Research Progress on Purification Process, Content Determination and Pharmacological Action of Atractylodin

Atractylodis Rhizoma comes from the dry rhizome of Atractylis lancea or Atractylodes chinensis in the Compositae family,and it is suitable for preventing and treating diseases such as cold,edema,night blindness and rheumatic arthralgia.Atractylodin is the main active component extracted and isolated from Atractylodis Rhizoma.A large number of studies have found that atractylodin has excellent drug activity in improving gastrointestinal emptying,anti-inflammation,inhibiting malignant tumor and reducing blood lipid.In this paper,the purification process and pharmacological activity of Atractylodin were summarized to provide a theoretical basis for basic research,clinical application and further development and utilization of atractylodin.

Purification Process,Content Determination,Pharmacological Activity and Molecular Mechanism of Neogambogic Acid

Neogambogic acid is characterized by broad antitumor spectrum,good antitumor effect and low toxicity and side effects.This paper reviews the purification process,content determination and pharmacologic activity of neogambogic acid,in order to provide a theoretical reference for the research and application of neogambogic acid.

Research Progress on Purification Process Optimization and Content Determination of Zeaxanthin

At present,the purification process of zeaxanthin mainly includes organic solvent extraction,ultrasonic-assisted extraction and enzyme extraction,and the content determination technology mainly includes ultraviolet-spectrophotometry and high performance liquid chromatography.In this paper,the purification process and content determination technology of zeaxanthin in recent years are reviewed in order to provide ideas and theoretical basis for further research and application of zeaxanthin.

THE STUDY ON THE WATER SOLUBLE PEPTIDES IN PAPAVER SOMNIFERUM POLLEN EXTRACTION,PURIFICATION,SEQUENCE DETERMINATION AND SYNTHESIS

Four peptides PSPP1,PSPP2,PSPP3 and PSPP4 were purified from the water-extract of Papaver somniferum pollen.Their sequences,with 21,17,13 and 16 amino acid residues respectively,have been determined by Edman degradation-N-terminal dansylation.PSPP2, PSPP3 and PSPP4 were synthesized using solid phase method.The immunopromotive activities of PSPP1,PSPP2,PSPP3,PSPP4 and the initially separated sample PSPP have been also observed by the methods of counting erythrocyte rosette forming cells(ERFC) and T-lymphocyte transformation test in vitro.

Simultaneous Determination of Four Pesticide Residues in Fruit Juice by HPLC

[Objective] This study was conducted to develop a system for simultaneous determination of imidacloprid, diflubenzuron, thiabendazole and carbendazim in fruit juice by HPLC. [Method] Using acetonitrile as the extraction solvent, the pesticides in fruit juice were purified through a NH2 solid phase extraction （SPE） cartridge, then detected by HPLC. [Result] There was a good linear relationship between the peak area and the concentrations of imidacloprid, diflubenzuron, thiabendazole and carbendazim in a range of 0.05-5.0 μg/ml, and the linear correlation coefficient varied in a range of 0.999 0-0.999 8; the limit of detection for imidacloprid, diflubenzuron, thiabendazole and carbendazim was 0.003, 0.005, 0.003 and 0.007 mg/kg, respectively. The recovery rate of imidacloprid, diflubenzuron, thiabendazole and carbendazim standards added at three levels （0.1, 0.5 and 1.0 mg/kg） ranged from 82% to 107%, with RSD less than 4.5%. [Conclusion] The sensitivity, accuracy and precision of this method were able to meet the requirements for pesticide residue analysis.

Determination of Phytoene Content in Tomato Ketchup by HPLC

[Objective] The aim was to establish an HPLC method for determination of phytoene content in tomato ketchup. [Method] The chromatographic conditions were as follows： column, Agilent AT C18 （250 mm×4.6 mm, 5 μm）; mobile phase, methanol-THF （75：25）; detection wavelength, 287 nm; flow rate, 1.0 ml/min; column temperature, 30 ℃; sample size, 50 μl. [Result] There was a good linear relation- ship in the phytoene content range of 0.186-1.116μg. The average recovery rate was 103.8% with RSD of 1.47%. The phytoene content in the tomato ketchup sample was determined as 10.7 rag/100 g simple, accurate, rapid and reliable, and phytoene content in tomato ketchup. [Conclusion] The established method is it can be used for the determination of

Simultaneous Determination of Ticlopidine Hydrochloride and Nitrendipine in a New Tablet Formulation by HPLC

An accurate, simple and sensitive high performance liquid chromarographic (HPLC) method for simultaneous determination of ticlopidine hydrochloride and nitrendipine in a new tablet formulation is described. Chromatographic separation of ticlopidine hydrochloride and nitrendipine was achieved on a Hypersil BDS C18 column using a mobile phase consisting of acetonitrile-methanol-10 mmol/L ammonium acetate (60∶10∶30(V/V), pH 6.5) at a flow rate of 1.0 mL/min. Absorbance was monitored at 236 nm where both drugs have significant absorption. The proposed method was validated with respect to linearity, selectivity, accuracy, precision, and limits of detection and quantitation. The linear ranges for ticlopidine hydrochloride and nitrendipine were found to be 75-750 μg/mL and 1-10 μg/mL, respectively. The mean recoveries were 100.1%(S R=0.6%,n=9) for ticlopidine hydrochloride and 99.9%(S R=0.7%,n=9) for nitrendipine. The within-day precision and between-day precision for ticlopidine hydrochloride and nitrendipine were 0.63% and 0.89%, and 0.74% and 1.0%, respectively. The proposed HPLC method can be used for the simultaneous determination of both drugs in pharmaceutical preparations.

Development of RP-HPLC method for simultaneous determination of docetaxel and curcumin in rat plasma: Validation and stability

The purpose of the present research was to develop a suitable, simple, precise, accurate,robust, and reproducible RP-HPLC method for a reliable simultaneous quantification of docetaxel(DTX) and curcumin(CCM) in rat plasma samples using paclitaxel(PTX) as an internal standard. The samples were assayed by the Agilent 1260 Infinity HPLC instrument using a Capcell Pak C8 column(4.6 mm × 150 mm, 5 μm) under isocratic conditions. The mobile phase consisted of acetonitrile and triple distilled water(40/60, v/v) with a flow rate of 1.0 ml/min. The eluent was monitored at 230 nm for simultaneous measurement of curcumin and docetaxel. The method was validated by determining system suitability, selectivity, sensitivity, linearity, inter-day and intra-day precision, accuracy, robustness, and stability in accordance with the guidelines of the United States Food and Drug Administration(FDA).The developed chromatographic method proved to be simple, precise, accurate, robust and reproducible. Moreover, the samples showed stability at room temperature over a period of 48 h. Thus, this method would be employed for routine simultaneous quantification of docetaxel and curcumin in rat plasma samples.

Content Determination of Astragaloside Ⅳ in Astragalus from Three Different Regions by HPLC

[Objectives] The content of astragaloside in Astragalus membranaceus(Fisch.) Bge.var.mongholicus(Bge.) Hisao from three different regions was determined.[Methods] Referring to the method recorded in the Chinese Pharmacopoeia(2015 edition),the content of astragaloside IV in A.membranaceus was determined by HPLC.[Results] There were great differences in the astragaloside IV content of A.membranaceus among different regions.The content of astragaloside IV in A.membranaceus cultivated in Inner Mongolia was highest(0.155%),followed by that(0.143%) in A.membranaceus cultivated in Gansu,and the content of astragaloside IV in A.membranaceus cultivated in Shanxi was lowest(0.080%).The contents of astragaloside IV in A.membranaceus from different regions were all in line with the standard(not less than 0.040%) of Chinese Pharmacopoeia(2015 edition).[Conclusions]The content of astragaloside IV in A.membranaceus cultivated in three different regions met the medicinal standards.

RP-HPLC Method for the Simultaneous Determination of Lisinopril and NSAIDs in API, Pharmaceutical Formulations and Human Serum

High performance liquid chromatographic method was developed valdated and applied for the simultaneous determi- nation of lisinopril and NSAIDs in bulk, pharmaceuticals formulations and human serum. A Purospher star C18 (5 μm, 25 × 0.46 cm) column was used with mobile phase consisting of methanol: water: acetonitrile (80:17.5:2.5 v/v, pH 3.0) and quantitative evaluation was performed at 225 nm with a flow rate of 1.0 mL?min–1. The retention time of lisinopril was 2.2 min while naproxen, flurbiprofen, diclofenac sodium and mefenamic acid were found to be 4.0, 4.5, 5.0 and 6.7 min respectively. Suitability of this method for the quantitative determination of the drugs was proved by validation in accordance with the requirements laid down by International Conference on Harmonization (ICH) guidelines. The method is selective, precise, accurate and can be used for analysis of pharmaceutical preparations in quality control and clinical laboratories.

Study on HPLC Fingerprint and Simultaneous Determination of Three Active Components of Dipsaci Radix Collected in Different Time

[Objectives] To establish methods of HPLC fingerprint and simultaneous determination of the three bioactive components(namely, asperosaponin VI, loganin and sweroside) of Dipsaci Radix, and explore the quality situation of this crude medicine collected in various months in autumn. [Methods] The analysis was performed on Thermo BDS Hypersil C_(18)(4.6 mm×250 mm, 5 μm) column with a mixture of acetonitrile and 0.1% phosphoric acid as mobile phase in gradient mood at a flow rate of 1.0 mL/min, the column temperature was set at 30 ℃, and the detection wavelength was 220 nm. [Results] The analysis methods for HPLC fingerprint and determination of the three components in Dipsaci Radix had been established. The results showed that 12 batches of samples, which were collected in four different places from September to November, possessed the similarities of greater than 0.976. Through the quantitative analysis of asperosaponin VI, loganin and sweroside in Dipsaci Radix, it was found that the quality variation of this herbal medicine and the different collected months of autumn showed a low correlation. [Conclusions] The established methods of HPLC characteristic fingerprint and simultaneous determination of three glycosides provided a new way for quality analysis of Dipsaci Radix. It was preliminarily indicated that collecting this plant medicine in different months of autumn would not affect its quality.

Research on Purification Process and Pharmacological Action of Andrographolide

This paper reviews the purification process,content determination methods and pharmacological action of Andrographolide,aiming to provide new ideas for the subsequent study of Andrographolide and its related drug development and application.

Simultaneous Determination of Six Constituents in Cangbaiqutong Capsule by HPLC Wavelength Switching Technique

[Objectives] To establish a high performance liquid chromatography (HPLC) wavelength switching method for the simultaneous determination of content of six constituents (phellodendrine chloride, gentiopicrin, paeoniflorin, tetrandrine, berberine hydrochloride and paeonol) in Cangbaiqutong capsules, and provide a scientific basis for the comprehensive evaluation of the quality of Cangbaiqutong capsule.[Methods] The chromatographic column of Waters XSELECT CSH-C 18 (4.6 mm × 150 mm, 5 μm), the mobile phase of acetonitrile-0.1% phosphate solution, gradient elution (0-15 min,10%-18% A;15-30 min,18%-50% A;30-35 min, 50%-10% A);the flow rate of 1.0 mL/min, wavelength switching of 284 (0-7 min, phellodendrine), 274 (7-10 min, gentiopicrin), 230 (10-14 min, paeoniflorin) and 274 nm (14-35 min, tetrandrine, berberine hydrochloride, paeonol), the injection volume of 10 μL.[Results] There was a good linear relationship between the area of chromatographic peak and the injection volume of phellodendrine chloride, gentiopicrin, paeoniflorin, tetrandrine, berberine hydrochloride and paeonol in the range of 0.150-1.504, 0.768-7.680, 1.096-10.960, 0.220-2.200, 0.296-2.956, 0.0345-0.345 μg, respectively;the average recovery rates ( n =6) were 98.3%, 99.2%, 98.8%, 98.8%, 99.1% and 98.2%, respectively;the RSD value was 1.32%, 1.46%, 1.08%, 1.31%, 1.26% and 1.21%, respectively.[Conclusions] The method can be used to determine many kinds of constituents at the same time, and the operation is simple, accurate and reproducible, and can be used for the quality control of compound Cangbaiqutong capsules.

Determination of Chlorogenic Acid,Geniposide,Total Flavonoids and Total Triterpenes in Wulan-13

[Objectives]This study was conducted to establish a method for the determination of chlorogenic acid,geniposide,total flavonoids and total triterpenes in Wulan-13.[Methods]The contents of chlorogenic acid and geniposide were determined by HPLC,and the contents of total flavonoids and total triterpenes were determined by an ultraviolet spectrophotometer.[Results]There was a good linear relation between the mass of chlorogenic acid reference substance and the peak area in the range of 0.05-0.45μg,and the regression equation was Y=2524.1X+3.1943,(r=0.9998).A good linear relationship was found between the mass of gardenoside reference substance and the peak area in the range of 0.776-6.984μg,and the regression equation was Y=1670.5X+64.804,(r=0.9998).There was also a good linear relation between the mass of rutin reference substance and its absorbance in the range of 0.00808-0.04848 mg,and the regression equation was Y=12.916X+0.014,(r=0.999).The mass of oleanolic acid reference substance had a good linear relation with its absorbance in the range of 0.00418-0.0209 mg,and the regression equation was Y=51.89X-0.0839,(r=0.9991).[Conclusions]The content determination method is simple,reliable and reproducible,and suitable for controlling the contents of chlorogenic acid,geniposide,total flavonoids and total triterpenes in Wulan-13.

Development of a novel stability indicating RP-HPLC method for quantification of Connexin43 mimetic peptide and determination of its degradation kinetics in biological fluids

Connexin43 mimetic peptide （Cx43MP） has been intensively investigated for its therapeutic effect in the management of inflammatory eye conditions, spinal cord injury, wound healing and iscbemia-induced brain damage. Here, we report on a validated stability-indicating reversed-phase high performance liquid chromatography （RP-HPLC） method for the quantification of Cx43MP under stress conditions. These included exposure to acid/base, light, oxidation and high temperature. In addition, the degradation kinetics of the peptide were evaluated in bovine vitreous and drug-free human plasma at 37℃. Detection of Cx43MP was carried out at 214 nm with a retention time of 7.5 min. The method showed excellent linearity over the concentration range of 0.9-250μg/mL （R2a0.998）, and the limits of detection （LOD） and quantification （LOQ） were found to be 0.90 and 2.98 μg/mL, respectively. The accuracy of the method determined by the mean percentage recovery at 7.8, 62.5 and 250μg/mL was 96.79%, 98.25% and 99.06% with a RSD of〈 2.2%. Accelerated stability studies revealed that Cx43MP was more sensitive to basic conditions and completely degraded within 24 h at 37℃（0% recovery） and within 12 h at 80℃ （0.34% recovery）. Cx43MP was found to be more stable in bovine vitreous （t1/2 slow= 171.8 min） compared to human plasma （t1/2slow = 39.3 min） at 37℃ according to the two phase degradation kinetic model. These findings are important for further pre-clinical development of Cx43MP.

RP-HPLC method for the simultaneous determination of daidzein,genistein and formonetin in Solanum Lyratum Thunb

A rapid method for the simultaneous determination of daidzein,genistein and formonetin in solanum Lyratum Thunb by high performance liquid chromatography(HPLC)was developed.Separation was achieved on a Diamonsil C18 column(250 mm×4.6 mm,5 μm)with isocratic elution,using a mobile phase of methanol-tetrahydrofuran-water(44∶3∶53,v/v).The wavelength was set at 260 nm and column was maintained at 35 ℃.The linear ranges of daidzein,genistein and formonetin were 1.0-40.0,0.1-4.0 and 0.1-4.0 μg/mL,respectively.The average recoveries were between 98.4% and 101.3%.This method could be used for the quality control of Solanum lyratum Thunb due to its simplification,reliability,rapidity and excellent precision.

Determination and pharmacokinetic study of catechin in rat plasma by HPLC

A high performance liquid chromatographic method was developed and validated for the quantitative determination of catechin in rat plasma and its pharmacokinetic study after intragastric administration of Catechu and Xiongdanjiangre Wan into SD rats. Plasma samples were prepared by protein precipitation using methanol-5% aqueous zinc sulfate （70：30, v/v） as precipitant. Chromatographic separation was achieved on Hypersil Cl8 column （250 mm~ 4.6 mm, 10 pm） with acetonitrile-water-triethylamine （6：94：0.3, v/v/v, pH 4.0＋0.1, adjusted with phos- phoric acid） as mobile phase, followed by a UV detection at 207 nm. Good linearity was obtained over the range of 0.143-7.15 mg/L of catechin, with correlation coefficient of 0.9992. The method was simple, sensitive, accurate and reproducible and＇ has been successfully applied to the pharmacokinetic study of catechin in rat plasma.

Simultaneous Determination of Four Ingredients in Guanxin Danshen Tablets by RP-HPLC

[Objectives]This study aimed to establish a method for simultaneous determination of four ingredients in Guanxin Danshen tablets by RP-HPLC.[Methods]HPLC chromatography was adopted with column of Thermo Hypersil-Keystone C 18 column(4.6 mm×250 mm,5μm),mobile phase of acetonitrile(A)-0.03 mol/L ammonium acetate(pH adjusted to 2.4 with formic acid)(B),gradient elution(0-5 min,5%A;5-10 min,5%A→19%A;10-40 min,19%A;40-68 min,19%A→36%A;68-90 min,36%A→95%A),flow rate of 1.0 mL/min and column temperature of 25℃.[Results]The content of salvianic acid A sodium,protocatechuic aldehyde,salvianolic acid B and tanshinone II A showed good linear relationship with chromatographic peak area in the range of 3.310-18.66,0.03950-0.2370,0.7500-4.500,0.05920-0.3550μg,respectively.The recovery rate(n=6)was 101.75%,96.86%,104.15%and 99.03%,respectively,and the RSD was 1.52%,2.81%,1.80%,and 1.37%respectively.The established method has good precision,reproducibility and stability.[Conclusions]This method can be used for the quality control of multiple ingredients of Guanxin Danshen tablets.

Determination of Oleanolic Acid in Achyranthes aspera L. of Different Production Areas by HPLC

[Objectives] To determine the content of oleanolic acid in 10 batches of Achyranthes aspera L. from different production areas,and to establish a high performance liquid chromatography( HPLC) method for the determination of oleanolic acid in A. aspera. [Methods]Agilent C18 liquid chromatography column( 250 mm × 4. 6 mm,5 μm) was used for gradient elution with acetonitrile∶ water = 61∶ 39 as the mobile phase. The flow rate was 1 mL/min,the detection wavelength was 210 nm,and the column temperature was 35℃. [Results] The oleanolic acid injection volume showed a good linear relationship with the peak area in the range of 0. 382-7. 640 μg. The linear equation of oleanolic acid was: A = 530. 76 C,R = 1. 000 0; the range of sample recovery rate was 96. 03%-102. 73%,and the RSD value was 2. 16%( n =9). The content of the oleanolic acid was the highest in A. aspera produced in Guilin City of Guangxi( 0. 92%),and the lowest oleanolic acid content was in sample of Sitang in Nanning City of Guangxi( 0. 27%). It is recommended that the oleanolic acid content of A. aspera should not be lower than 0. 20%. [Conclusions]The HPLC method is accurate,reliable,easy to operate,and has good resolution. It is suitable for the determination of the content of oleic acid in A. aspera. It can provide reference for the quality control and standard drafting of A. aspera.

Determination of Naringin Content in Peel of Guangxi Citrus maxima (Burm.) Merr by HPLC

[Objectives]To establish a method for determining the naringin content in the peel of Guangxi Citrus maxima(Burm.)Merr.[Methods]The high performance liquid chromatography(HPLC)method was applied.The chromatographic column was Agilent HC-C18(4.6 mm×250 mm,5μm);the mobile phase was acetonitrile-0.1%phosphoric acid(18∶82);the flow rate was 1.0 mL/min;the column temperature was 25℃;the detection wavelength was 283 nm.[Results]Naringin showed a good linear relationship in the range of 0.164-3.27μg,r=0.9999.The average recovery rate was 98.66%,and RSD=1.80%(n=6).[Conclusions]This method is simple,feasible,reproducible,and accurate,so it can be used for the determination of naringin content in the peel of Guangxi C.maxima.