Diffusosides C and D, two new iridoid glucosides from Oldenlandia diffusa.

During our continual investigations on Oldenlandia diffusa (Willd.) Roxb., two new iridoid glucosides, diffusosides C (1) and D (2), were isolated and their structures were elucidated through cumulative analysis of NMR and HRESIMS spectroscopic data as well as computational studies. Both isolated compounds displayed no obvious neuroprotective activity on H2O2-induced injury PC12 cells at 50 µM in vitro.

San Wu Huangqin decoction regulates inflammation and immune dysfunction induced by influenza virus by regulating the NF-κB signaling pathway in H1N1-infected mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The San Wu Huangqin Decoction (SWHD), which is made from the dried root of Sophora flavescens Aiton (Kushen in Chinese), the dried root of Scutellaria baicalensis Georgi (Huangqin in Chinese), and the dried root tuber of Rehmannia glutinosa (Gaertn.) DC. (Dihuang in Chinese), is a traditional Chinese formula used to treat prolonged fever and inflammatory diseases in clinics and proven to inhibit influenza virus effectively in our previous study. AIM OF THE STUDY: This work was performed to study the regulation of SWHD on inflammation and immune dysfunction induced by the influenza virus and the underlying mechanism in the treatment of SWHD. METHODS: In this study, the influenza virus A/PR/8/34 (H1N1)-infected mouse model was used to investigate the regulation of SWHD on inflammation and immune dysfunction induced by H1N1. The pathological changes, the capacity of proliferation of T and B lymphocytes, the cytotoxicity of natural killer (NK) cells, and the levels of IL-6, TNF-α, IL-1ß, IL-4, and IFN-γ in the serum, bronchoalveolar lavage fluid (BALF), and lung were analyzed. The effects of type 1 T helper cell (Th1) and type 2 T helper cell (Th2) immune responses were discussed indirectly. In addition, the expression levels of p-p65, p65, IKKα/ß, p-IκBα, and IκBα in relation to the NF-κB pathway were measured using Western blot analysis, or immunohistochemical assay. RESULTS: SWHD decreased the pathological changes in lung tissues, promoted the proliferation of T and B lymphocytes, enhanced NK cell activity, and accelerated the phagocytic function of macrophages in H1N1-infected mice. At the same time, SWHD decreased the levels of IL-6, TNF-α, IL-1ß, IFN-γ, and increased the level of IL-4 in the serum, BALF, and lung of model mice. Moreover, the p-p65, p65, and IκBα protein expression levels were inhibited, whereas the p-IκBα protein expression levels were improved in the lungs of H1N1-infected mice. CONCLUSIONS: SWHD can inhibit the replication of the H1N1 virus and reduced the excessive inflammation and immune dysfunction induced by the H1N1 virus in the body. This work provides rich experimental basis for further anti-inflammation research of SWHD and sets the foundation for the development of a viral inflammation drug of traditional Chinese medicine.

San Wu Huangqin Decoction, a Chinese Herbal Formula, Inhibits Influenza a/PR/8/34 (H1N1) Virus Infection In Vitro and In Vivo.

The San Wu Huangqin Decoction (SWHD), a traditional Chinese medicine formula, is used to treat colds caused by exposure to wind-pathogen, hyperpyrexia, infectious diseases and cancer; moreover, it is used for detoxification. The individual herbs of SWHD, such as Sophora flavescens and Scutellaria baicalensis, exhibit a wide spectrum of antiviral, anti-inflammatory, antibacterial, anticancer and other properties. The Chinese compound formula of SWHD is composed of S. flavescens, S. baicalensis and Rehmannia glutinosa. However, the effect of SWHD on the influenza virus (IFV) and its mechanism remain unknown. The aim of this study was to evaluate, for the first time, whether SWHD could be used to treat influenza. Results showed that SWHD could effectively inhibit influenza A/PR/8/34 (H1N1) virus at different stages of viral replication (confirmed through antiviral effect assay, penetration assay, attachment assay and internalization assay) in vitro. It could reduce the infection of the virus in a dose- and time-dependent manner, as confirmed by observing the cell cytopathic effect and calculating the cell viability (p < 0.05). SWHD demonstrated better antiviral activity than oseltamivir in the evaluation of antiviral prophylaxis on influenza (p < 0.05). The antiviral activity of SWHD may be related to its regulation ability on the immune system. Western blot, real-time polymerase chain reaction and indirect immunofluorescence assay showed that the expression of the four target viral proteins of the IFV (namely, haemagglutinin (HA), neuraminidase (NA), nucleoprotein (NP) and matrix-2 (M2)) reduced significantly (p < 0.05). Moreover, SWHD (23.40 and 11.70 g/kg) significantly alleviated the clinical signs, reduced the mortality and increased the survival time of infected mice (p < 0.05). The lung index, virus titres, pathological changes in lung tissues and the expression of key proteins of the IFV in mice also decreased (p < 0.05). In conclusion, SWHD possessed anti-influenza activity. This work provided a new view of complementary therapy and drug discovery for clinical treatment.

Antipyretic Effect of Herba Ephedrae-Ramulus Cinnamomi Herb Pair on Yeast-Induced Pyrexia Rats: A Metabolomics Study.

OBJECTIVE: To investigate the antipyretic mechanism of Herba Ephedrae (Eph)-Ramulus Cinnamomi (RC) herb pair on yeast-induced pyrexia in rats. METHODS: Totally 30 qualified male SD rats were randomly assigned to the normal control (NC) group, the pyrexia model (model) group, the Eph, RC and Eph-RC treatment groups by a random digital table, 6 rats in each group. Each rat received a 20% aqueous suspension of yeast (10 mL/kg) except the NC group. The 3 treatment groups were administered 8.1, 5.4 and 13.5 g/kg Eph, RC and Eph-RC respectively at 5 and 12 h after yeast injection, the NC group and the model groups were administered equal volume of distilled water. Rectal temperatures were measured at 0, 6, 8, 10, 12, 15, 18, 24 and 30 h and urine was collected prior to yeast injection and at 6, 10, 18, 24, 30, and 36 h after yeast injection. Then urine metabolomic profiling by gas chromatography tandem mass spectrometry, coupled with multivariate statistical analysis and pattern recognition techniques were used to explore the antipyretic effects of Eph-RC. Partial least squares discriminate analysis was used to analyze the metabolomics dataset including classification and regression in metabolomics plot profiling. RESULTS: Compared with the NC group, rectal temperatures were significantly higher in the model group (P<0.01), while 3 treatment groups decreased significantly compared with the model group (P<0.05 or P<0.01). Rectal temperatures of Eph-RC-treated rats started to go down at 6 h, and markedly decreased at 8, 12, 15, 18 and 24 h (P<0.05 or P<0.01), while those of the Eph and RC groups had decreased firstly at 8 h and were markedly lower at 12 h (P<0.05 or P<0.01). Seventeen potential biomarkers related to pyrexia were confirmed and identified, including pyruvic acid, L-phenylalanine, L-tyrosine, phenylacetic acid, hippuric acid, succinic acid, citrate and so on. Eight potential alterations of metabolic pathways including phenylalanine metabolism, citrate cycle, tryptophan metabolism, biosynthesis of valine, leucine and isoleucine, were identified in relation to the antipyretic effects of Eph-RC using MetPA software. CONCLUSION: The antipyretic effect of Eph-RC herb pair on yeast-induced pyrexia in rats involved correction of perturbed amino acid, fatty acid, and carbohydrate metabolism according to the metabolic pathway analysis with MetPA.

Comparative pharmacokinetics of three phenylpropanoids in rat plasma after oral administration of Ramulus Cinnamomi and Ramulus Cinnamomi-Ephedrae Herba herb-couple extract.

OBJECTIVE: To investigate the pharmacokinetic characteristics of three phenylpropanoids (cinnamic acid, cinnamic alcohol and coumarin) in Ramulus Cinnamomi (GZ) and Ramulus Cinnamomi-Ephedrae Herba (MH) herb-couple (GZMH). METHODS: Twelve male Sprague-Dawley rats were randomly and equally divided into the GZ and GZMH herb-couple groups. Blood samples were collected at 0, 0.08, 0.25, 0.5, 0.75, 1.5, 3, 4, 6, 8, 12, 24, 36 and 48 h after oral administration. The three phenylpropanoids in rat plasma were quantified using an ultra-performance liquid-chromatography with tandem mass spectrometry (UPLC-MS/MS) method for pharmacokinetic study. RESULTS: In GZMH group, the area under the curve (AUC), mean retention time (MRT) of cinnamic acid and coumarin were increased significantly (P<0.01, respectively), and biological half-life (t1/2z) was obviously shorter (P<0.05) compared with the GZ group. There were no significant differences in the mean retention time from 0 to ∞ (MRT0-∞), the peak concentration (Cmax), the time to peak (Tmax) and t1/2z, except for AUC and MRT0-t (the mean retention time from 0 to t) of cinnamic alcohol in the GZMH group by comparison to the GZ group (P<0.01, respectively). The AUC, MRT (both P<0.01) and t1/2z (P<0.05) of coumarin were increased significantly, while Cmax, and Tmax were decreased slightly by comparison to the GZ group (P>0.05). CONCLUSIONS: There were statistically significant differences in some pharmacokinetic parameters of the three compounds between GZ and GZMH groups, which meant that MH could affect the absorption and elimination of the three compounds.

Antitumorpharmacological mechanism of the oral liquid of Poriacocos polysaccharide.

ETHNOPHARMACOLOGICAL RELEVANCE: The liquid oral formulation of Poria cocos polysaccharides is composed of polysaccharides of Lentinusedodes, Ganodermalucidum and Poria cocos(1:1:2), which are all fungi used in traditional Chinese medicine. Polysaccharides extracted from these fungi have been reported to exhibit an antitumor effect by modulating the immune system. AIM OF THE STUDY: The present study aimed to clarify the antitumor mechanism of an orally administered liquid containing Poriacocos and to further provide clinical guidance. MATERIALS AND METHODS: In this study, the effects of an orally administered liquid containing Poriacocos polysaccharides on the solid tumors formed from sarcoma 180 cells in mice were evaluated. The protein expression of Bcl-2, caspase-3, and caspase-9in the thymus, spleen and liver tissues in the mice was determined by Western blot analysis. In addition, hematoxylin-eosin（H&E）staining and immunohistochemistry were performed on thymus, spleen and liver tissue and the positive staining rate was calculated for the three protein expression. RESULTS: The liquid oral formulation of Poriacocos polysaccharides reduced Bcl-2 protein levels and increased caspase-3 and -9 protein levels in sarcoma 180 cells. CONCLUSION: The mechanism underlying the antitumor effects of the oral liquid formulation of Poriacocos polysaccharides involved inhibition of Bcl-2 expression and activation of caspase-9 expression in sarcoma 180 cells. Furthermore, the downstream caspase-3 promoter cascade was activated and cell apoptosis was activated in sarcoma 180 cells.

Comparative Study on the Antivirus Activity of Shuang-Huang-Lian Injectable Powder and Its Bioactive Compound Mixture against Human Adenovirus III In Vitro.

Shuang-Huang-Lian injectable powder (SHL)-a classical purified herbal preparation extracted from Scutellaria baicalensis, Lonicera japonica, and Forsythia suspense-has been used against human adenovirus III (HAdV3) for many years. The combination herb and its major bioactive compounds, including chlorogenic acid, baicalin, and forsythia glycosides A, are effective inhibitors of the virus. However, no comprehensive studies are available on the antiviral effects of SHL against HAdV3. Moreover, it remains unclear whether the mixture of chlorogenic acid, baicalin, and forsythia glycosides A (CBF) has enhanced antiviral activity compared with SHL. Therefore, a comparative study was performed to investigate the combination which is promising for further antiviral drug development. To evaluate their antivirus activity in parallel, the combination ratio and dose of CBF were controlled and consistent with SHL. First, the fingerprint and the ratio of CBF in SHL were determined by high performance liquid chromatography. Then, a plaque reduction assay, reverse transcription polymerase chain reaction (PCR), real-time polymerase chain reaction (qPCR), and enzyme-linked immunosorbent assay (ELISA) were used to explore its therapeutic effects on viral infection and replication, respectively. The results showed that SHL and CBF inhibited dose- and time-dependently HAdV3-induced plaque formation in A549 and HEp-2 cells. SHL was more effective than CBF when supplemented prior to and after viral inoculation. SHL prevented viral attachment, internalization, and replication at high concentration and decreased viral levels within and out of cells at non-toxic concentrations in both cell types. Moreover, the expression of tumor necrosis factor alpha (TNF)-α, interleukin (IL)-1ß, and IL-6 was lower and the expression of interferon (IFN)-γ was higher in both cell types treated with SHL than with CBF. In conclusion, SHL is much more effective and slightly less toxic than CBF.

Evaluation of antinociceptive and anti-inflammatory effects of aqueous extract of Armadillidium vulgare Latreille.

OBJECTIVE: To assess the antinociceptive and anti-inflammatory properties of the aqueous extract of Armadillidium vulgare (AV). METHODS: The antinociceptive effect of AV (400, 600 and 800 mg/kg) was investigated in mice using the acetic acid-induced writhing, formalin-induced nociceptive, and hot plate tests. Phlogogen-induced paw edema using carrageenan, dextran, or compound 48/80 as phlogogen was used as inflammatory models to evaluate AV's anti-inflammatory effect. Additionally, the bioactive substances glucosamine (GLcN) and taurine in AV were determined using high-performance liquid chromatography. RESULTS: Oral treatment of the mice with AV (600 and 800 mg/kg) significantly reduced the number of writhes in the acetic acid-induced writhing test (P<0.01) but not the hot plate test (P>0.05). All doses tested significantly inhibited paw-withdrawal during the second phase of the formalin-induced nociceptive model (P<0.01). AV demonstrated a strong anti-inflammatory effect in all those inflammatory models (P<0.05). CONCLUSIONS: AV has antinociceptive and anti-inflammatory effects, providing scientific evidence of the efficacy of its traditional use in pain treatment. Furthermore, GLcN and taurine contribute, at least in part, to the anti-inflammatory activity of AV.

Anticancer Activities of C18-, C19-, C20-, and Bis-Diterpenoid Alkaloids Derived from Genus Aconitum.

Cancer is one of the most common lethal diseases, and natural products have been extensively studied as anticancer agents considering their availability, low toxicity, and economic affordability. Plants belonging to the genus Aconitum have been widely used medically in many Asian countries since ancient times. These plants have been proven effective for treating several types of cancer, such as lung, stomach, and liver cancers. The main effective components of Aconitum plants are diterpenoid alkaloids-which are divided into C18-, C19-, C20-, and bis-diterpenoid alkaloids-are reportedly some of the most promising, naturally abundant compounds for treating cancer. This review focuses on the progress of diterpenoid alkaloids with different structures derived from Aconitum plants and some of their derivatives with potential anticancer activities. We hope that this work can serve as a reference for further developing Aconitum diterpenoid alkaloids as anticancer agents.

Comparative tissue distribution and excretion study of alkaloids from Herba Ephedrae-Radix Aconiti Lateralis extracts in rats.

Herba Ephedrae-Radix Aconiti Lateralis, composed of Ephedrae (Mahuang in Chinese) and Radix Aconiti Lateralis (Fuzi in Chinese), is a classical herbal combination proven to be effective in treating common cold, asthma, and rheumatoid arthritis. Alkaloids, bioactive components of the herbal extract, have been associated with many side effects. Nine alkaloids, including norephedrine, norpseudoephedrine, ephedrine, pseudoephedrine, methylephedrine, hypaconitine, benzoylaconine, benzoylmesaconine and benzoylhypaconine, were simultaneously quantified within 14.5min, by a validated ultra-performance liquid chromatography-tandem mass spectrometry method in various rat tissues, urine, and feces after oral administration of Mahuang-Fuzi and single-herb extracts. The results indicated that the alkaloids were widely distributed in the heart, liver, spleen, lung, kidney, and brain. Lower bioavailability and higher clearance of some alkaloids were observed for the herbal combination, but hypaconitine showed a longer residence time and lower clearance. Elimination kinetics demonstrated that ephedra and aconitum alkaloids were mainly excreted in urine and feces, respectively. The tissue distribution and excretion of ephedra and aconitum alkaloids are comprehensively reported for the first time for the Mahuang-Fuzi combination. Compared with single-herb extracts, lower extraction efficiencies of alkaloids in vitro were observed which may result in their lower intake. However, the combination showed a prolonged residence time and delayed elimination of aconitum alkaloids, which increases the risk of drug accumulation. The study demonstrated potential risks of intoxication with aconitum alkaloids, associated with the use of Fuzi in combination with Mahuang. Mahuang-Fuzi is a classical combination used in clinics, further investigation is needed.

Stereoselective metabolism of amygdalin-based study of detoxification of Semen Armeniacae Amarum in the Herba Ephedrae-Semen Armeniacae Amarum herb pair.

ETHNOPHARMACOLOGICAL RELEVANCE: The Mahuang-Xingren (MX) herb pair, the combination of Herba Ephedrae (Mahuang in Chinese) and Semen Armeniacae Amarum (Xingren in Chinese), is a core component of traditional Chinese medicine formulations used to treat asthma and bronchitis. Although Xingren is considered to be toxic, MX is widely used in the clinic and has few adverse effects. The mechanism underlying detoxification of Xingren by Mahuang in MX remains unknown and merits investigation. AIM OF THE STUDY: To determine the mechanism underlying detoxification of Xingren by Mahuang in MX. MATERIALS AND METHODS: Acute toxic effects were evaluated in mice after oral administration of Mahuang, Xingren, and MX aqueous extracts. Synergism, additivity, and antagonism were quantified by determining the CI (combination index) and DRI (dose-reduction index), which were calculated by the median effect method. High performance liquid chromatography analysis of bioactive compounds (ephedrine, pseudoephedrine and amygdalin) in aqueous extracts and data from previous pharmacokinetic studies in rats were combined to explore the potential mechanism of toxicity antagonism by the components of MX. Moreover, the cytotoxic effects of amygdalin and amygdalin activated by ß-glucosidase (including different proportions of l-amygdalin and d-amygdalin) were also investigated. RESULTS: Mahuang prevented and antagonized the acute toxicity of Xingren and allowed escalation of the Xingren dose. Pearson correlation analysis indicated that the proportion of d-amygdalin was closely correlated with the antagonism of Xingren toxicity. The antagonism of its acute toxicity was primarily attributed to stereoselective metabolism of amygdalin. Interestingly, the process was facilitated by Mahuang, which led to reduced levels of the d-prunasin in vivo and thus reduced toxicity. Furthermore, the mechanism was also evaluated by testing the cytotoxicity of amygdalin. Metabolism of d-amygdalin was a major cause of cytotoxicity and no stereoselective metabolism occurred in culture medium. CONCLUSIONS: A comprehensive study of Xingren detoxification in the context of the MX combination suggested that stereoselective metabolism of amygdalin facilitated by Mahuang may be the crucial mechanism underlying detoxification of Xingren in the MX combination. Therefore, Mahuang acts to enhance and control the effects of Xingren in the MX combination. These results illustrate the rationale behind the combination of Mahuang and Xingren.

Antipyretic and anti-asthmatic activities of traditional Chinese herb-pairs, Ephedra and Gypsum.

OBJECTIVE: Mahuang-Shigao herb-pair is a famous formula composed of Ephedra and Gypsum. The herb-pair is frequently used for treating cold symptoms and bronchial asthma in the clinical practice of Chinese medicine (CM). In the present study, we evaluated evidence for the benefit of combined use of Ephedra and Gypsum by analyzing the antipyretic and anti-asthmatic activities of Ephedra-Gypsum. METHODS: The antipyretic effects of Ephedra-Gypsum were evaluated in yeast-induced hyperthermia test. Thirty male Wistar rats were randomly divided into 5 groups, including control group, standard aspirin group, and 3 Ephedra- Gypsum groups of different doses (6, 12, 24 g/kg). Ephedra-Gypsum extract and asprin were administered orally 6 h after the injection of yeast solution and body temperature was measured every 1 h for 8 h. The antiasthmatic effects of Ephedra-Gypsum were evaluated using an ovalbumin (OVA)-induced asthmatic rat model. Thirty-six male SD rats were randomly divided into 6 groups. Rats were alternately sensitized and OVA+Al(OH) challenged by exposure to mists of ovalbumin. Ephedra-Gypsum extracts (6, 12, 24 g/kg) or dexamethasone were administered 45 min prior to the allergen challenge for 8 days. Latent period and the weight of wet to dry ratio of lung were determined. In addition, the eosinophils in blood and white blood cell (WBC) were counted by an YZ-Hemavet Analyzer. RESULTS: The Ephedra-Gypsum extracts at test dose (6, 12, 24 g/kg) significantly and dose-dependently attenuated yeast-induced fever in rats. The Ephedra-Gypsum extracts also prolonged the latent period, reduced OVA-induced increases in eosinophils and WBC, and decreased the wet and dry weight ratio of the lungs in the anti-asthmatic test. CONCLUSIONS: These findings indicate that the Ephedra-Gypsum extract has antipyretic and anti-asthmatic properties. Hence, the results support additional scientific evidence in prescriptions.

[Changes in plasma pharmacokinetics and urinary excretion characteristics before and after combined administration of Ephedrae Herba-Gypsum Fibrosum].

In this study, UPLC-MS/MS was adopted to determine the contents of five ephedrine alkaloids (Norephedrine, Norpseudoephedrine, Ephedrine, Pseudoephedrine, Methylephedrine) in plasma and urine in rats after the combined administration of Ephedrae Herba-Gypsum Fibrosum and calculate relevant pharmacokinetic parameters, in order to discuss the effect of the combined administration of Ephedrae Herba-Gypsum Fibrosum on plasma pharmacokinetics and urinary excretion characteristics. According to the results, after being combined with Gypsum, the five ephedrine alkaloids showed similar pharmacokinetic changes, such as shortened t(max), accelerated absorption rate, but reduced AUC(0-t) and V(z)/F, which may be related to the increase in urine excretion. Besides, Gypsum was added to enhance C(max) of Pseudoephedrine and prolong MRT(0-t) of Methylephedrine, so as to enhance the anti-asthmatic effect of Ephedrae Herba and resist the toxic effect of Norephedrine and Ephedrine. This study proved the scientific compatibility of Ephedrae Herba-Gypsum Fibrosum and provided a reference for studies on the prescription compatibility regularity and relevant practices.

Evaluation of the anaphylactoid potential of Andrographis paniculata extracts using the popliteal lymph node assay and P815 cell degranulation in vitro.

BACKGROUND: The anaphylactoid reactions induced by andrographis injection have repeatedly been reported. The aim of our study was to evaluate the immuno-sensitizing potential of extracts from Andrographis paniculata Nees and to screen for the constituent that is responsible for inducing the anaphylactoid reaction. METHODS: In the direct popliteal lymph node assay (D-PLNA), female BALB/c mice were randomly divided into several groups with ten mice per group according to the experiment design, the right hind footpads of mice received a single subcutaneous injection of Andrographis paniculata (50 µl), and the left hind footpads received the same volume of vehicle. Seven days later, the mice were sacrificed by cervical dislocation, and the popliteal lymph nodes from both the left and right sides were removed. The weight (WI) and cellularity indices (CI) of the popliteal lymph nodes (PLNs) were then calculated, and the pathological changes of the PLNs were measured. In addition, P815 mast cells were collected for the in vitro cell degranulation experiment. The level of histamine, the percentage of cell degranulation and the ratio of ammonia glycosidase released were measured to further evaluate the potential allergenicity. RESULTS: Alcohol extract (AEE), ethyl acetate extract (EAE) and n-butanol extract (NBE) significantly increased the weight (WI > 2) and cell number (CI > 5) of PLNs (P < 0.05, P < 0.01). Additionally, all the three monomers of andrographis, namely NAD, AND, and DDA, significantly increased the weight (WI > 2) and cell number (CI > 5) of the PLNs (P < 0.05, P < 0.01). In the cell model, all of the different extract fractions (AEE, EAE and NBE) and the three monomers of andrographis markedly elevated the level of histamine, the percentage of cell degranulation and the ratio of ammonia glycosidase released. CONCLUSION: The diterpene lactone compounds of Andrographis paniculata Nees (total lactones of andrographolide) may have a potential sensitizing capacity in andrographis injection.

Concurrent quantification and comparative pharmacokinetic analysis of bioactive compounds in the Herba Ephedrae-Semen Armeniacae Amarum herb pair.

The Mahuang-Xingren herb-pair (MX), the combination of Herba Ephedrae (Mahuang in Chinese) and Semen Armeniacae Amarum (Xingren in Chinese), is a classical combination used in traditional Chinese Medicine to treat asthma and bronchitis. A simple and reliable ultra-performance liquid chromatography-tandem mass spectrometry method was developed to simultaneously quantify and compare the pharmacokinetics of 5 ephedra alkaloids and epimers of amygdalin and prunasin in rat plasma after oral administration of Mahuang, Xingren, and MX aqueous extracts. Samples were pretreated by a single-step protein precipitation with acetonitrile, and diphenhydramine hydrochloride and puerarin were used as internal standards. Pharmacokinetic parameters were investigated using DAS 3.2.2 (Mathematical Pharmacology Professional Committee of China, Shanghai, China). The validated method demonstrated adequate sensitivity, selectivity, and process efficiency for the bioanalysis of 8 compounds, including 3 pairs of epimers. MX administration improved the bioavailability of amygdalin and prunasin. Furthermore, MX facilitated intake of lower doses of ephedra alkaloids and increased elimination rates in comparison with Mahuang alone. These results illustrate the rationale behind the preferred use of the combination of Mahuang and Xingren. To our knowledge, this is the first report of stereo-selective metabolism of amygdalin. Further, the metabolic mechanism underlying this phenomenon merits future research attention.

Neuroprotective effect of gui zhi (ramulus cinnamomi) on ma huang- (herb ephedra-) induced toxicity in rats treated with a ma huang-gui zhi herb pair.

Herb Ephedra (Ma Huang in Chinese) and Ramulus Cinnamomi (Gui Zhi in Chinese) are traditional Chinese herbs, often used together to treat asthma, nose and lung congestion, and fever with anhidrosis. Due to the adverse effects of ephedrine, clinical use of Ma Huang is restricted. However, Gui Zhi extract has been reported to decrease spontaneous activity in rats and exert anti-inflammatory and neuroprotective effects. The present study explored the possible inhibitory effect of Gui Zhi on Ma Huang-induced neurotoxicity in rats when the two herbs were used in combination. All Ma Huang and Ma Huang-Gui Zhi herb pair extracts were prepared using methods of traditional Chinese medicine and were normalized based on the ephedrine content. Two-month-old male Sprague-Dawley rats (n = 6 rats/group) were administered Ma Huang or the Ma Huang-Gui Zhi herb pair extracts for 7 days (ephedrine = 48 mg/kg), and locomotor activity was measured. After 7 days, oxidative damage in the prefrontal cortex was measured. Gui Zhi decreased hyperactivity and sensitization produced by repeated Ma Huang administration and attenuated oxidative stress induced by Ma Huang. The results of this study demonstrate the neuroprotective potential of Gui Zhi in Ma Huang-induced hyperactivity and oxidative damage in the prefrontal cortex of rats when used in combination.

Simultaneous quantification and pharmacokinetics of alkaloids in Herba Ephedrae-Radix Aconiti Lateralis extracts.

The combination of Herba Ephedrae (Mahuang in Chinese) and Radix Aconiti Lateralis (Fuzi in Chinese) is a classical preparation in traditional Chinese medicine and used for treating colds and rheumatic arthralgia. However, herbal medicines containing ephedrines and Aconitum alkaloids are strictly regulated because of the potential for adverse effects on the cardiovascular system and the central nervous system. We aimed to investigate the pharmacokinetics of 11 alkaloids in the Mahuang-Fuzi combination and single-herb extracts after oral administration in rats. The alkaloids were norephedrine, norpseudoephedrine, ephedrine, pseudoephedrine, methylephedrine, aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine and benzoylhypaconine. Simultaneous determination of the alkaloids, including two pairs of diastereomers, was achieved in 14.5 min by a simple, rapid and sensitive ultra-performance liquid chromatography-tandem mass spectrometry method. The separation was performed on a Zorbax SB-Aq column (100 mm × 2.1 mm, 3.5 µm) at a flow rate of 0.3 mL/min using acetonitrile-0.1% formic acid aqueous solution as the mobile phase. The validated method demonstrated adequate sensitivity, selectivity and process efficiency for the quantitative analysis of complex herbal components. Compared with single-herb extracts, alkaloids in plasma (except methylephedrine, benzoylmesaconine and benzoylhypaconine) showed slower elimination (the mean residence time or half-life was longer), although the maximum plasma concentration and area under the plasma concentration curve values decreased. Accumulation may occur with continuous drug intake. These results suggest that drug monitoring may be essential for the safe use of the Mahuang-Fuzi combination.

Pharmacokinetic comparisons of five ephedrine alkaloids following oral administration of four different Mahuang-Guizhi herb-pair aqueous extracts ratios in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Herba Ephedra (Mahuang in Chinese), is derived from dried Ephedra sinica Stapf stems and has been widely used to treat the common cold, coughs, asthma, and edema for thousands of years. The Mahuang-Guizhi herb-pair is a famous formula composed of Mahuang and Ramulus Cinnamomi (Guizhi in Chinese, the dried twig of Cinnamomum cassia Presl.), used to improve pharmacological effects and reduce toxicity. In order to investigate the influence of Mahuang-Guizhi herb-pair ratios on bioavailability, the plasma pharmacokinetics profiles of five ephedrine alkaloids were compared following oral administration of four different ratios to rats. MATERIALS AND METHODS: Sprague-Dawley rats were randomly assigned to four groups and orally administered Mahuang-Guizhi (ratios 3:0; 3:1; 3:2; 3:4, w/w). Assays for five ephedrine alkaloids (ephedrine, pseudoephedrine, methylephedrine, norephedrine, and norpseudoephedrine) were developed and validated using ultra-high-performance liquid chromatography tandem mass spectrometry coupled with liquid-liquid extraction. RESULTS: Key pharmacokinetic parameters of the five ephedrine alkaloids (maximal plasma concentration, mean residence time, and half-life) were significantly different (p<0.05) after oral administration of Mahuang-Guizhi herb-pair ratios, as compared to those of Mahuang. CONCLUSION: Ephedrine alkaloid pharmacokinetic differences in rat plasma could help explain previous findings of pharmacological and toxicity differences between Mahuang and Mahuang-Guizhi herb-pair preparations. These results could facilitate future studies to increase the efficacy and decrease the toxicity of Mahuang and Guizhi.

[Pharmacokinetic and Tissue Distribution Study of Ephedrae Herba and Herbal Pair of Ephedrae Herba-Atractylodis Macrocephalae Rhizoma in Rats].

OBJECTIVE: Five kinds of Ephedra alkaloids (NME, NMP, E, PE and ME) in Ephedrae Herba extracts and Ephedrae Herba-Atractylodis Macrocephalae Rhizoma herbal pair extracts of plasma pharmacokinetic and tissue distribution study in rats were carried out, to discuss the changes of Ephedrae Herba compatibility with Atractylodis Macrocephalae Rhizoma before and after METHODS: HPLC- MS method was used and the condition was as flollows: ZORBAX SB-C18 column (100 mm x 2.1 mm, 3.5 µm), column temperature of 35 °C, mobile phase of ACE-0.1% formic solution in gradient elution mode, flow rate at 0.4 mL/min; MRM positive ion detection mode. RESULTS: The distribution trends of Ephedra alkaloids were changing in plasma and tissues of rats after Ephedrae Herba compati- bility with Atractylodis Macrocephalae Rhizoma. CONCLUSION: Ephedrae Herba compatibility with Atractylodis Macrocephalae Rhizoma may increase drug efficacy and reduce the toxicity of Ephedra alkaloids at the same time.