OBJECTIVE:To study the metabolites,distribution,metabolic type and the possible activity of harpagide which is the active component from Scrophularia ningpoensis in rats in vivo. METHODS:4 SD rats were divided into blank group (ul-tra-pure water) and administration group (harpagide reference solution),2 in each group,ig,160 mg/kg,twice a day,for 3 d. Urine and feces were collected every 12 h before administration and the first administration;sample blood 8 mL was taken after 0.5,1 h of last administration;heart,liver,spleen,lung,kidney,stomach and small intestine were taken. The blood,urine,fe-ces and other tissue solutions were prepared,HPLC-MS was conducted to detect and identify the harpagide metabolites in rats in vi-vo and presume metabolic pathways,and PharmMapper software was used to predict metabolites activity. RESULTS:12 harpagide metabolites were identified in rats in vivo,the form of prototypes and metabolites were distributed in heart,liver,spleen,lung, kidney,stomach and small intestine. The metabolic type mainly included hydrolysis,dehydration,reduction,methylation,sul-fation,glucuronic acid binding,grade A coumaric acid binding,etc. The 12 compounds may have activities in the treatment of epi-lepsy,amyotrophic lateral sclerosis,diabetes,stroke,etc. CONCLUSIONS:Harpagide may be effective in the form of prototypes and metabolites. The study has provided basis for attributing the origins of metabolite,studying the effective form of S. ningpoensis clarifying its pharmacological mechanism and processing mechanism.

OBJECTIVE:To explore the effects and mechanism of extracts,active constituents and constituent combination of Sinopodophylli Fructus on cell proliferation of human breast cancer. METHODS:Acid phosphatase method was conducted to deter-mine the effects of 4 extracts [ethanol extract (Xc),petroleum ether extract from ethanol extract (Xp),ethyl acetate extract from ethanol extract (Xe),n-butanol extract from ethanol extract (Xz)],5 active constituents [podophyllotoxin (S1),deoxypodophyllo-toxin (S2),4-desmethyl deoxypodophyllotoxin (S3),8-isopentenyl kaempferol (S4),8,2′-diisoprenyl quercetin-3-methyl ether (S5)] and 3 active constituent combination [combination 1,S1-S2-S3-S4-S5 (2:4:1:4:32),Z1;combination 2,S2-S4 (1:1),Z2;combination 3,S3-S4(1:4),Z3] on the MDA-MB-231,MCF-7 cell proliferation;flow cytometry was adopted to detect the effects of above-mentioned samples on MDA-MB-231,MCF-7(T47D)cell cycle and mitochondrial membrane potential. RESULTS:The active constituent combination Z1 showed significant inhibitory effects on MDA-MB-231,MCF-7 cells,the half inhibitory concen-trations(IC50)were(0.27±0.2),(0.11±0.1)μg/mL;extracts Xc,Xp,Xe,active constituents S2,S4 and active constituent combi-nation Z2,Z3 showed relatively strong inhibitory effects on MDA-MB-231,MCF-7 (T47D) cell proliferation (IC50<15 μg/mL). Both extracts and active constituents can block MDA-MB-231,MCF-7 cell cycle in G2/M phase;all active constituents can block MDA-MB-231,T47D cell cycle in G0/G1 phase,and can reduce MDA-MB-231,T47D cell mitochondrial membrane potential. CONCLUSIONS:The active constituents and constituent combination of Sinopodophylli Fructus can inhibit cell proliferation of breast cancer by affecting cell cycle and mitochondrial mem-brane potential.

OBJECTIVE:To establish a method of simultaneous determination of inulicin and deacetylinulicin in Inulae Flos. METHODS:HPLC was performed on the column of Zorbax SB-C18 with mobile phase of acetonitrile-water(gradient elution)at a flow rate of 1.0 ml/min,the column temperature was 25 ℃,the detection wavelength was 210 nm,and the injection volume was 10 μl. RESULTS:The linear range was 0.000 2-0.005 μg/ml(r=0.999 8)for inulicin and 0.000 1-0.001 7 μg/ml(r=0.999 4)for deacetylinulicin;RSDs of precision,stability and reproducibility tests were lower than 2.0%;recoveries were 99.63%-103.56%(RSD=1.26%,n=9)and 95.98%-101.21%(RSD=1.84%,n=9),respectively. CONCLUSIONS:The method is simple,accurate and reliable,and can be used for the quality evaluation of Inulae Flos.

OBJECTIVE:To establish a method for the contents determination of 9 components in Guizhi decoction,and com-pare the effects of traditional decoction method and the extracting machine decoction method on these contents in Guizhi decoction. METHODS:HPLC was performed on the column of Agilent Zorbax SB-C18 with mobile phaseof acetonitrile- 0.1% phosphoric ac-id(gradient elution)at a flow rate of 1.0 ml/min,the detection wavelength was 230 nm,254 nm and 280 nm,the column tempera-ture was 25℃,and the injection volume was 10μl. RESULTS:The linear range was 0.410 2-210.0μg/ml for gallic acid(r=0.999 9), 0.994 0-254.5μg/ml for albiflorin(r=0.999 9),1.636 0-1 675.0μg/ml for paeoniflorin(r=0.999 9),0.988 3-506.0μg/ml for liquiri-tin(r=0.999 6),0.987 3-31.59 μg/ml for coumarin(r=0.999 5),0.486 8-124.6 μg/ml for cinnamic acid(r=0.999 5),2.458 0-314.6μg/ml for cinnamaldehyde(r=0.999 5),0.034 3-1.096 μg/ml for 2-methoxy cinnamaldehyde(r=0.999 8),and 1.711 0-219.0 μg/ml for glycyrrdhizic acid (r=0.999 7);RSDs of precision,stability and reproducibility tests were lower than 5%,recoveries were 93.56%-103.19%(RSD=4.00%,n=9)、101.51%-107.32%(RSD=2.21%,n=9)、95.08%-103.76%(RSD=2.87%,n=9)、100.82%-105.73%(RSD=1.85%,n=9)、85.08%-89.12%(RSD=1.40%,n=9)、92.31%-99.12%(RSD=2.71%,n=9)、99.17%-102.32%(RSD=1.24%,n=9)、100.15%-103.98%(RSD=1.18%,n=9)、99.93%-102.61%(RSD=1.03%,n=9). The content of total effective components from the extracting machine decoction method was 4 565μg/g,that from the traditional decoc-tion method was 2 742 μg/g.CONCLUSIONS:The method is simple,stable and reproducible,and can be used for the simultaneous determination of 9 componentsin Guizhi decoction. The contents of gallic acid,albiflorin and 2-methoxy cinnamaldehyde are first re-ported. The total effective components from the extracting machine decoction method are higher than that from the traditional decoc-tion method.

OBJECTIVE:To establish the HPLC characteristic chromatogram of pheretima,and compare the differences of the main ingredient contents of Guangdong pheretima and Shanghai pheretima and the chromatogram differences among pheretima and 3 other animal drugs. METHODS:Pheretima HPLC characteristic chromatogram method was adopted to determine the characteris-tic chromatograms of 16 Guangdong pheretima,8 Shanghai pheretima,3 eupolyphaga,3 hirudo and 3 catharsius. Similarity evalua-tion and t test were used to analyze the differences of chromatogram data of 5 animal drugs. RESULTS:The established HPLC char-acteristic chromatogram method firstly identified 11 common characteristic peaks,including 6 nucleosides,4 nucleobase and 1 ami-no acid;and it could be used for the identification of pheretima from eupolyphaga,hirudo and catharsius;the differences of main ingredient contents in the characteristic chromatogram of Guangdong pheretima and Shanghai pheretima were firstly studied. The contents of xanthine and adenosine in Guangdong pheretima were higher than Shanghai pheretima,while the contents of uridine, guanosine and 2′-deoxy guanosine in Shanghai pheretima were higher than Guangdong pheretima. A new index S,calculated by these 5 constituents,was successfully applied to distinguish the 2 kinds of pheretima. CONCLUSIONS:The characteristic chro-matogram can be used for the identification of pheretima,and can provide reference for the pharmacodynamic differences study of Guangdong pheretima and Shanghai pheretima.

The blood concentrations of the pharmacodynamic substances of traditional Chinese medicines (TCMs) are usually very low. How can they exert pharmacological actions, in which forms (original form, metabolite or the both) do they exert the actions. To answer these questions, we proposed a new concept ofEffective Formsof pharmacodynamic substances of TCMs and a hypothesis of additive effect of multiple constituents of TCMs. The hypothesis includes that the aggregate or summation of Effective Forms of pharmacodynamic substances of TCMs is the core material base of the effi-cacy of TCMs, and the additive effect of the blood concentrations of different Effective Forms is one part of the action mechanism. The additive effect of the different Effective Forms of a TCMs means an additive effect of numerous con-stituents or/and metabolites on a same target, and therefore the efficacy brought by the addition of the concentrations of all these compounds, which different from the synergy effect of multi-constituents on multi-targets. Studies on the disposition of TCMs showed that a constituent can be biotransformed to many metabolites (up to more than 50 metabolites);different constituents can produce the same metabolites;many metabolites (up to 10 compounds for each metabolite) are isomers or homologues; some constituents can be converted to each other in vivo; and some metabolites are bioactive. These com-pounds having the similar structure are likely to have the same pharmacological effects on the same target, which could provide experimental evidences for the concept ofEffective Formsand the hypothesis ofAdditive Effect. We suggest that the Effective Forms and Additive Effects of the pharmacodynamic substances of TCMs should be extensively investi-gated in the future, and the results of such researches will help us further understand the pharmacodynamic substances and action mechanism of TCMs, and give a new explanation 'Toxicities Scattering Effect' for 'Why the toxicities of TCMs are low', and propose a new strategy for quality control of TCMs.

OBJECTIVETo compare the chemical differences in 4 commercial specifications of Scutellaria Radix, research the affection of decayed central xylem part on the crude drug's chemical composition and provide scientific data for production, processing, sale and clinical applications of Scutellariae Radix.

METHODMacroscopical identification method was used for observation of different specifications of Scutellariae Radix, including Qinwang, Tiaoqin both in 1st class and 2nd class and inferior samples. HPLC fingerprint method was used to analyze chemically the decayed central xylem part and non-decayed part as well as complete sample, and the results were described by the relative peak area.

RESULTThe morphological characteristics of 4 specifications are greatly different from one another mainly in root diameters, root lengths and the proportions of decayed central xylem part in the root, and so the authors classified Qinwang and Tiaoqin in 1st class as Kuqin for all samples of them which have decayed central xylem; and classified Tiaoqin in 2nd class and the inferior samples as Ziqin, for having little decayed central xylem. The 4 specifications collected from the same producing area have similar HPLC fingerprint profile to one another, while they are different in relative peak area. The peak area ratios of aglycone to their glucuronide (baicalein/baicalin, wogonin/wogonoside, oroxylin A/oroxylin A-7-O-glucuronide) from Kuqin were significantly higher than those of Ziqin. The total area of the peaks in HPLC fingerprint chromatographs of decayed central xylem part were quite lower than that of non-decayed part, whereas peak areas of the characteristic peaks and the 3 peak area ratios of decayed central xylem were significantly higher than those of non-decayed part which could be used as characteristic parameters to distinguish Kuqin and Ziqin.

CONCLUSIONFour commercial specifications of Scutellariae Radix can be classified as Kuqin and Ziqin respectively according to morphological characteristics and the proportions of decayed central xylem part in the root. The chemical characteristics of Kuqin and Ziqin are different from each other, so it's worth clarifying the similarities and differences of Kuqin and Ziqin in future. The result in this research can be used as references for identification and quality control of Scutellariae Radix specifications, and investigation on effective components of Kuqin and Ziqin.

Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; standards ; Quality Control ; Scutellaria baicalensis ; chemistry

OBJECTIVETo assess the anti-nociception and anti-inflammation pharmacodynamics of Asarum heterotropoides var. mandshuricum and A. sieboldii.

METHODBoth the writhing test and hot plate test were conducted to assess the anti-nociceptive effect of Asarum and Xylene-induced mouse ear edema was conducted to assess the anti-inflammatory effect of Asarum.

RESULTTwelve samples of A. heterotropoides var. mandshuricum and A. sieboldii from different producing areas showed anti-nociceptive and anti-inflammatory effects. Specifically, 27% to 61% of the seven samples of A. heterotropoides var. mandshuricum showed anti-nociceptive effect and while 34% to 48% of A. sieboldi showed anti-nociceptive effect. The inflammatory inhibition rate of A. heterotropoides var. mandshuricum produced in six producing areas (38%-57%) is higher than that of A. heterotropoides var. mandshuricum produced in five producing areas (34%-48%). The same kind of Asarum produced in different areas showed significant differences. A. heterotropoides var. mandshuricum produced in Jilin province (38%-57%) showed better anti-nociceptive effect than sample produced in Heilongjiang province (34%) in writhing test. A. heterotropoides var. mandshuricum produced in Heilongjiang (43%) province showed a better anti-nociceptive effect than samples produced in Liaoning province (29%-36%) in hot plate test. A. sieboldii produced in Shaanxi province (47%-49%) showed a better anti-nociceptive effect than samples produced in Hubei province (40%) in writhing test. A. sieboldii produced in Shaanxi province (45%-59%) showed better anti-nociceptive effect than samples produced in Chongqing (40%) in hot plate test. A. heterotropoides var. mandshuricum produced in Jilin province (51%-63%) showed better anti-inflammatory effect than samples produced in Heilongjiang province (50%). In totality, the results from analysis of geoherbalism showed that famous-region A. heterotropoides var. mandshuricum and A. sieboldii had a better anti-nociception effect than Asarum produced in other producing areas, famous-region A. heterotropoides var. mandshuricum had a better effect than those produced in other producing areas in anti-inflammation. But famous-region A. sieboldii showed no obvious difference from those produced in other producing areas in anti-inflammation.

CONCLUSIONAll samples of Asarum showed anti-nociceptive and anti-inflammatory effects, but with significant differences among Asarum produced in different areas, indicating the eoherbalism to some extent.

Analgesics ; pharmacology ; Animals ; Anti-Inflammatory Agents ; pharmacology ; Asarum ; Female ; Male ; Mice ; Mice, Inbred ICR ; Plant Extracts ; pharmacology

To study the chemical constituents of Asarum himalaicum, fifteen compounds were isolated from a 70% ethanol extract by using a combination of various chromatographic techniques including column chromatography over silica gel, Sephadex LH-20, and semi-preparative HPLC. By spectroscopic techniques including 1H NMR, 13C NMR, and HR-ESI-MS, these compounds were identified as 4-demethoxyaristolochic acid BII (1), aristolochic acid I (2), aristolochic acid Ia (3), 7-hydroxyaristolochic acid I (4), aristolochic acid IV (5), aristolic acid II (6), debilic acid (7), aristololactam I (8), 9-hydroxyaristololactam I (9), 7-methoxyaristololactam IV (10), (2S)-narigenin-5, 7-di-O-beta-D-pyranosylglucoside (11), 4-hydroxybenzoic acid (12), 3, 4-dihydroxybenzoic acid (13), 4-hydroxycinnamic acid (14), and beta-sitosterol (15). All of these compounds (1-15) were obtained from A. himalaicum for the first time. Among them, 1 was identified as a new compound, and compounds 3-6, 9, 12-14 were isolated from Asarum genus for the first time. Since the kidney toxicity of aristolochic acids and aristololactams has been reported, the result of this investigation suggests that it should be cautioned to use A. himalaicum as a medicine.

OBJECTIVETo develop a method for the determination of harpagide and harpagoside in Scrophulariae Radix (Xuanshen) by HPLC-UV under double wavelength, and to study the changes of these two constituents during processing, and to set the limitation of harpagide and harpagoside contents in crude drug and sliced pieces of Xuanshen.

METHODThe analyses were performed on an Agilent Technologies ZORBAX SB-C18 (4.6 mm x 250 mm, 5 microm) eluted with acetonitrile-water (containing 0.03% phosphoric acid) in gradient model. The flow rate was 1.0 mL x min(-1) . The column temperature was 25 degrees C. The UV detector wavelength was set at 210 nm before 13 min and then changed to 280 nm.

RESULTHarpagide and harpagoside were separated well. The linear calibration curves were obtained over of 0.0549 - 1.46 microg for harpagide (r = 0.9999, n =7) ,0.0225 - 0.900 microg for harpagoside (r = 0.9998, n = 9). The recoveries ( +/- RSD)% were 98.1 (+/- 2.4)% for harpagide and 98.8 (+/- 4.3)% for harpagoside. The contents of harpagide were 0. 277% - 0.620%, harpagoside were 0.078% - 0.362% in Xuanshen, and harpagide were 0.276% - 1.059%, harpagoside were 0. 059% - 0.183% in sliced Xuanshen, respectively. After the processing of Scrophulariae Radix, the content of harpagide increases 13.7% - 96.0%, while harpagoside decreases 11.0%-73.9%.

CONCLUSIONThis method is simple, accurate, and can be used for the quality control of Scrophulariae Radix. We propose that the total content of harpagide and harpagoside in either crude drug or sliced pieces of Scrophulariae Radix should not be less than 0.45%.

Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; analysis ; isolation & purification ; Glycosides ; analysis ; isolation & purification ; Iridoid Glycosides ; Magnoliopsida ; chemistry ; Plant Roots ; chemistry ; Pyrans ; analysis ; isolation & purification ; Spectrophotometry, Ultraviolet ; methods