, sagittatoside B, 2＂-O-rhamnosyl icariside II, and baohuoside I in Epimedium from different sources.

Quantitative determination of polysaccharide in Curcuma wenyujin, a traditional Chinese medicine, by a phenol-sulfuric acid method

Curcuma wenyujin has been widely used as a traditional medicine in China. In this paper a strategy for the quantitative determination of the polysaccharide by a phenol-sulfuric acid method was described. Involved in three factors, 5% phenol volume, H2SO4 volume, and temperature of water bath, we adopted the L9（3）3 orthogonal array design to gain the optimal colorimetric method. 3.0 ml of polysaccharide solution, 1.0 ml, 5% phenol and 7.0 ml H2SO4 were mixed with constant stirring in a glass vessel, and then kept in a water bath at 40 ℃. After cooling to room temperature for 20 min, the absorbance values were recorded by the UV-2501 PC spectrometer at the wavelength range of 485 nm. The polysaccharide content in Curcuma wenyujin were 3.21%, 3.23%, 3.20%, 3.18~/0, 3.22% and 2.38%, respectively. All results showed that this method was adequate, valid and applicable, may be applied to the determination of other bacterial polysaccharide as well.

Simultaneous determination of ten components in Dianxiankang Capsules by HPLC QAMS

Objective:To establish an high performance liquid chromatography-quantitative analysis of multi components by single marker(HPLC-QAMS)method for simultaneous determination of gastrodin,parishin E,parishin B,parishin,tenuifolin,onjisaponin B,methylophiopogonanone A,methylophiopogonanone B,β-asarone andαasarone in Dianxiankang Capsules.Methods:Waters Symmetry C_(18)column was used with acetonitrile-0.05%phosphoric acid solution as mobile phase for gradient elution.Multiwavelength switching detection.The contents of gastrodin,parishin E,parishin B,parishin,onjisaponin B,methylophiopogonanone A,methylophiopogonanone B,β-asarone andα-asarone were calculated by relative correction factor.At the same time,the contents of 10 components in 12 batches of Dianxiankang Capsules were determined by external standard method(ESM).Results:An HPLC-QAMS method was established tenuifolin as the internal reference substance was established.The relative correction factors of gastrodin,parishin E,parishin B,parishin,onjisaponin B,methylophiopogonanone A,methylophiopogonanone B,β-asarone andα-asarone were 0.8238,0.7239,1.0229,1.1881,0.7272,1.3108,0.9314,0.6549 and 1.0572,respectively.The relative correction factors had good repeatability and no significant difference with ESM(P>0.05).Conclusion:HPLC-QAMS can be used for simultaneous determination of multi-index components in Dianxiankang Capsules.

Quantitative Determination of Tetrachlorantraniliprole by(1)^H NMR Spectroscopy with Internal Standard Method

[Objective]This study was to establish a rapid,specific and simple method for quantitative determination of tetrachlorantraniliprole by 1H NMR.[Method]1H NMR spectroscopy was acquired with deuterium DMSO as the solvent and maleic acid as internal standard under the conditions of temperature 25℃,pulses width 8.0μs,delay time 5 s,and scanning times 8.[Result]The hydrogen proton peaks of tetrachlorantraniliprole(δ=10.55)and maleic acid(δ=6.27)were taken as quantitative peaks.The peak area ratio y(As/Ar)and mass ratio x(ms/mr)were linearly regressed,and the correlation coefficient was 0.9999.The RSD value of repeatability test was 0.38%,and the RSD value of stability test was 0.77%.The content of tetrachlorantraniliprole was determined as 99.6%.[Conclusion]1H NMR spectroscopy can be used for quantitative determination of tetrachlorantraniliprole without standard reference,which is rapid,accurate and simple.

A Mathematical Calculation Model Using Biomarkers to Quantitatively Determine the Relative Source Proportion of Mixed Oils

It is difficult to identify the source（s） of mixed oils from multiple source rocks, and in particular the relative contribution of each source rock. Artificial mixing experiments using typical crude oils and ratios of different biomarkers show that the relative contribution changes are non-linear when two oils with different concentrations of biomarkers mix with each other. This may result in an incorrect conclusion if ratios of biomarkers and a simple binary linear equation are used to calculate the contribution proportion of each end-member to the mixed oil. The changes of biomarker ratios with the mixing proportion of end-member oils in the trinal mixing model are more complex than in the binary mixing model. When four or more oils mix, the contribution proportion of each end-member oil to the mixed oil cannot be calculated using biomarker ratios and a simple formula. Artificial mixing experiments on typical oils reveal that the absolute concentrations of biomarkers in the mixed oil cause a linear change with mixing proportion of each end-member. Mathematical inferences verify such linear changes. Some of the mathematical calculation methods using the absolute concentrations or ratios of biomarkers to quantitatively determine the proportion of each end-member in the mixed oils are deduced from the results of artificial experiments and by theoretical inference. Ratio of two biomarker compounds changes as a hyperbola with the mixing proportion in the binary mixing model, as a hyperboloid in the trinal mixing model, and as a hypersurface when mixing more than three end- members. The mixing proportion of each end-member can be quantitatively determined with these mathematical models, using the absolute concentrations and the ratios of biomarkers. The mathematical calculation model is more economical, convenient, accurate and reliable than conventional artificial mixing methods.

HPLC Fingerprint with Multi-components Analysis for Quality Consistency Evaluation of Traditional Chinese Medicine Si-Mo-Tang Oral Liquid Preparation

Si-Mo-Tang（SMT） oral liquid preparation, a traditional Chinese medicine, was prepared from four crude herbal drugs, Fructus Aurantii Submaturus, Radix Aucklandiae, Semen Arecae and Radix Linderae Aggregatae. A combinative method using HPLC fingerprint and quantitative analysis was developed and validated for quality consistency evaluation of SMT. Individual HPLC chromatograms were evaluated against the mean chromatogram generated via a similarity evaluation computer program. Data from chromatographic fingerprints were also processed with principal component analysis（PCA） and hierarchical cluster analysis（HCA）. Additionally, six components （naringin, isonaringin, hesperidin, neohesperidin, norisoboldine and potassium sorbate） in SMT were simultaneously determined to interpret the quality consistency. For fingerprint analysis, 20 peaks were selected as the characteristic peaks to evaluate the similarities of 26 SMT collected from different manufacturers. Among the 20 characteristic peaks, 10 peaks were assigned to be naringin, hesperidin, neohesperidin, isonaringin, neoeriocitrin, tangeretin, nobiletin, norisoboldine, 5-（ethoxymethyl）furan-2-carbaldehyde and potassium sorbate, respectively. The results of similarity analysis, PCA and HCA, indicate that the samples from different manufacturers were consistent with each other in composition. The results from the quantitative data show that the contents of six compounds were significantly different in SMT oral liquid preparations from different manufacturers. The combinative method of chromatographic fingerprint with quantitative analysis developed here offered an efficient way for the quality consistency evaluation of the traditional Chinese medicine SMT.

Simultaneous determination of 10 nucleosides and nucleobases from different cultivation years of Fritillaria unibracteata var. wabuensis by HPLC-DAD

Determination of nucleosides and nucleobases is important for the quality control of Fritillaria unibracteata Hsiao et K.C. Hsia var. wabuensis （FUW） due to their physiological and pharmacological actions. In the present study, we developed a sensitive and reliable HPLC-diode-array detection method to simultaneously determine ten nucleosides and nucleobases, including cytosine, uracil, cytidine, uridine, thymine, adenine, inosine, guanosine, thymidine and adenosine. Complete separation of all the analytes was achieved on a Zorbax 300 A 300 Extend C18 column with a gradient of methanol-ultrapure water at a flow rate of 1 mL/min in less than 30 min. The diode-array detector wavelength was set at 260 nm for the UV detection of nucleosides and nucleobases. The optimized method provided good linearity （R2〉0.9993 for all the analytes）, satisfactory precision （RSD〈3.715%）, good repeatability （RSD_〈3.748%） and good recovery （RSD from 97.688% to 102.923%）. In addition, the developed method was successfully applied to simultaneous determination of ten nucleosides and nucleobases from FUW, and their content changes of various cultivation time （1-7 years） were further analyzed for the first time. Our findings were useful for ensuring the cultivation time choice of artificial cultivation, quality control, pharmaceutical studies and clinical efficacy of FUW.

Simultaneous determination of five major components of Glycyrrhizae Radix et Rhizoma in Xiaoer Zhike Tangjiang with standardized reference extract

In this study, we developed a novel and simple HPLC-DAD method for simultaneous qualitative and quantitative determination of five major components of Glycyrrhizae Radix et Rhizoma （GRR） in Xiaoer Zhike Tangjiang （XEZKTJ） with standardized reference extract （SRE）. The five analytes （liquiritin apioside, liquiritin, isoliquiritin apioside, liquirigenin and glycyrrhizic acid） were well separated with good linearity, precision, stability and repeatability. The recovery rates ranged from 95.69% to 100.80%. The content of the five compounds in 34 batches of commercial XEZKTJ products was determined using standardized GRR extract （SRE method） and individual chemical reference standards （CRS method）. Highly similar results were obtained from the two methods, demonstrating the feasibility of the proposed SRE method. Taken together, we proposed an efficient and low-cost way to perform multi-component quality control of XEZKTJ in this study.

Simultaneous determination of five bioactive phenolic acids in Salvia yunnanensis and Salvia miltiorrhiza by HPLC

A reverse-phase HPLC method was developed for the simultaneous separation and determination of five bioactive phenolic acids,yunnaneic acid E,rosmarinic acid,lithospermic acid,salvianolic acid B and salvianolic acid A in eight different samples of Salvia yunnanensis collected in Yunnan Province.For comparison,the sample of Salvia miltiorrhiza was included. All the samples were extracted for 60 min with 50%methanol in an ultrasonic bath.The optimal separation was achieved on a YMC-Pack Pro C18 column,with a gradient of 0.1%（v/v） phosphoric acid and acetonitrile,at a flow rate of 1.0 mL/min and at a detection wavelength of 280 nm.The separation was obtained within 65 min for five bioactive phenolic acids.All calibration curves showed good linearity（r^2〉0.999） within test ranges.The relative standard deviation of the method was less than 5%for intra- and inter-day assays.The mean recovery of the method was in the range from 97%to 104%,with RSD less than 5%.This assay was successfully applied to the quantitative determination of five bioactive phenolic acids in nine resource samples. The results showed that the developed HPLC assay was suitable for the quality control of S.yunnanensis and it can be used to differentiate S.yunnanensis from S.miltiorrhiza.

Application of a quantitative ^1H NMR method for rapid extraction and determination of the content of paeonol in Cynanchum paniculatum

Cynanchum paniculatum(Bunge) Kitagawa is usually used as an herbal medicine for treating many diseases. Paeonol is the main active component, and its content is the key indicator for quality control of C. paniculatum. In the present study, we developed a rapid, accurate and precise method for quantitation of paeonol in C. paniculatum using 1 H NMR spectra. The deuterated solvent of methanol-d4 enabled satisfactory separation of the signals to be integrated in 1 H NMR spectrum. H-6(δ 7.78) of 1 H NMR spectrum of C. paniculatum was selected as the feature signal for quantitation, and trimesic acid(TMA) was selected as an internal standard. Validation of the quantitative method was performed in terms of linearity, specificity, repeatability and stability. This is the first time to report quantitative 1 H NMR(qHNMR) applied to determine the content of paeonol in C. paniculatum and showed a wider linearity range than the reported quantitation of paeonol in others. The simple extraction of paeonol from C. paniculatum was rapid and will prompt the application of the developed method. This work implied that qHNMR represented a feasible alternative to HPLC-based methods for quantitation of paeonol in C. paniculatum, and it was suitable for the quality control of C. paniculatum.

Determination of inulin-type fructo-oligosaccharides in inulin by HPLC-ELSD

To develop an HPLC-ELSD method for the determination of nine inulin-type fructo-oligosaccharides in inulin,the HPLC-ELSD system consisted of Waters XBridge■ Amide column(4.6 mm×250 mm,5μm)with a gradient elution mobile phase consisting of acetonitrile and water at a flow rate of 1.2 mL/min at 30°C.The detector was an Agilent Technologies 380-ELSD.The drift tube temperature for the ELSD was set at 55°C with a nitrogen flow rate of 1.8 L/min.The injection volume was 15μL.The results showed that the detection range for the nine inulin-type fructo-oligosaccharides was 3.81–30.60μg R^(2)=0.99969 for kestose,3.73–29.97μg R^(2)=0.99981 for nystose,3.82–30.69μg R^(2)=0.99993 for fructosylnystose,3.80–30.48μg R^(2)=0.99995 for GF5,3.73–29.96μg R^(2)=0.99993 for GF6,3.78–30.30μg R^(2)=0.99983 for GF7,3.82–30μg R^(2)=0.99989 for GF8,3.71–29.80μg R^(2)=0.99974 for GF9,3.61–29.00μg R^(2)=0.99970 for GF10,respectively.The recovery of the nine oligosaccharides ranged between 96.48%–100.84%(n=6).The method was simple,accurate,and reproducible that it could be used as an analytical method for evaluating the quality of inulin effectively.

Simultaneous Determination of Two Major Triterpenoid Saponins:Celosins Ⅰ and Ⅱ in Celosiae Semen by HPLC-ELSD

Objective To establish an analytical method for the simultaneous determination of celosin Ⅰ and celosin Ⅱ, two major triterpenoids in Celosia Semen and compare the contents of celosin Ⅰ and celosin Ⅱ from different habitats to screen the resources of elite germplasm for further applications. Methods A sensitive and simple high- performance liquid chromatography coupled with evaporative light-scattering detector（HPLC-ELSD） method was developed for the first time for the simultaneous determination of celosin Ⅰ and celosin Ⅱ. Using this method, 21 batches of Celosiae Semen were determined from different habitats in China. Results There was an obvious difference in the contents of celosin Ⅰ and celosin Ⅱ among Celosiae Semen species from various habitats across China. The crude drug from Yongzhou, Hunan province, Zhuhai, Guangdong province, and Nanning, Guangxi province showed the highest contents of all habitats, while Anguo, Hebei province, Haidian, Beijing, and Zhengzhou, Henan province showed the lowest content. The results also showed that geographical location had a great influence on the contents of the two compounds. The batches from lower latitudes were higher in contents of celosin Ⅰ and celosin Ⅱ. Conclusion The sun light may be a key factor influencing the contents of the two saponins, indicating that the environment has a great impact on the quality of Celosiae Semen.

Simultaneous quantitative determination of eight index constituents and compatibility changes in Longchai Decoction by UPLC–Q-TOF-MS

The goal of this research was to develop a simple,rapid and sensitive method for simultaneous quantitative determination of salidroside,gardenoside,liquiritin,baicalin,wogonoside,wogonin,saikosaponin A and saikosaponin D in Longchai Decoction by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry(UPLC–Q-TOF-MS),in order to control the quality of Longchai Decoction and to analyze the changes of chemical components before and after the compatibility of the component herb drugs.The chromatographic separation was performed on the Waters ACQUITY BEH C18 column(2.1 mm
100 mm,1.8μm)using the mixture of acetonitrile and 0.1%(v/v)methanoic acid as mobile phase with a gradient elution program at the flow rate of 0.3 mL/min and the column temperature of 301C.The eight components of the standards achieved baseline separation.Regression analysis revealed a linear relationship(r 240.9998)between the contents and the peak areas of the mixed standard substances.The average recovery rates were between 99.72%and 102.13%with RSD values were less than 2.82%(n¼5).The obtained results indicated that the content of index components were higher in co-decoction compared to mixed decoction.This method with a good resolution and high precision can be used for the quality control of Longchai Decoction.

Simultaneous determination of eight bioactive bakkenolides of Petasites tatewakianus Kitam by HPLC–UV

A high-performance liquid chromatography coupled with variable wavelength detection(HPLC–UV)was developed to evaluate the quality of Petasites tatewakianus Kitam through a simultaneous determination of eight bakkenolides:bakkenolide-L,bakkenolide-D,bakkenolide-B,bakkenolide-Ⅰa,bakkenolide-Ⅶa,bakkenolide-Ⅳa,bakkenolide-Ⅲa and homofukinolide.With a C18 analytical column,the eight analytes were efficiently separated using tetrahydrofuran–acetonitrile–water as the mobile phase in a constant program.The limits of detection and limits of quantitation of the method ranged 0.42–2.56μg/mL and 1.22–8.40μg/mL,respectively.The intra-and inter-day precisions of the method were all less than 1.83%.All the recoveries for the spiked analytes ranged 97.8%–102.4%.There were statistically significant differences among the contents of the eight bakkenolides in different parts and origins of P.tatewakianus(Po0.01).The content of total bakkenolides in rhizome was higher than that in other parts of the plant.The content of total bakkenolides in rhizome from Baishan was higher than those in other localities.The result suggested that rhizome may be the most valuable part of P.tatewakianus,and P.tatewakianus from Baishan may be the best plant resource.Our results might serve as a sound foundation for further study and application of this plant.

Origin of potential errors in the quantitative determination of terahertz optical properties in time-domain terahertz spectroscopy

We demonstrate theoretically and experimentally how changes of a terahertz （THz） beam induced by the sample affect the accuracy of the determination of THz dielectric properties in THz time-domain transmission spectros- copy （TDTS）. We apply a Gaussian beam and the ABCD matrix formalism to describe the propagation of the THz beam in a focused beam setup. The insertion of the sample induces a focus displacement which is absent in the reference t without a sample. We show how the focus displacement can be corrected. The THz optical properties after focus displacement correction reported in this Letter are in quantitative agreement with those obtained using collimated beam THz-TDTSinpreviouswork.

Determination of gas adsorption capacity in organic-rich marine shale: a case study of Wufeng-Lower Longmaxi Shale in the southeast Sichuan Basin

Determination of gas adsorption capacity under geological conditions is essential in evaluating shale gas resource potential.A quantitative determination of gas adsorption capacity was proposed through 1)investigating controlling geological factors(including both internal ones and external ones)of gas adsorption capacity in organic-rich marine shale with geochemical analysis,XRD diffraction,field-emission scanning electron microscopy,and methane sorption isotherms;2)defining the relationship between gas adsorption capacity and single controlling factor;3)establishing a comprehensive determination model with the consideration of all these controlling factors.The primary controlling factors of the sorption capacity for the studied O3wLower S1l shale are TOC,illite and quartz,temperature,pressure,Ro,and moisture(water saturation).Specifically,TOC,thermal maturity,illite,and pressure are positively correlated with sorption capacity,whereas,quartz and temperature contribute negatively to the sorption capacity.We present the quantitative model along with application examples from the Wufeng-Lower Longmaxi Shale in the southeast Sichuan Basin,west China,to demonstrate the approach in shale gas evaluation.The result shows that the comprehensive determination model provides a good and unbiased estimate of gas adsorption capacities with a high correlation coefficient(0.96)and bell-shaped residues centered at zero.

Simultaneous quantification of ten phenolic acids in Lonicerae Japonicae Flos by HPLC-DAD

A reverse-phase HPLC-DAD method was developed for simultaneous quantification of ten phenolic acids （caffeic acid, chlorogenic acid, neochlorogenic acid, 4-O-caffeoylquinic acid, 3,4-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid, 4,5-O-dicaffeoylquinic acid, 3,5-O-dicaffeoylquinic acid methyl ester, 3,4-O-dicaffeoylquinic acid methyl ester, and 4,5-0- dicaffeoylquinic acid methyl ester） in the dried flower buds of Lonicera japonica Thunb. （Lonicerae Japonicae Flos; LJF）. An optimal sample preparation method was established as 30-min ultrasonication with 100 times 50% （v/w） ethanol aqueous solution based on the orthogonal test results. The chromatographic separation of the ten phenolic acids was achieved with an AQ-C18 column （4.6 mm x 250 mm, 5 p.m） and a gradient elution of acetonitrile, methanol and 0.1% formic acid aqueous solution within 55 rain. All calibration curves showed good linearity （r2〉0.999） within test ranges. The average recoveries were in the range of 98.57%-103.22% with RSD less than 3%. The method developed was accurate, sensitive and reproducible for determination of ten phenolic acids in LJF.

Establishment of one-step approach to detoxification of hypertoxic aconite based on the evaluation of alkaloids contents and quality

Aconite is a valuable drug and also a toxic material, which can be used only after detoxification processing. Although traditional processing methods can achieve detoxification effect as desired, there are some obvious drawbacks, including a significant loss of alkaloids and poor quality consistency. It is thus necessary to develop a new detoxification approach. In the present study, we designed a novel one-step detoxification approach by quickly drying fresh-cut aconite particles. In order to evaluate the technical advantages, the contents of mesaconitine, aconitine, hypaconitine, benzoylmesaconine, benzoylaconine, benzoylhypaconine, neoline, fuziline, songorine, and talatisamine were determined using HPLC and UHPLC/Q-TOF-MS. Multivariate analysis methods, such as Clustering analysis and Principle component analysis, were applied to determine the quality differences between samples. Our results showed that traditional processes could reduce toxicity as desired, but also led to more than 85.2% alkaloids loss. However, our novel one-step method was capable of achieving virtually the same detoxification effect, with only an approximately 30% alkaloids loss. Cluster analysis and Principal component analysis analyses suggested that Shengfupian and the novel products were significantly different from various traditional products. Acute toxicity testing showed that the novel products achieved a good detoxification effect, with its maximum tolerated dose being equivalent to 20 times of adult dosage. And cardiac effect testing also showed that the activity of the novel products was stronger than that of traditional products. Moreover, particles specification greatly improved the quality consistency of the novel products, which was immensely superior to the traditional products. These results would help guide the rational optimization of aconite processing technologies, providing better drugs for clinical treatment.

Chemical fingerprint analysis of Gentianae Radix et Rhizoma by high-performance liquid chromatography

Gentianae Radix et Rhizoma(also called "Longdan" in Chinese)is commonly used for eliminating damp-heat and quenching the fire of liver and gall bladder in traditional Chinese medicine.In this study,a novel and reliable method using high-performance liquid chromatography(HPLC)was developed both for quantitative analysis of four bioactive compounds(loganic acid,swertiamarin,gentiopicroside and sweroside)and chemical fingerprint analysis of "Longdan".In quantitative analysis,four compounds showed good regressions(R^(2)>40.9987)within the test ranges and the recovery of the method was in the range 97.61-102.49%.In fingerprint analysis,ten characteristic peaks were selected to evaluate the similarities of the crude drugs,and the HPLC chromatograms of twenty samples from different regions of China showed similar patterns.The results demonstrated that the combination of the quantitative and chromatographic fingerprint analyses offered an efficient way to evaluate the quality consistency of Gentianae Radix et Rhizoma.

Insights into pH-dependent transformation of gibberellic acid in aqueous solution:Transformation pathway,mechanism and toxicity estimation

Gibberellic acid(GA_(3))is widely used in agriculture and maybe transfer with groundwater flow,which is an endocrine disruptor,but few studies have focused on the transformation pathway and toxicity assessment of GA_(3)and its products.Here,GA_(3)and its transformation products in aqueous solution were identified and quantified by liquid chromatography mass spectrometry hybrid ion trap time-of-flight(LCMS-IT-TOF)and high-performance liquid chromatography(HPLC),respectively.The results showed that the half-life of GA_(3)transformation in ultrapure water was 16.1–24.6 days at p H=2.0–8.0,with the lowest half-life occurring at p H=8.0 and highest half-life occurring at p H=3.3.Isomerized gibberellic acid(Iso-GA_(3))and gibberellenic acid(GEA)were the main transformation products with a little hydroxy gibberellic acid(OH-GA_(3)).In North China groundwater,the mass balance of GA_(3)and its products was 76.2%,including Iso-GA_(3)(58%),GEA(7.9%),GA_(3)(7.3%)and OH-GA_(3)(3%)after reaching transformation equilibrium.Using Gaussian 09 for chemical computation,it was found that the transformation mechanism of GA_(3)was dependent upon the bond energy and the stereochemical feature of its molecular structure.GA_(3)always isomerized from theγ-lactone ring due to the lowest bond energy between the oxygen terminus of theγ-lactone ring and A ring.While GA_(3)and its transformation products all had developmental toxicity,the predicated LC 50(96 hr)and LD 50 of the main products of GA_(3)were much lower than those of GA_(3),indicating GA_(3)would be transformed into higher toxicity derivatives in water environments,posing a significant health risk to humans and the environment.