Indole alkaloids from the aerial parts of Kopsia fruticosa and their cytotoxic, antimicrobial and antifungal activities.

A chemical investigation on the 80% EtOH extract of the aerial parts of Kopsia fruticosa led to five new indole alkaloids, kopsifolines G-K (1-5), and one known alkaloid, kopsifoline A (6). Structural elucidation of all the compounds were performed by spectral methods such as 1D and 2D (1H-1H COSY, HMQC, and HMBC) NMR spectroscopy, in addition to high resolution mass spectrometry. The isolated components were evaluated in vitro for cytotoxic activities against seven tumor cell lines, antimicrobial activities against two Gram-positive bacteria and five Gram-negative bacteria, and antifungal activities against five pathogens. As a result, alkaloids 3-5 exhibited some cytotoxicity against all of seven tested tumor cell lines (HS-1, HS-4, SCL-1, A431, BGC-823, MCF-7, and W480) with IC50 values of 11.8-13.8, 10.3-12.5, and 7.3-9.5 µM, respectively. Alkaloids 3-5 also possessed significant antimicrobial and antifungal activities which was reported for the first time for the alkaloids isolated from Kopsia genus.

Antiaging effect of Cordyceps sinensis extract.

This experiment studied the effect of Cordyceps sinensis extract (CSE) on mice aged by d-galactose and castrated rats to analyse its antiaging effect. Water maze and step-down type avoidance tests were used to examine the effect of CSE on learning and memory. CSE shortened escape latency, prolonged step-down latency and decreased the number of errors in mice aged by d-galactose. The effect of CSE on the sexual function of castrated rats was evaluated by measuring the penis erection latency, mount latency and ejaculation latency. CSE appeared to shorten penis erection latency and mount latency in castrated rats. The study also measured the effect of CSE on the activity of age-related enzymes. The results showed that CSE improved the activity of superoxide dismutase, glutathione peroxidase and catalase and lowered the level of lipid peroxidation and monoamine oxidase activity in the aged mice. The study demonstrated that CSE can improve the brain function and antioxidative enzyme activity in mice with d-galactose-induced senescence and promote sexual function in castrated rats. All of these findings suggest that CSE has an antiaging effect.

Establishment of a cell-based assay to screen regulators of the hypoxia-inducible factor-1-dependent vascular endothelial growth factor promoter.

In this study, we established a drug screening system based on transcriptional regulation of vascular endothelial growth factor (VEGF) under hypoxia-inducible factor-1alpha control. We cloned the neomycin-resistance gene into the plasmid GL (pGL)3-promoter vector to generate the pGL3-promoter-neo vector. The 3 copies of the 47-bp fragment that contained the hypoxia response element of VEGF were synthesized and inserted in front of the minimal promoter of the pGL3-promoter-neo vector to generate p3HRE-luc-neo. The recombinant reporter gene vectors were transfected into EAhy926 cells, and stable cell lines were obtained. The positive cell line was selected for its ability to express luciferase in response to hypoxia. We demonstrated that CoCl(2) significantly enhances luciferase activity in a concentration-dependent fashion. We then optimized the cell density and incubation time under hypoxia which were used to screen. The assay exhibited a low background and was an ideal model for high-throughput screening for human VEGF regulators.

Echinacoside prevents the striatal extracellular levels of monoamine neurotransmitters from diminution in 6-hydroxydopamine lesion rats.

We investigated the effects of echinacoside, a phenylethanoid glycoside isolated and purified from the stems of Cistanche salsa, a Chinese herbal medicine, on the striatal extracellular levels of dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in 6-hydroxydopamine (6-OHDA) lesion rats. Seven days after 6-OHDA was injected into the right striatum of rats, the striatal extracellular levels of DA, DOPAC and HVA fell significantly (P<0.01 vs. vehicle), as demonstrated by the method of cerebral microdialysis and high performance liquid chromatography with electrochemical detection. However, simultaneous treatment with echinacoside (7.0, 3.5mg/kg) attenuated the diminution of them (P<0.01 vs. model). The results implied that echinacoside could protect the striatal dopaminergic neurons from injury induced by 6-OHDA and may be useful in the prevention and treatment of Parkinson's disease (PD).

Unique spirocyclopiperazinium salt III: further investigation of monospirocyclopiperazinium (MSPZ) salts as potential analgesics.

Two novel classes of monospirocyclopiperazinium salts were designed, synthesized, and evaluated for their in vivo analgesic activities. Some interesting structure-activity relationships are revealed: (1) Spirocyclopiperazinium moiety is favorable to improve the analgesic activity; (2) The size and conformation of spirocyclopiperazinium moiety significantly affects the analgesic activity; (3) Phenylethyl group of 3d is a crucial pharmacophore. Among the compounds synthesized, 3d exhibited the most potent activity with low toxicity. Further antinociceptive mechanism studies of 3d showed that these compounds will be a new kind of analgesics.

[Experimental study on primary pharmacodynamics of Niuhuang Qingwei wan].

OBJECTIVE: To study the primary effects of Niuhuang Qingwei wan on the gastrointestinal function in aninmal for justifying its efficacies in clinic. METHOD: Mice were twice administered with Niuhuang Qingwei wan (0.83, 1.67, 3.33 g x kg(-1), ig) and rats were twice administered with Niuhuang Qingwei wan (0.59, 1.18, 2.36 g x kg(-1), ig). The effects on the stomach function were evaluated by the gastric emptying test in mice and the gastric analysis in rats. The effect on the intestinal function were evaluated by the propulsive motility of the total gastrointestinal tract test in mice by recording the time and frequency of excreting carbo medicinalis. Its analgesia was explored by using the abdominal constriction test induced by acetic acid. RESULT: Niuhuang Qingwei wan decreased the activity and secretion of pepsin in a dose-dependent manner (P < 0.01, P < 0.05), the gastric juice volume at middle and high doses (P <0.01, P <0.05), and the gastric acid volume at high dose (P <0.05); However, it had no significant effects on the gastric emptying in normal mice and the acidity in gastric juice. It shortened the excreting time of feces and increased the frequency of defecation (P < 0.01, P < 0.05). It also inhibited abdominal constriction responses at high dose, and the inhibition rate was 40.0% (P <0.01). CONCLUSION: Niuhuang Qingwei wan can promote gastrointestinal motility, decrease gastric acid volume and activity of pepsin and show certain analgesia effect. Those findings are consistent with its treating stomach heat in clinic.

Antinociceptive effects of the novel spirocyclopiperazinium salt compound LXM-10 in mice.

The compound LXM-10 (2,4-dimethyl-9-beta-phenylethyl-3-oxo-6, 9-diazaspiro [5.5]undecane chloride) is a new spirocyclopiperazinium salt compound. This is the first article to evaluate its antinociceptive effect in the abdominal constriction test induced by acetic acid and the hot-plate test. In the abdominal constriction test, LXM-10 had a significant dose-response effect, and the maximal inhibition ratio was 79.2%. In the hot-plate test, LXM-10 had significant dose-response and time-response effects. The antinociceptive effect began at 1.0 h, peaked at 2.0 h, and persisted 3.0 h after s.c. administration. The hot-plate latency was increased by 126.8% at the dose of 12.0 mg/kg. The antinociceptive effect of LXM-10 was blocked by mecamylamine (a central and peripheral neuronal nicotinic acetylcholine receptor antagonist, 0.25, 0.5, 1.0 mg/kg, i.p.), hexamethonium (a peripheral neuronal nicotinic acetylcholine receptor antagonist, 0.2, 1.0, 5.0 mg/kg, i.p.), atropine (a central and peripheral muscarinic acetylcholine receptor antagonist, 0.2, 1.0, 5.0 mg/kg, i.p.), and atropine methylnitrate (a peripheral muscarinic acetylcholine receptor antagonist, 0.2, 1.0, 5.0 mg/kg, i.p.) in a dose-dependent fashion. In contrast, the effect was not blocked by naloxone (a non-selective opioid receptor antagonist, 2.0 mg/kg, i.p.) or yohimbine (a alpha(2)-adrenergic receptor antagonist, 1.0, 2.5, 5.0 mg/kg, i.p.) in the hot-plate test. Therefore, the antinociceptive effects of LXM-10 involve the peripheral neuronal nicotinic and muscarinic acetylcholine receptors; they are not related to opioid receptors or alpha(2)-adrenergic receptors. LXM-10 did not affect motor coordination, spontaneous activity, or body temperature. These findings with LXM-10 suggest that spirocyclopiperazinium derivatives could provide insight on new analgesics.

[Protective effect of hydroxysafflor yellow A on experimental cerebral ischemia in rats].

AIM: To investigate the protective effect of hydroxysafflor yellow A (HSYA), a soluble element extracted from Carthamus tinctorius L., on focal cerebral ischemia in rats. METHODS: Focal cerebral ischemia in male Wistar-Kyoto (WKY) rats were induced by permanent middle cerebral artery occlusion (MCAO). Three doses of 1.5, 3.0 and 6.0 mg x kg(-1) of HSYA were administrated to three groups of rats, separately, via sublingular vein injection 30 min after the onset of ischemia. 24 h after ischemia in rats, neurological deficit scores were evaluated and the infarction area of brain was assessed by quantitative image analysis. The in vitro neuroprotective effect of HSYA was tested in cultured fetal cortical neurons exposed to glutamate and sodium cyanide (NaCN). RESULTS: HSYA at doses of 3.0 and 6.0 mg x kg(-1) exerted significant neuroprotective effects on rats with focal cerebral ischemic injury as expressed by neurological deficit scores and reduced the infarct area as compared with saline group, and the potency of HSYA at dose of 6.0 mg x kg(-1) was similar to that of 0.2 mg x kg(-1) of nimodipine. In vitro studies, HSYA significantly inhibited neurons damage induced by exposure to glutamate and NaCN in cultured fetal cortical cells. CONCLUSION: HSYA has potential neuroprotective action against focal cerebral ischemia in rats and cultured rat fetal cortical neurons as well.

[An experimental study on anti-aging action of Cordyceps extract].

OBJECTIVE: To investigate anti-aging effect and mechanism of Cordyceps extract(CSE) on aged mice induced by D-galactose. METHOD: The aged mice were induced by D-galactose. Meanwhile, they were treated with three doses of CSE. Then the ability of learning and memory, the activity of antioxidase in the different tissue, the contents of MDA of brain and liver were measured after 6 weeks. RESULT: CSE could significantly increase the ability of learning and memory, improve the activity of SOD of red blood cells, brain and liver, the activity of Na(+) -K(+) -ATPE of brain, the activity of CAT and GSH-Px of blood, and remarkably decrease the activity of MAO of brain and the contents of MDA of brain and liver. CONCLUSION: CSE has good anti-aging effects on the aged mice, which is probably due to effects of improving antioxidation and removing free radicals.

Protective effect of verbascoside on 1-methyl-4-phenylpyridinium ion-induced neurotoxicity in PC12 cells.

The neuroprotective effects of verbascoside, one of phenylpropanoid glucoside isolated from the Chinese herbal medicine Buddleja officinalis Maxim, on 1-methyl-4-phenylpyridinium ion (MPP(+)) induced apoptosis and oxidative stress in PC12 neuronal cells were investigated. Treatment of PC12 cells with MPP(+) for 48 h induced apoptotic death as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry, the activation of caspase-3 measured by the caspase-3 activity assay kit, the reduction in mitochondrial membrane potential with laser scanning confocal microscopy and the increase in the extracellular hydrogen peroxide level. Simultaneous treatment with verbascoside markedly attenuated MPP(+)-induced apoptotic death, increased extracellular hydrogen peroxide level, the activation of caspase-3 and the collapse of mitochondrial membrane potential. These results strongly indicate that verbascoside may provide a useful therapeutic strategy for the treatment of oxidative stress-induced neurodegenerative disease such as Parkinson's disease.

[Variation of endogeneous ET, CGRP and NO in myocardial ischemia in rats and the regulatory effect of xinshuping].

OBJECTIVE: To observe the variation of ET and CGRP contents in ischemic heart, and NO level in serum of myocardial damaged rats, and their regulation when with the protection of Xinshuping, a traditional Chinese medicine compound. METHOD: The models of myocardial ischemia were prepared by subcutaneous injection of isoproterenol or by ligation of coronary artery. RESULT: ET content in myocardium was significantly increased (P < 0.01), and CGRP content as well as NO level in serum was not changed obviously in the model induced by isoproterenal. However, NO level in serum of rats treated with Xinshuping (ig bid x 2.5 d) was markedly raised (P < 0.01), neither ET not CGRP contents were affected by it. LDH and CK levels in serum of rats were evidently lowered by Xinshuping treatment. S-T segment's elevation of ECG was significantly inhibited and myocardial infarction size was reduced markedly by Xinshuping treatment in rats subjected to coronary artery ligature. CONCLUSION: ET, CGRP or NO is involved in myocardial infarction caused by isoproterenol. The ischemic damage or dysfunction in different models is obviously protected by Xinshuping. The promotion of NO release from vascular endothelium is probably related with this protective effect.

[Mechanical research on effects of yishenqing on membranous nephropathy].

OBJECTIVE: To investigate the therapeutic effect and mechanism of Yishenqing on rabbit membranous nephropathy. METHOD: The rabbit membranous nephropathy model was induced by cationic bovine serum albumin, and the immunity of mice together with urine volume and haemorheological property of rats were also estimated. RESULT: Yishenqing could significantly reduce the urinary protein content, preserve the renal function and pathologically restore the glomeruli. Moreover, its effects also include immunity enhancement, diuretic property and blood stasis amelioration. CONCLUSION: Yishenqing has good effects on the rabbit membranous nephropathy, which is probably due to the effects of diuretic property, immunity enhancement and blood stasis amelioration.