Bear Bile Powder Ameliorates LPS-Induced Acute Lung Injury by Inhibiting CD14 Pathway and Improving Intestinal Flora: Exploration of "Fei (Lung)-Dachang (Large Intestine) Interaction".

OBJECTIVE: To explore the effect of bear bile powder (BBP) on acute lung injury (ALI) and the underlying mechanism. METHODS: The chemical constituents of BBP were analyzed by ultra-high-pressure liquid chromatography-mass spectrometry (UPLC-MS). After 7 days of adaptive feeding, 50 mice were randomly divided into 5 groups by a random number table (n=10): normal control (NC), lipopolysaccharide (LPS), dexamethasone (Dex), low-, and high-dose BBP groups. The dosing cycle was 9 days. On the 12th and 14th days, 20 µL of Staphylococcus aureus solution (bacterial concentration of 1 × 10-7 CFU/mL) was given by nasal drip after 1 h of intragastric administration, and the mice in the NC group was given the same dose of phosphated buffered saline (PBS) solution. On the 16th day, after 1 h intragastric administration, 100 µL of LPS solution (1 mg/mL) was given by tracheal intubation, and the same dose of PBS solution was given to the NC group. Lung tissue was obtained to measure the myeloperoxidase (MPO) activity, the lung wet/dry weight ratio and expressions of CD14 and other related proteins. The lower lobe of the right lung was obtained for pathological examination. The concentrations of inflammatory cytokines including interleukin (IL)-6, tumour necrosis factor α (TNF-α ) and IL-1ß in the bronchoalveolar lavage fluid (BALF) were detected by enzyme linked immunosorbent assay, and the number of neutrophils was counted. The colonic contents of the mice were analyzed by 16 sRNA technique and the contents of short-chain fatty acids (SCFAs) were measured by gas chromatograph-mass spectrometer (GC-MS). RESULTS: UPLC-MS revealed that the chemical components of BBP samples were mainly tauroursodeoxycholic acid and taurochenodeoxycholic acid sodium salt. BBP reduced the activity of MPO, concentrations of inflammatory cytokines, and inhibited the expression of CD14 protein, thus suppressing the activation of NF-κB pathway (P<0.05). The lung histopathological results indicated that BBP significantly reduced the degree of neutrophil infiltration, cell shedding, necrosis, and alveolar cavity depression. Moreover, BBP effectively regulated the composition of the intestinal microflora and increased the production of SCFAs, which contributed to its treatment effect (P<0.05). CONCLUSIONS: BBP alleviates lung injury in ALI mouse through inhibiting activation of NF-κB pathway and decreasing expression of CD14 protein. BBP may promote recovery of ALI by improving the structure of intestinal flora and enhancing metabolic function of intestinal flora.

A novel strategy for active compound efficacy status identification in multi-tropism Chinese herbal medicine (Scutellaria baicalensis Georgi) based on multi-indexes spectrum-effect gray correlation analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Bioactive substance identification is always the focal point and the main challenge in Chinese herbal medicine (CHM). Most CHM present multiple efficacies and multiple tropisms, which has improved the application accuracy of CHM, and is worthy of further study. In this article, the concept of "multi-tropism efficacy of CHM" has been proposed for the first time. In addition, it is hypothesized that the different components in CHM can be classified based on their efficacy status. AIM OF THE STUDY: The spectrum-effect relationship between the fingerprint and efficacy was established to identify the efficacy status of components. This provided a practical, efficient and accurate way to identify the bioactive substances from a complex CHM system. MATERIALS AND METHODS: The network pharmacology approach was applied to preliminarily analyze the potential antibacterial compounds and mechanisms of HQ. Furthermore, its chemical fingerprint was established and the characteristic peaks were identified by LC-MS/MS. The antibacterial and anti-inflammatory bioactivities of HQ were determined to evaluate its pharmacological effect of heat-clearing and detoxification, and its anticoagulation activity was determined to evaluate its heat-clearing and tocolysis effects. The spectrum-effect relationships were assessed by gray correlation analysis to discriminate the status of active components in HQ with different efficacies. RESULTS: Network pharmacology analysis revealed apigenin, wogonin, baicalein, acacetin, ß-sitosterol, baicalin, eugenol, moslosooflavone, palmitic acid, oroxylin-A 7-O-glucuronide, and scutevulin as the potential active compounds responsible for the efficacy of HQ against both E. coli and S. aureus. The spectrum-effect relationship was utilized to reveal the orientation activities, with the results as follows: 1) The main basic-efficacy components in HQ with antibacterial, anti-inflammatory, and anticoagulant effects were P5, P8, P9, P15, P18, P19, P20; while the general basic-efficacy components were P2, P3, P6, P7, P11, P14, P21, P22, P28. 2) The main efficacy-oriented components in HQ with antibacterial effects on E. coli were P1, P12, P17, while the general efficacy-oriented compound was P10, P24, P25, P26, P27; the main efficacy-oriented in HQ with antibacterial effects on S. aureus were P14 and the general efficacy-oriented components were P1, P12, P26, P29, P30, respectively. 3) The main efficacy-oriented components with anti-inflammatory activity were P14, P24, P25, P27, and P30, while the general efficacy-oriented components were P13, P23, P26. 4) The main efficacy-oriented compounds in HQ with effects on anticoagulation were P6 and P22; these acted by prolonging APTT through the intrinsic coagulation pathway and PT through the extrinsic coagulation pathway, respectively. 5) The pharmacodynamic status classification of Scutellaria baicalensis ingredients were confirmed by nine reference compounds exemplarily. CONCLUSION: This work established a novel strategy for active compound efficacy status identification in multi-tropism Chinese herbal medicine (Scutellaria baicalensis Georgi) based on multi-indexes spectrum-effect gray correlation analysis, the method is scientific feasible and can be applied to the effective substances identification and quality control of other CHM.

Protective Effect of Fresh/Dry Dandelion Extracts on APAP-Overdose-Induced Acute Liver Injury.

OBJECITVIE: To compare the liver protective activity of fresh/dried dandelion extracts against acetaminophen (APAP)-induced hepatotoxicity. METHODS: Totally 90 Kunming mice were randomly divided into 10 groups according to body weight (9 mice for each group). The mice in the normal control and model (vehicle control) groups were administered sodium carboxymethyl cellulose (CMC-Na, 0.5%) only. Administration groups were pretreated with high and low-dose dry dandelion extract (1,000 or 500 g fresh herb dried and then decocted into 120 mL solution, DDE-H and DDE-L); low-, medium- and high-dose dandelion juice (250, 500, 1,000 g/120 mL, DJ-L, DJ-M, and DJ-H); fresh dandelions evaporation juice water (120 mL, DEJW); dry dandelion extract dissolved by pure water (1 kg/120 mL, DDED-PW); dry dandelion extract dissolved by DEJW (120 g/120 mL, DDED-DEJW) by oral gavage for 7 days at the dosage of 0.5 mL solution/10 g body weight; after that, except normal control group, all other groups were intraperitonealy injected with 350 mg/kg APAP to induce liver injury. Twenty hours after APAP administration, serum and liver tissue were collected and serum alanine aminotransferase (AST), aspartate transaminase (ALT), alkaline phosphatase (AKP), malondialdehyde (MDA), glutathione (GSH), superoxide dismutase (SOD) activities were quantified by biochemical kits; tumor necrosis factor (TNF-α), interleukin (IL)-2, and IL-1 ß contents in liver tissue were determined by enzyme linked immunosorbent assay kits. Histopathological changes in liver tissues were observed by hematoxylin and eosin staining; TUNEL Assay and Hoechst 33258 staining were applied for cell apoptosis evaluation. The expressions of heme oxygenase-1 (HO-1), nuclear factor erythroid-2-related factor 2 (Nrf-2), caspase-9, B-cell leukemia/lymphoma 2 (Bcl-2), Bax and p-JNK were determined by Western blot analysis. RESULTS: Pretreatment with fresh dandelion juice (FDJ, including DJ-L, DJ-M, DJ-H, DEJW and DDED-DEJW) significantly decreased the levels of serum ALT, AST, AKP, TNF-α and IL-1ß compared with vehicle control group (P<0.05 or P<0.01). Additionally, compared with the vehicle control group, FDJ decreased the levels of hepatic MDA and restored GSH levels and SOD activity in livers (P<0.05 or P<0.01). FDJ inhibited the overexpression of pro-inflammatory factors including cyclooxygenase-2 and inducible nitric oxide synthase in the liver tissues (P<0.05 or P<0.01). Furthermore, Western blot analysis revealed that FDJ pretreatment inhibited activation of apoptotic signaling pathways via decreasing of Bax, and caspase-9 and JNK protein expression, and inhibited activation of JNK pathway (P<0.05 or P<0.01). Liver histopathological observation provided further evidence that FDJ pretreatment significantly inhibited APAP-induced hepatocyte necrosis, inflammatory cell infiltration and congestion. CONCLUSIONS: FDJ pretreatment protects against APAP-induced hepatic injury by activating the Nrf-2/HO-1 pathway and inhibition of the intrinsic apoptosis pathway, and the effect of fresh dandelion extracts was superior to dried dandelion extracts in APAP hepatotoxicity model mice.

[Preparation of nanoemulsion spray from Moslae Herba volatile oil and its antibacterial activity].

Moslae Herba is a commonly used aromatic Chinese medicinal with volatile oil as the main effective component and exhibits broad-spectrum antibacterial and antiviral effects. However, the irritation and instability of Moslae Herba volatile oil necessitate the preparation into a specific dosage form. In this study, the steam distillation method was employed to extract the Moslae Herba volatile oil. The content of thymol and carvacrol in Moslae Herba volatile oil was determined by HPLC as(0.111 9±0.001 0) and(0.235 4±0.004 7) mg·mL~(-1), respectively. Pseudo-ternary phase diagrams and surfactants compounding were applied in the selection of the optimal excipients(surfactant and cosurfactant). On this basis, a nanoemulsion was prepared from the Moslae Herba volatile oil and then loaded into pressure vessels to get sprays, whose stability and antibacterial activity were evaluated afterward. With clarity, viscosity, smell and body feeling as comprehensive indexes, the optimal formulation of the Moslae Herba volatile oil nanoemulsion was determined as follows: Moslae Herba volatile oil∶peppermint oil∶cremophor EL∶absolute ethanol∶distilled water 7.78∶1.58∶19.26∶6.15∶65.23. The as-prepared nanoemulsion was a light yellow transparent liquid, with Tyndall effect shown under the irradiation of parallel light. It has the pH of 5.50, conductivity of 125.9 µS·cm~(-1), average particle size of 15.45 nm, polydispersity index(PDI) of 0.156, and Zeta potential of-17.9 mV. Under a transmission electron microscope, the Moslae Herba volatile oil nanoemulsion was presented as regular spheres without adhesion and agglomeration. Stability test revealed that the Moslae Herba volatile oil nanoemulsion was stable at 4-55 ℃, which was free from demulsification and stratification within 30 days. After the centrifugation at 12 000 r·min~(-1) for 30 min, there was no stratification either. The nanoemulsion had good inhibitory effects on Escherichia coli, Staphylococcus aureus and resistant S. aureus strains, with the minimum inhibitory concentrations of 0.39, 3.12 and 1.56 mg·mL~(-1), respectively. The above results demonstrated that the nanoemulsion was prepared feasibly and showed stable physical and chemical properties and good antibacterial effects. This study provides a practicable technical solution for the development of anti-epidemic and anti-infection products from Moslae Herba volatile oil.

Dandelion polyphenols protect against acetaminophen-induced hepatotoxicity in mice via activation of the Nrf-2/HO-1 pathway and inhibition of the JNK signaling pathway.

We investigated the liver protective activity of dandelion polyphenols (DP) against acetaminophen (APAP; Paracetamol)-induced hepatotoxicity. Mice were acclimated for 1 week and randomly divided into the following groups (n = 9 per group): Control, APAP, APAP + DP (100 mg·kg-1), APAP + DP (200 mg·kg-1), and APAP + DP (400 mg·kg-1) groups. Mice were pretreated with DP (100, 200, and 400 mg·kg-1) by oral gavage for 7 d before being treated with 350 mg·kg-1 APAP for 24 h to induced hepatotoxicity. Severe liver injury was observed, and hepatotoxicity was analyzed after 24 h by evaluation of biochemical markers, protein expressions levels, and liver histopathology. Pretreatment with DP was able to restore serum liver characteristics (aspartate transaminase, AST; alanine aminotransferase, ALT; alkaline phosphatase, AKP), improve redox imbalance (superoxide dismutase, SOD; glutathione, GSH; malondialdehyde, MDA), and decrease inflammatory factors (tumor necrosis factor-α, TNF-α; interleukin-1ß, IL-1ß). Pretreatment with DP also significantly inhibited the expression levels of nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Furthermore, DP pretreatment could inhibit the apoptosis of liver cells caused by APAP through up-regulation of Bcl-2 and down-regulation of Bax and caspase-9 protein. DP also down-regulated p-JNK protein expression levels to inhibit APAP-induced mitochondrial oxidative stress and up-regulated the expression of Nrf-2 and its target gene HO-1. The histopathological staining demonstrated that DP pretreatment could inhibit APAP-induced hepatocyte infiltration, congestion, and necrosis. Our results demonstrate that DP pretreatment could protect against APAP-induced hepatic injury by activating the Nrf-2/HO-1 pathway and inhibition of the intrinsic apoptosis pathway.

Novel application of neural network modelling for multicomponent herbal medicine optimization.

The conventional method for effective or toxic chemical substance identification of multicomponent herbal medicine is based on single component separation, which is time-consuming, labor intensive, inefficient, and neglects the interaction and integrity among the components; therefore, it is necessary to find an alternative routine to evaluate the components more efficiently and scientifically. In this study, sodium aescinate injection (SAI), obtained from different manufacturers and prepared as "components knockout" samples, was chosen as the case study. The chemical fingerprints of SAI were obtained by high-performance liquid chromatography to provide the chemical information. The effectiveness and irritation of each sample were evaluated using anti-inflammatory and irritation tests, and then "Gray correlation" analysis (GCA) was applied to rank the effectiveness and irritability of each component to provide a preliminary judgment for product optimization. The prediction model of the proportions of the expected components was constructed using the artificial neural network. The results of the GCA showed that the irritation sorting of each SAI component was in the order of B > A > G > J > I > H > D > F > E > C and the effectiveness sorting of SAI components was in the order of D > C > B > A > F > E > H > I > G > J; the predictive proportion of SAI was optimized by the BP neural network as A: B: C: D: E: F = 0.7526: 0.5005: 5.4565: 1.4149: 0.8113: 1.0642. This study provided a scientific, accurate, reliable, and efficient approach for the proportion optimization of multicomponent drugs, which has a good prospect of popularization and application in product upgrading and development of herbal medicine.

Anti-inflammatory and analgesic effects of Bi-yuan-ling granules.

Bi-yuan-ling granule (BLG) is a traditional Chinese medicine compound composed mainly of baicalin and chlorogenic acid. It has been demonstrated to be clinically effective for various inflammatory diseases such as acute rhinitis, chronic rhinitis, atrophic rhinitis and allergic rhinitis. However, the underlying mechanisms of BLG against these diseases are not fully understood. This study aimed to explore the anti-inflammatory and analgesic activities of BLG, and examine its protective effects on mouse acute lung injury (ALI). The hot plate test and acetic acid-induced writhing assay in Kunming mice were adopted to evaluate the pain-relieving effects of BLG. The anti-inflammatory activities of BLG were determined by examining the effects of BLG on xylene-caused ear swelling in Kunming mice, the cotton pellet-induced granuloma in rats, carrageenan-induced hind paw edema and lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. The results showed that BLG at 15.5 mg/g could significantly relieve the pain by 82.5% (P<0.01) at 1 h after thermal stimulation and 91.2% (P<0.01) at 2 h after thermal stimulation. BLG at doses of 7.75 and 15.5 mg/g reduced the writhing count up to 33.3% (P<0.05) and 53.4% (P<0.01), respectively. Additionally, the xylene-induced edema in mice was markedly restrained by BLG at 7.75 mg/g (P<0.05) and 15.5 mg/g (P<0.01). BLG at 5.35 and 10.7 mg/g significantly reduced paw edema by 34.8% (P<0.05) and 37.9% (P<0.05) at 5 h after carrageenan injection. The granulomatous formation of the cotton pellet was profoundly suppressed by BLG at 2.68, 5.35 and 10.7 mg/g by 15.4%, 38.2% (P<0.01) and 58.9% (P<0.001), respectively. BLG also inhibited lung W/D ratio and the release of prostaglandin E2 (PGE2) in ALI mice. In addition, the median lethal dose (LD50), median effective dose (ED50) and half maximal inhibitory concentration (IC50) of BLG were found to be 42.7, 3.2 and 12.33 mg/g, respectively. All the findings suggest that BLG has significantly anti-inflammatory and analgesic effects and it may help reduce the damage of ALI.

[A novel method for testing sterility of injections based on biothermodynamics].

This study aims at trying to establish a novel method of sterility test for injections based on biothermodynamics, in order to overcome the deficiencies of routine sterility tests such as long detecting cycle, low sensitivity and prone to misjudgments. A biothermodynamics method was adopted to rapidly detect the microorganism contamination of injections by monitoring the heat metabolism during the growth of microbe. The growth rate equal to or greater than zero and the heat power difference of P(i) and P(0) with three folds higher than the noise of baseline were chosen as indexes to study the heat change rule of microbe. In this way, the effectiveness of the new method to detect strains required by conventional sterility test or in injection samples was also investigated. Results showed that the Gram-positive bacteria, Gram-negative bacteria and fungi demanded by sterility testing methodology could be detected by biothermodynamics method within 10 hours, with the sensitivity lower than 100 CFU x mL(-1). Meanwhile, this method was successfully applied to the sterility test of Compound Yinchen injection (FFYC), Shuanghuanglian powder injection (SHL) and Compound Triamcinolone injection (TAND) which were sterilized with different degrees. Therefore, the biothermodynamics method, with advantages of fast detection and high sensitivity, could be a complementary solution for conventional sterility tests.

[Compatibility evaluation of Chinese medicines injections based on isothermal titration calorimetry].

The paper is to report the establishment of a method for quickly evaluating compatibility of Chinese medicines injections. Isothermal titration calorimetry (ITC) was used to evaluate compatibility of Yiqifumai (YQFM) and vitamin C injection (Vc)/5% glucose injection (5% GS). The diversification of Gibbs free energy (deltaG), enthalpy (deltaH) and entropy (deltaS) were used to decide reaction types of colliquefaction procedures of different injections. The reactive profiles were used to determine signs and quantity of heat. And high performance liquid chromatography (HPLC) was used as a supportive method for ITC. Then, feasible binding sites were analyzed based on the information of spatial structures of major compositions. During the colliquefaction procedure of YQFM and Vc, [deltaH] > T[deltaS], so, the reaction is enthalpy-driving. And the reactive profile showed that a big deal of heat was given out during the procedure. Obviously, chemical reactions happened and the major compositions changed. On the other side, the reaction of YQFM mixed with 5% GS was entropy-driving, because [deltaH] < T[deltaS]. The reactive profile showed there was a little heat gave out. So, non-chemical reactions happened, and the major compositions did not change. The conformity existed between the results of ITC and HPLC. ITC could be used to evaluate the compatibility of Chinese medicines injections because of the advantages of ITC, such as real time, fast, sensitive and having more parameters.

Applications of microcalorimetry in the antibacterial activity evaluation of various Rhizoma coptidis.

CONTEXT: Various Rhizoma coptidis are commonly used in clinical applications due to their strong antibacterial effect. OBJECTIVE: The antibacterial effect of various R. coptidis were evaluated by microcalorimetry together with statistical analysis. MATERIALS AND METHODS: The dried roots of Coptis chinensis Franch, Coptis deltoidea C.Y. Cheng et Hsiao, and Coptis teeta Wall (Ranunculaceae) were identified by Prof. Xiao-He Xiao. The herbs were tested using thermal activity monitor (TAM) air isothermal microcalorimeter, ampoule mode, to evaluate the biological activity. The dynamic energy metabolism information about the Staphylococcus aureus growth at 37°C was analyzed with principal component analysis. RESULTS: From the principle component analysis on quantitative parameters, the action of various R. coptidis on S. aureus growth at 37°C could be easily evaluated by analyzing the change of values of the two main parameters: the growth rate constant of first stage (k(1)) and the maximum heat rating of the first stage (P(1)). And the results of paired t-test show that there were no notable differences between inhibitory ratios calculated with k(1) and that of P(1). Using microdilution method, the minimal inhibitory concentrations (MIC) of C. chinensis, C. deltoidea, and C. teeta were detected, and they were 77.8 ± 2, 116.6 ± 4, 93.3 ± 2 µg/mL, respectively. DISCUSSION AND CONCLUSION: This work provided a useful method for the evaluation of antibacterial activity of various R. coptidis.

Study on the cold and hot properties of medicinal herbs by thermotropism in mice behavior.

It is a common sense that chewing a mint leaf causes a cold feeling, while masticating a piece of ginger root is associated with a hot sensation. The Traditional Chinese Medicine has termed this phenomenon as cold and hot properties of herbs and applied them in treating certain human diseases successfully for thousands of years. Here, we have developed an Animal Thermotropism Behavior Surveillance System, and by using this device and other approaches, we not only verified the existence of, but also characterized and quantitated the cold and hot properties of medicinal herbs in animal behavioral experiments. The results suggested that the hot and cold properties of herbal drugs indeed correlated with the alteration of animal behavior in search for residence temperature.

[Quality control of shuanghuanglian freeze-dried powder for injection based on its HPLC-ELSD fingerprints and biological profiles].

This paper is aimed to provide the methods of quality control and bioassay of traditional Chinese medicine injections including bioassay method. Shuanghuanglian freeze-dried powder for injection (SFPI) was chosen as study object. HPLC-ELSD fingerprints of SFPI had been established and the samples were differentiated by similarity calculation. Meanwhile, biological profiles of SFPI on Escherichia coli had been established by microcalorimetry. The similarity values were calculated using the correlation coefficient, based on quantitative thermo-kinetic parameters (T2m, Tj, I%). The results indicated that HPLC-ELSD fingerprints, which showed content changes of chemical components, could not monitor minimal variation of different samples, especially that of biological pollutants, while biological profiles could sensitively detect antibiotic activity alterations of the samples, which were kept under specific conditions. In conclusion, characterized by two-dimension, microcalorimetry could supply thermograms as biological profiles characterized to describe the bioactivity of drugs. This study could clearly demonstrate that the correlative detection was proposed as an efficient strategy for quality control of SFPI, based on HPLC-ELSD fingerprints and biological profiles, which could detect quality fluctuation of samples early and quickly and predict the potential adverse drug events (ADE) for ensuring clinical safety.

[Hemagglutination activity of radix isatidis detected by microcalorimetry].

In this study, microcalorimetry was adopted to establish a novel method for detecting the hemagglutination process of Radix Isatidis (Banlangen in Chinese, BLG), and to evaluate the hemagglutination activity diversity of BLG from various habitats. The hemagglutination biothermokinetics curves of positive reagent (phytohemagglutinin, PHA) and 8 batches BLG from different regions of the hemagglutination with 20% rabbit erythrocyte were recorded by microcalorimetry, then biothermokinetics parameters were abstracted, the hemagglutination utility of samples were calculated and analyzed by principal component analysis (PCA) and cluster analysis (CA), meanwhile the results were authenticated by micro-plate agglutination. It showed that the hemagglutination was an exothermic reaction, the reaction rate constant (k: 0.039-73.6 min(-1)), maximum reaction power (Pmax: -1 140.2 - 988.2 microW) and reaction enthalpy (Hi: -529.9 - 717.9 microJ) had good linear correlation with BLG extraction concentration (0.2-1.0 g mL(-1), r > 0.97), and PCA showed Pmax (531-1 335 microW) and Ht (585.2-989.2 microJ) could represent the hemagglutination activity diversity of BLG samples, just confirming with the results of micro-plate agglutination (the agglutination dilution was 3-11 respectively). According to the hemagglutination utility, the BLG samples from Good Agriculture Practice (GAP) regions, main producing area and general regions could be clustered correctly; meanwhile, the biothermokinetics curves with perfect distinctive fingerprint and specificity could give out more information for the quality control and evaluation for BLG. In conclusion, the microcalorimetry method established for detecting the hemagglutination activity of BLG samples on rabbit erythrocyte is sensitive and reliable, and could be adopted as an effective technique in detection aggulatination precisely, quantitatively and consecutively; and provide a novel approach for examining and evaluating quality for Chinese herbal medicine with aggulatinative activity such as BLG.

[Differences between cold and hot natures of processed Radix ginseng rubra and Panax quinquefolius L. based upon mice temperature tropism].

OBJECTIVE: To establish an objective method to estimate the disparity between the cold and hot natures on the basis of an intrinsic correlation between temperature tropism of mice and the cold and hot natures of Chinese medicines. METHODS: Male KM mice were randomly divided into 7 groups of 6 each, namely the normal group (NM), the weak model group (WM), the strong model group (SM), the weak model plus Radix ginseng rubra group (WM + RG), the weak model plus Panax quinquefolius L. group (WM + PQ), the strong model plus Radix ginseng rubra group (SM + RG) and the strong model plus Panax quinquefolius L. group (SM +PQ). The specific herbal drugs were administered intragastricly. To induce the weak model, mice were fed with a limited supply of feed and forced to swim in cold water until almost drowning while the strong model induced by feeding a high-protein diet with an unlimited feed access. The doses of Radix ginseng rubra and Panax quinquefolius L. were 35 mg/g of body weight per day (counted by the quantity of crude material) and lasting for seven days. The NM and model groups without dosing were intragastricly administered with physiological saline of the same volume to the dosing groups. The percentage of the remaining time of mouse on a high temperature (40 degrees C) pad to the total monitoring time was recorded by a self-designed intelligent animal behavior monitoring system. Meanwhile, the drinking volume of mice in each group was measured. Immediately after experiment, the activities of Na(+)K(+)-ATPase and superoxide dismutase (SOD) in liver tissue were measured by assay kits of phosphorus and xanthine oxidase methods respectively. RESULTS: The features of deficient and cold symptom, such as fatigue, stagnant weight growth, decreased water intake, cold limbs and tail etc, were observed in WM group. And the features of heat symptom, such as increased weight and water intake, hyperactivity etc, were observed in SM group. The percentage of time that the mouse remained on 40 degrees C pad of the WM group within the seven days experiment was significantly higher than that of the normal group (70.6% +/- 21.3% vs 52.1% +/- 6.5%, P < 0.05). While the value of SM group (45.7% +/- 4.6% ) was significantly lower than that of the normal group (P < 0.05); the value of WM + RG group and WM + PQ group were 65.6% +/- 7.8% and 75.3% +/- 13.0% respectively (both P < 0.05 compared with WM group); the values of SM + RG group and SM + PQ group were 36.1% +/- 15.5% and 55.5% +/- 7.7% respectively (both P < 0.05 compared with SM group). The activities of Na(+)K(+)-ATPase and SOD of WM mice treated with either Radix ginseng rubra or Panax quinquefolius L. were found to have a significant up-regulation (P < 0.05) as compared with untreated WM mice. But only the Panax quinquefolius L. showed an up-regulating effect upon Na(+)K(+) ATPase and SOD in SM mice. CONCLUSIONS: The external cold and hot natures of Radix ginseng rubra and Panax quinquefolius L. can be represented in an ethological way by the changes of animal's temperature tropism. And such a tropism may be internally regulated by body's energy metabolism.

Action of palmatine on Tetrahymena thermophila BF5 growth investigated by microcalorimetry.

Using a thermal activity monitor (TAM) air isothermal microcalorimeter with ampoule mode, the thermo-genic curves of the metabolism of Tetrahymena thermophila BF(5) growth at 28 degrees C were obtained and the action of palmatine on it was investigated. Meanwhile, the biomass change during the process of T. thermophila BF(5) growth coexisted with palmatine was studied by a haemacytometer. The results showed that a low concentration (50 microg/mL) of palmatine began to inhibit the growth of T. thermophila BF(5), and when the concentration of palmatine reached 600 microg/mL, T. thermophila BF(5) could not grow at all. The relationship between the growth rate constant (k) and the concentration c was almost linear with the correlation coefficient of 0.9957, showing the strong toxic action of palmatine on T. thermophila BF(5) growth. The biomass during T. thermophila BF(5) growth decreased obviously by the addition of palmatine at different concentrations. The investigation of biomass agreed well with the results obtained by means of microcalorimetry.

[Determination of colistin based on biothermal activity detection method].

A biothermal activity detection method has been established to determine the potency of colistin. The biothermal activity fingerprints of E. coli with colistin were determined. There was a good linear relationship (r = 0.993) between logarithm concentration of colistin (lgC) and lag rate of growing time (Deltat%) when the concentrations of colistin ranged from 17.0 to 41.6 u x mL(-1). The average recovery rate was 100.3% (n = 9). Using this method, there was no significant difference between results of colistin potency measurement and those using cup-plate method (P > 0.05). As a result, biothermal activity detection method is sensitive, accurate, rapid, convenient and feasible to determine the potency of colistin. This method can also be applied in real time and online to monitor the process of bacterial growth and could be complementary to the cup-plate method.

[COLD and HOT nature of Coptis & Evodia and their prescriptions investigated with diet restriction/cold-water swimming mice models].

To establish a new method to evaluate the COLD and HOT nature of Coptis & Evodia and their prescriptions Zuojinwan and Fanzuojinwan. Physical models of mice were established by diet restriction with cold-water swimming (weak model, WM) and fed with high protein animal feeds (strong model, SM). An instrument with cold and hot pads was used to investigate the variation of temperature tropism among SM and WM groups of mice affected by drugs. Meanwhile, the oxygen consumption and activity of adenosine triphosphatase (ATPase) were detected, in order to investigate the mechanism of energy metabolism which might be affected by these drugs. The results showed that the drug effects gradually changed in an order of "Coptis-->Zuojinwan--> Fanzuojinwan-->Evodia". In detail, Coptis increased the remaining rate (RR) of mice on hot pad, decreased oxygen consumption and ATPase activity (n=6, P < 0.01 or P < 0.05), while Evodia performed inversely; which indicated the COLD nature of Coptis and HOT nature of Evodia, and confirmed with their traditional definition in medicinal works. In conclusion, the methods applied in this work, can objectively and directly express the nature disparity between the two herbs and predict the tendency of changes of the nature of their combination, which brings a new approach in investigation of the nature theory of traditional Chinese medicine.

[Effects of different drying methods and conditions on anthraquinones and tannins in water extract from Radix et Rhiroma Rhei].

OBJECTIVE: To investigate effects of different drying methods on the content of anthraquinones and tannins in water extract from Radix et Rhiroma Rhei (DHY). METHOD: DHY was dried by freeze drying, vacuum drying, drying under normal press and spray drying respectively until its moisture has been 5%. The content of anthraquinones and tannins of samples by different drying methods was determined and compared with. RESULT: The content of total anthraquinones, free anthraquinones, conjugated anthraquinones and tannins of samples by different drying methods was some different. Samples with freeze drying were highest and samples with drying under normal press at 100 degrees C were low. CONCLUSION: Temperature is an important pole in drying process water extract from Radix et Rhiroma Rhei. In our study water extract from Radix et Rhiroma Rhei was stable under 60 degrees C on the whole and unstable when drying exceed in 90 degrees C.

[HPLC fingerprint analysis of cordyceps and mycelium of cultured cordy].

OBJECTIVE: To establish HPLC fingerprints of cordyceps and mycelium of cultured cordy for quality control. METHOD: The analysis was carried out on a Diamonsil C18 column eluted with phosphate buffer (pH 6.5) and methanol as mobile phase, the gradient elution program was used, and the detection wavelength was 260 nm; The similarity of fingerprints was evaluated, then, cluster analysis was studied. RESULT: The method of HPLC fingerprint analysis was validated and in keeping with the requirement. Ten common peaks were found in HPLC fingerprint. Cordyceps from different areas and mycelium of cultured cordy could be distinguished by the HPLC fingerprint. CONCLUSION: All results above exhibit that this method is practicable, reproducible, and reliable as a method for the quality control of cordyceps and mycelium of cultured cordy at present.