Kornkanok Ingkaninan

High-performance liquid chromatography with on-line coupled UV, mass spectrometric and biochemical detection for identification of acetylcholinesterase inhibitors from natural products.

A high-performance liquid chromatography (HPLC) method with on-line coupled ultraviolet (UV), mass spectrometry (MS) and biochemical detection for acetylcholinesterase (AChE) inhibitory activity has been developed. By combining the separation power of HPLC, the high selectivity of biochemical detection, and the ability to provide molecular mass and structural information of MS, AChE inhibitors can be rapidly identified. The biochemical detection was based on a colorimetric method using Ellmans reagent. The detection limit of galanthamine, an AChE inhibitor, in the HPLC-biochemical detection is 0.3 nmol. The three detector lines used, i.e., UV, MS and Vis for the biochemical detection were recorded simultaneously and the delay times of the peaks obtained were found to be consistent. This on-line post-column detection technique can be used for the identification of AChE inhibitors in plant extracts and other complex mixtures such as combinatorial libraries.

Cognitive enhancement and neuroprotective effects of Bacopa monnieri in Alzheimer's disease model

ETHNOPHARMACOLOGICAL RELEVANCE Bacopa monnieri (L.) Wettst., a plant belonging to the family Scrophulariaceae, has been used in the traditional system of Ayurvedic medicine to improve intelligence and memory for a long time. Therefore, the potential of this plant to protect against Alzheimers disease has been raised but less supported document is available. AIM OF THE STUDY To determine the effect of alcoholic extract of Bacopa monnieri on cognitive function and neurodegeneration in animal model of Alzheimers disease induced by ethylcholine aziridinium ion (AF64A). MATERIALS AND METHODS Male Wistar rats were orally given the alcoholic extract of Bacopa monnieri at doses of 20, 40 and 80 mg/kg BW via feeding needle for a period of 2 weeks before and 1 week after the intracerebroventricular administration of AF64A bilaterally. Rats were tested for spatial memory using Morris water maze test and the density of neurons and cholinergic neurons was determined using histological techniques 7 days after AF64A administration. RESULTS Bacopa monnieri extract improved the escape latency time (p<.01) in Morris water maze test. Moreover, the reduction of neurons and cholinergic neuron densities were also mitigated. CONCLUSION These findings suggest that Bacopa monnieri is a potential cognitive enhancer and neuroprotectant against Alzheimers disease.

Neuroprotective effect of Bacopa monnieri on beta-amyloid-induced cell death in primary cortical culture.

AIM OF THE STUDY Bacopa monnieri (Brahmi) is extensively used in traditional Indian medicine as a nerve tonic and thought to improve memory. To examine the neuroprotective effects of Brahmi extract, we tested its protection against the beta-amyloid protein (25-35) and glutamate-induced neurotoxicity in primary cortical cultured neurons. MATERIALS AND METHODS Neuroprotective effects were determined by measuring neuronal cell viability following beta-amyloid and glutamate treatment with and without Brahmi extract. Mechanisms of neuroprotection were evaluated by monitoring cellular oxidative stress and acetylcholinesterase activity. RESULTS Our result demonstrated that Brahmi extract protected neurons from beta-amyloid-induced cell death, but not glutamate-induced excitotoxicity. This neuroprotection was possibly due to its ability to suppress cellular acetylcholinesterase activity but not the inhibition of glutamate-mediated toxicity. In addition, culture medium containing Brahmi extract appeared to promote cell survival compared to neuronal cells growing in regular culture medium. Further study showed that Brahmi-treated neurons expressed lower level of reactive oxygen species suggesting that Brahmi restrained intracellular oxidative stress which in turn prolonged the lifespan of the culture neurons. Brahmi extract also exhibited both reducing and lipid peroxidation inhibitory activities. CONCLUSIONS From this study, the mode of action of neuroprotective effects of Brahmi appeared to be the results of its antioxidant to suppress neuronal oxidative stress and the acetylcholinesterase inhibitory activities. Therefore, treating patients with Brahmi extract may be an alternative direction for ameliorating neurodegenerative disorders associated with the overwhelming oxidative stress as well as Alzheimers disease.

Hypoglycemic effect of bitter melon compared with metformin in newly diagnosed type 2 diabetes patients

ETHNOPHARMACOLOGICAL RELEVANCE Bitter melon (Momordica charantia L.) has been widely used as an traditional medicine treatment for diabetic patients in Asia. In vitro and animal studies suggested its hypoglycemic activity, but limited human studies are available to support its use. AIM OF STUDY This study was conducted to assess the efficacy and safety of three doses of bitter melon compared with metformin. MATERIALS AND METHODS This is a 4-week, multicenter, randomized, double-blind, active-control trial. Patients were randomized into 4 groups to receive bitter melon 500 mg/day, 1,000 mg/day, and 2,000 mg/day or metformin 1,000 mg/day. All patients were followed for 4 weeks. RESULTS There was a significant decline in fructosamine at week 4 of the metformin group (-16.8; 95% CI, -31.2, -2.4 μmol/L) and the bitter melon 2,000 mg/day group (-10.2; 95% CI, -19.1, -1.3 μmol/L). Bitter melon 500 and 1,000 mg/day did not significantly decrease fructosamine levels (-3.5; 95% CI -11.7, 4.6 and -10.3; 95% CI -22.7, 2.2 μmol/L, respectively). CONCLUSIONS Bitter melon had a modest hypoglycemic effect and significantly reduced fructosamine levels from baseline among patients with type 2 diabetes who received 2,000 mg/day. However, the hypoglycemic effect of bitter melon was less than metformin 1,000 mg/day.

Rapid screening and identification of antioxidants in aqueous extracts of Houttuynia cordata using LC-ESI-MS coupled with DPPH assay

Abstract Houttuynia cordata Thunb. has been used traditionally as immune stimulant and anticancer agent. An aqueous extract of H. cordata tea showed high radical scavenging activity determined by off-line DPPH assays. Then, it was screened for its antioxidant components via an on-line DPPH radical scavenging technique coupled with a liquid chromatography–electrospray ionization mass spectrometer (LC–ESI–MS). Based on their mass spectra and fragmentation patterns; the antioxidant compounds were identified as quinic acid derivative, caffeic acid derivatives, procyanidin B, neo-chlorogenic acid, catechin, chlorogenic acid, crypto-chlorogenic acid and quercetin hexoside. LC–MS/MS in multiple reactions monitoring (MRM) mode was used to quantify these antioxidant compounds. Chlorogenic acid was found to be a major component in H. cordata tea.

Acetylcholinesterase inhibitors from Stephania venosa tuber

Acetylcholinesterase (AChE) inhibitors have lately gained interest as potential drugs in the treatment of Alzheimers disease. Three AChE inhibitors were isolated from tubers of a Thai medicinal plant, Stephania venosa (Bl) Spreng. They were identified as quaternary protoberberine alkaloids, stepharanine, cyclanoline and N‐methyl stepholidine. They expressed inhibitory activity on AChE with IC50 values (concentration that caused 50% inhibition of activity) of 14.1K ± 0.81, 9.23 ± 3.47 and 31.30 ± 3.67 μM, respectively. The AChE inhibitory activity of these compounds was compared with those of the related compounds, palmatine, jatrorrhizine and berberine, as well as tertiary protoberberine alkaloids isolated from the same plant, stepholidine and corydalmine. The results suggest that the positive charge at the nitrogen of the tetrahydroisoquinoline portion, steric substitution at the nitrogen, planarity of the molecule or substitutions at C‐2, −3, −9, and −10 affect the AChE inhibitory activity of protoberberine alkaloids.

Vobasinyl‐iboga bisindole alkaloids, potent acetylcholinesterase inhibitors from Tabernaemontana divaricata root

The roots of the Thai medicinal plant, Tabernaemontana divaricata (L.) R. Br. ex Roem. & Schult., were investigated for their content of acetylcholinesterase inhibitors. Bioassay‐guided fractionation using the Ellman colorimetric method led to the isolation of two bisindole alkaloids, 19,20‐dihydrotabernamine and 19,20‐dihydroervahanine A. The compounds showed higher inhibitory activity on acetylcholinesterase in comparison with galanthamine, a well‐known acetylcholinesterase inhibitor. The inhibitory activity of 19,20‐dihydroervahanine A was proved to be specific, reversible and competitive. During the separation process, two inactive bisindole alkaloids, conodurine and tabernaelegantine A, were also isolated. The data suggest that the substitutions at the carbons 11′, 12′ and 16′ might affect the acetylcholinesterase inhibitory activity.

Zingiber officinale Mitigates Brain Damage and Improves Memory Impairment in Focal Cerebral Ischemic Rat

Cerebral ischemia is known to produce brain damage and related behavioral deficits including memory. Recently, accumulating lines of evidence showed that dietary enrichment with nutritional antioxidants could reduce brain damage and improve cognitive function. In this study, possible protective effect of Zingiber officinale, a medicinal plant reputed for neuroprotective effect against oxidative stress-related brain damage, on brain damage and memory deficit induced by focal cerebral ischemia was elucidated. Male adult Wistar rats were administrated an alcoholic extract of ginger rhizome orally 14 days before and 21 days after the permanent occlusion of right middle cerebral artery (MCAO). Cognitive function assessment was performed at 7, 14, and 21 days after MCAO using the Morris water maze test. The brain infarct volume and density of neurons in hippocampus were also determined. Furthermore, the level of malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) in cerebral cortex, striatum, and hippocampus was also quantified at the end of experiment. The results showed that cognitive function and neurons density in hippocampus of rats receiving ginger rhizome extract were improved while the brain infarct volume was decreased. The cognitive enhancing effect and neuroprotective effect occurred partly via the antioxidant activity of the extract. In conclusion, our study demonstrated the beneficial effect of ginger rhizome to protect against focal cerebral ischemia.

Acetylcholinesterase-inhibiting activity of pyrrole derivatives from a novel marine gliding bacterium, Rapidithrix thailandica.

Acetylcholinesterase-inhibiting activity of marinoquinoline A (1), a new pyrroloquinoline from a novel species of a marine gliding bacterium Rapidithrix thailandica, was assessed (IC50 4.9 μM). Two related pyrrole derivatives, 3-(2′-aminophenyl)-pyrrole (3) and 2,2-dimethyl-pyrrolo-1,2-dihydroquinoline (4), were also isolated from two other strains of R. thailandica. The isolation of 3 from a natural source is reported here for the first time. Compound 4 was proposed to be an isolation artifact derived from 3. The two isolated compounds were virtually inactive in the acetylcholinesterase-inhibitory assay (enzyme inhibition < 30% at 0.1 g L−1).

Acetylcholinesterase-inhibiting steroidal alkaloid from the sponge Corticium sp.

A new stigmastane-type steroidal alkaloid, 4-acetoxy-plakinamine B (1), was isolated from the Thai sponge Corticium sp. The compound was subjected to the acetylcholinesterase inhibitory activity determination to reveal a high inhibitory activity (IC(50) 3.75+/-1.69 microM). The kinetics of enzyme inhibition showed a decrease in V(max), whereas K(m) was increased, thus suggesting an unusual mixed-competitive mode of inhibition. Compound 1 is the first steroidal alkaloid bearing a stigmastane skeleton ever been reported to exhibit such good potency in the acetylcholinesterase inhibition bioassay.