Antioxidant and Acetylcholinesterase Inhibitor Potentials of the Stem Extract of Pternandra galeata / Potenciales antioxidantes e inhibidores de la acetilcolinesterasa del extracto de tallo de Pternandra galeata

Background: Pternandra galeata belongs to the family Melastomataceae. It is a native flowering plant in Borneo Island that serve as food for monkey habitat. There has been limited study on the medicinal and chemical properties of this plant. Objectives: We investigated the acetylcholinesterase inhibitory activity and evaluated the antioxidant activity of the ethanolic extract of Pternandra galeata stem. The total phenolic content in the sample was also determined. Methods: The acetylcholinesterase inhibitory assays were performed using Ellman's method. Two different methods were used to evaluate the antioxidant activity of the extract by 2,2-diphenyl-1-picryl hydrazyl (DPPH) and 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assays. The total phenolic content was determined by the Folin-Ciocalteu method by employing gallic acid as a reference. Results: The ethanolic extract of the P. galeata stems inhibited the AChE enzyme with an IC50 value of 74.62 ± 0.89 µg/mL.The sample exhibited antioxidant activity in the DPPH assay with an IC50 value of 20.21 ± 0.08 µg/mL and 7.68 ± 0.09 µg/mL in the ABTS scavenging assay. The total phenolic content was 164.71 ± 3.33 mg GAE/g extract. Conclusion: The ethanolic extract of the P. galeata stem can be a promising cholinesterase inhibitor and antioxidant for treating Alzheimer's disease

Antecedentes: Pternandra galeata pertenece a la familia Melastomataceae. Se trata de una planta con flores nativa de la isla de Borneo que sirve de alimento para el hábitat de los monos. Se han realizado pocos estudios sobre las propiedades medicinales y químicas de esta planta. Objetivos: Se investigó la actividad inhibidora de la acetilcolinesterasa y se evaluó la actividad antioxidante del extracto etanólico del tallo de Pternandra galeata. También se determinó el contenido fenólico total de la muestra. Métodos: Los ensayos de inhibición de la acetilcolinesterasa (AChE) se realizaron mediante el método de Ellman. Se utilizaron dos métodos diferentes para evaluar la actividad antioxidante de los ensayos de 2,2-difenil-1-picril hidrazilo (DPPH) y 2,2'-azinobis-(ácido 3-etilbenzotiazolina-6-sulfónico) (ABTS). El contenido fenólico total se determinó por el método de Folin-Ciocalteu empleando el ácido gálico como referencia. Resultados: El extracto etanólico de los tallos de P. galeata inhibió la enzima AChE con un valor IC50 de 74.62 ± 0.89 µg/mL. La muestra mostró actividad antioxidante en el ensayo DPPH con un valor IC50 de 20.21 ± 0.08 µg/mL y 7.68 ± 0.09 µg/mL en el ensayo de barrido ABTS. El contenido fenólico total fue de 164.71 ± 3.33 mg GAE/g de extracto. Conclusión: El extracto etanólico del tallo de P. galeata puede ser un prometedor inhibidor de la colinesterasa y antioxidante para el tratamiento de la enfermedad de Alzheimer.

Diterpenoid alkaloids from roots of Aconitum kongboense / 中国中药杂志

The chemical constituents from the roots of Aconitum kongboense were studied. Twenty-five diterpenoid alkaloids were isolated from the 95% methanol extract of the roots of A. kongboense by silica gel, reverse-phase silica gel and basic alumina column chromatography. They included a new aconitine-type diterpenoid alkaloid, named as kongboensenine(1), and twenty-four known ones(2-25), i.e., acotarine F(2), acotarine G(3), 14-acetyltalatisamine(4), talatisamine(5), indaconitine(6), yunaconitine(7), chasmanine(8), 6-epi-foresticine(9), homochasmanine(10), 8-deacetyl-yunaconitine(11), chasmaconitine(12), ajaconine(13), franchetine(14), ezochasmanine(15), crassicautine(16), 14-O-deacylcrassicausine(17), genicunine A(18), falconeridine(19), sachaconitine(20), liljestrandisine(21), 8-methyl-14-acetyltalatisamine(22), kongboendine(23), 14-benzoylchasmanine(24) and pseudaconine(25). Their structures were elucidated by common spectroscopic methods including high-resolution electrospray ionization mass spectrometry(HR-ESI-MS) and nuclear magnetic resonance(NMR) techniques. Compounds 2-4, 10, 13, 15-19 and 21-22 were isolated from this plant for the first time. Experimental results showed that all compounds did not have a significant inhibitory activity against acetylcholinesterase(AChE).

Determination of donepezil in human plasma using ultra performance liquid chromatography-tandem mass spectrometry

An ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the quantification of donepezil in human plasma. Donepezil and donepezil-D4 were extracted from human plasma by liquid-liquid extraction using a mixture of hexane and ethyl acetate (70:30 v/v). The extracted samples were analyzed using a Thermo Hypersil Gold C18 column with 5% acetic acid in 20 mM ammonium acetate buffer (pH 3.3) and 100% acetonitrile as a mobile phase with the 60:40 (v:v) isocratic method, at a flow rate of 0.3 mL/min. The injection volume was 3 µL, and the total run time was 3 min. Inter- and intra-batch accuracies ranged from 98.0% to 110.0%, and the precision was below 8%. The developed method was successfully applied to the quantification of donepezil in human plasma. The mean (standard deviation) maximum concentration and the median (range) time to maximum concentration were 8.6 (2.0) ng/mL and 2.0 h (1.0~5.0 h), respectively, in healthy Koreans after oral administration of 5 mg donepezil.

Study on preparation of immobilized acetylcholinesterase microreactor and its application in CMM inhibitors screening / 中草药

Objective To prepare an immobilized acetylcholinesterase (AChE) microreactor and establish a rapid screening method for Chinese materia medica (CMM) acetylcholinesterase inhibitors (AChEIs). Methods A novel immobilized AchE microreactor was prepared by crosslinking with glutaraldehyde, using aminated magnetic microspheres as carrier. The characterizations were conducted by physicochemical properties and chromatographic performance. The immobilized AChE reactor was used to screen AChEIs from the Huperzia Serratum extracts. Results In the enzyme reaction system, the optimum substrate concentration was 50 μmol/L, and the incubation time was 5 min, respectively. IR characterization, specificity verification, enzyme kinetics, and stability study results all demonstrated the effectiveness of the enzyme reactor. The CMM AChEI, huperzine A, was obtained from the screening of H. Serratum extracts. Conclusion A high throughput screening method for AChEIs is established in this paper, which will be further applied and popularized.

Synthesis and Acetylcholinesterase Inhibitory Activity of 6-Benzyl-3-aryl-7H-thiazolo3, 2-b-1, 2, 4 -triazin- 7-one Derivatives / 中国药学杂志

OBJECTIVE: To explore the synthesis and acetylcholinesterase inhibitory activity of 6-benzyl-3-aryl-7H-thiazolo[3, 2 - b]-1, 2, 4 -triazin-7-one derivatives. METHODS: Benzaldehyde and acetylglycine were used as raw materials and underwent Erlenmeyer- Plöchl reaction, condensation reaction, hydrolysis reaction, condensation reaction to obtain 6-benzyl-3-thioxo-3, 4 -dihydro-1, 2, 4-triazin-5(2H) -ones derivatives. The derivatives reacted with substituted α-phenacyl chlorides to generate 6-benzyl-3-(hydroxylaryl) - 7H-thiazolo[3, 2 -b]-1, 2, 4 -triazin-7-ones derivatives. Then, Williamson reaction was used to yield 6-benzyl-3-aryl-7H-thiazolo[3, 2 - b]-1, 2, 4 -triazin-7-ones as target compounds. RESULTS: Nine 6-benzyl-3-aryl-7H-thiazolo[3, 2 -b]-1, 2, 4 -triazin-7-ones were prepared as target compounds. All target compounds exhibited inhibitory activities against human AChE in vitro, five of which had inhibitory rates above 50% at 10 μmol•L-1. CONCLUSION: Based on the screening results of AChE inhibitory activity in vitro and docking studies, there are some interactions between 6-benzyl-3-aryl-7H-thiazolo[3, 2 -b]-1, 2, 4 -triazin-7-one derivatives and the anionic binding site and PAS zones of AChE, and the target compounds have exhibited AChE inhibitory activities.

Design, synthesis and evaluation of 4-pyridinylthiazole-2-amines as acetylcholinesterase inhibitors / 药学学报

Alzheimer's disease (AD) is a degenerative disease of the nervous system. Compound I reported to have inhibitory activity on AChE was used as a lead compound in this study, and 4-pyridinylthiazole-2-amines were designed by optimizing compound I structure. The new compounds were synthesized from acetylpyridines through five-steps of reaction, and their inhibition activities on AChE were measured in vitro by Ellman method. The new compounds exhibited a clear inhibitory activity on AChE in vitro. The bioactivity of compound 13c was the best among them, and its IC50 value was 0.15μmol·L-1, which was better than that of rivastigmine and compound I in the control. Meanwhile, it exhibited little inhibition on butyrylcholinesterase. So the selective inhibitory activities of 4-pyridinylthiazole-2-amines to acetylcholinesterase were worth of studying furtherly.

Clinical Significance of Repetitive Compound Muscle Action Potentials in Patients with Myasthenia Gravis: A Predictor for Cholinergic Side Effects of Acetylcholinesterase Inhibitors

BACKGROUND AND PURPOSE: Acetylcholinesterase inhibitors (AChEIs) are widely used to treat myasthenia gravis (MG). Although AChEIs are usually tolerated well, some MG patients suffer from side effects. Furthermore, a small proportion of MG patients show cholinergic hypersensitivity and cannot tolerate AChEIs. Because repetitive compound muscle action potentials (R-CMAPs) are an electrophysiologic feature of cholinergic neuromuscular hyperactivity, we investigated the clinical characteristics of MG patients with R-CMAPs to identify their clinical usefulness in therapeutic decision-making. METHODS: We retrospectively reviewed the clinical records and electrodiagnostic findings of MG patients who underwent electrodiagnostic studies and diagnostic neostigmine testing (NT). RESULTS: Among 71 MG patients, 9 could not tolerate oral pyridostigmine bromide (PB) and 17 experienced side effects of PB. R-CMAPs developed in 24 patients after NT. The highest daily dose of PB was lower in the patients with R-CMAPs (240 mg/day vs. 480 mg/day, p<0.001). The frequencies of PB intolerance and side effects were higher in the patients with R-CMAPs than in those without R-CMAPs [37.5% vs. 0% (p<0.001) and 45.8% vs. 12.8% (p=0.002), respectively]. The MG Foundation of America postintervention status did not differ significantly between MG patients with and without R-CMAPs, and the response to immunotherapy was also good in both groups. CONCLUSIONS: Side effects of and intolerance to AChEIs are more common in MG patients with R-CMAPs than in those without R-CMAPs. AChEIs should be used carefully in MG patients with R-CMAPs. The presence of R-CMAPs after NT may be a good indicator of the risks of PB side effects and intolerance.

Active anti-acetylcholinesterase component of secondary metabolites produced by the endophytic fungi of Huperzia serrata

Background An endophytic fungus lives within a healthy plant during certain stages of, or throughout, its life cycle. Endophytic fungi do not always cause plant disease, and they include fungi that yield different effects, including mutual benefit, and neutral and pathogenic effects. Endophytic fungi promote plant growth, improve the host plant's resistance to biotic and abiotic stresses, and can produce the same or similar biologically active substances as the host. Thus, endophytic fungal products have important implications in drug development. Result Among the numerous endophytic fungi, we identified two strains, L10Q37 and LQ2F02, that have anti-acetylcholinesterase activity, but the active compound was not huperzine A. The aim of this study was to investigate the anti-acetylcholinesterase activity of secondary metabolites isolated from the endophytic fungi of Huperzia serrata. Microbial cultivation and fermentation were used to obtain secondary metabolites. Active components were then extracted from the secondary metabolites, and their activities were tracked. Two compounds that were isolated from endophytic fungi of H. serrata were identified and had acetylcholine inhibitory activities. In conclusion, endophytic fungal strains were found in H. serrata that had the same anti-acetylcholinesterase activity. Conclusion We isolated 4 compounds from the endophytic fungus L10Q37, among them S1 and S3 are new compounds. 6 compounds were isolated from LQ2F02, all 6 compounds are new compounds. After tested anti acetylcholinesterase activity, S5 has the best activity. Other compounds' anti acetylcholinesterase activity was not better compared with huperzine A.

Single Photon Emission Computerized Tomography and Neuropsychological Tests That Predict a Good Response to Donepezil Therapy for Alzheimer's Disease

BACKGROUND AND PURPOSE: Cholinesterase inhibitors (ChEIs) are effective in Alzheimer's disease (AD) treatment. The aim of this study is 1) to find neuropsychological factors that affect the functional response to ChEI therapy and 2) to determine whether regional cerebral blood flow (rCBF) pretreatment predicts a cognitive change in response to ChEI. METHODS: We prospectively recruited 32 patients diagnosed with probable AD and treated them with donepezil, a ChEI, over one year. The patients were divided into stable (s-AD) and declined (d-AD) AD groups, based on changes in Korean version of Mini-Mental State Examination (K-MMSE) scores. Patients were assessed using the Alzheimer's Disease Co-operative Study-Activities of Daily Living (ADCS-ADL) and Seoul Neuropsychologic Screening Battery, as well as brain single photon emission computerized tomography (SPECT) at baseline and last medical evaluations. The predictors of therapeutic responses were analyzed using general linear models. RESULTS: Based on their cognitive function changes, AD patients were classified into two groups: s-AD (n=14, annual change in K-MMSE score or =0.9). The s-AD at baseline showed significantly better ADCS-ADL function (p=0.04) and had a tendency to preserve frontal function compared to the d-AD group. Global Statistical Parametric Mapping analysis revealed no significant decrease of rCBF between baseline and follow-up SPECT, in either the s-AD or the d-AD groups. However, on regional perfusion analysis of baseline SPECT, the d-AD group demonstrated perfusion deficits in the supramarginal gyrus, inferior occipital gyrus, and rolandic operculum compared with the s-AD group. CONCLUSIONS: Highly preserved ADCS-ADLs predicted a better improvement in MMSE scores in response to ChEI therapy and a more positive functional response in the group with preserved frontal function. rCBF provided hints to the variable response to donepezil therapy with ChEI treatment.

Biological Predictors of Alzheimer's Disease Treatment

Variety of biomarkers that are related to the Alzheimer's disease and its diagnosis and progress have been found. However, research lacks in predicting the reaction of the treatment. In addition, there is no definite treatment reaction to the disease but rather it is varied. The purpose of this review article is to study the research of the biomarkers that are able to predict the treatment reaction. There was a research that illustrated a relationship between plasma amyloid beta peptide, cerebrospinal fluid tau, neuroanatomical biomarkers and acetylcholinesterase inhibitors. Polymorphisms in genes of the cholinergic markers AChE, BuChE, ChAT and PON-1 were found to be associated with better clinical response to acetylcholinesterase inhibitors. Many pharmacogenetic studies have been conducted to evaluate the impact of the lipoprotein apolipoprotein E (APOE) genotype on treatment response to acetylcholinesterase inhibitor. However, there is no significant influence of the APOE genotypes on treatment response. Further research is needed to find other predictors of treatment with acetylcholinesterase inhibitors in patients with Alzheimer's disease.

Research on quality specification of the seeds of Peganum harmala L. of a uygur traditional medicine / 中国药学杂志

OBJECTIVE: To develop the quality specification of seeds of Peganum harmala. METHODS: According to the Chinese Pharmacopoeia (2010 Version, Volume 1) and its appendix method, the water, total ash, acid insoluble ash, 50% ethanol extractives, and heavy metal were analyzed for seeds of P. harmala. TLC method was used to separate harmaline (HAL), harmine (HAR) and vasicine (VAS) in seed samples using mixture of ethyl acetate-methanol-ammonia water (10:1.5:0.5) as a developing solvent on high performance silica G pre-coated plate with 254 nm fluorescent (GF254) and to identify them inspected under UV 366 nm, 254 nm, visualized by spraying with both Dragendorff reagent and by bioautographic assay. In the HPLC method, HAL and HAR were separated on a C18 column with acetonitrile-ammonium acetate water (19:81) as the mobile phase and detected at 330 nm. The HPLC fingerprints were performed on the same C18 column and eluted by using a linear gradient of acetonitrile (A) and 0.1 mmol · L-1 ammonium acetate buffer under the flow rate at 0.7 mL · min-1 and detected at 280 nm. RESULTS: In the TLC procedures, 254 and 366 nm fluorescent, Dragendorff reagent, and bioautographic assay for the detection of acetylcholinesterase inhibitor can be used for qualitative identification of the active ingredients. For the HPLC quantitative method, the calibration curve of HAR displayed ideal linearity over the range of 1.97-198.68 μg · mL-1 with average recovery of 99.69% (RSD of 1.89%). HAL displayed ideal linearity over the range of 1.70-345.30 μg · mL-1 with average recovery of 100.66% (RSD of 1.78%). The contents of HAL and HAR in 11 batches of seeds of P. harmala were 3.234% and 3.755%. In the characteristic fingerprints of seeds of P. harmala, four common peaks were identified. CONCLUSION: The results indicated that the water, total ash, acid insoluble ash, and 50% ethanol extractives were not more than 9.0%, 8.0%, 1.0%, and 22.0%, respectively. The heavy metal of plumbum, cadmium, arsenic, mercury, and copper were not more than 5 × 10-6, 3 × 10-6, 2 × 10-6, 2 × 10-6 and 20 × 10-6, respectively. The content limit of the sum of HAL and HAR was not lower than 5.5%. With the peak of HAL as reference peak, the variance of relative retention time of the four common peaks, in the characteristic fingerprints of seeds off. harmala, should be fluctuated in the range of 5% of the specified value. The qualitative and quantitative method established was suitable for the quality evaluation and assessment of seeds of P. harmala.

Clinical and Electrophysiologic Responses to Acetylcholinesterase Inhibitors in MuSK-Antibody-Positive Myasthenia Gravis: Evidence for Cholinergic Neuromuscular Hyperactivity

BACKGROUND AND PURPOSE: Patients with muscle-specific tyrosine kinase (MuSK) antibody (MuSK-Ab)-positive myasthenia gravis (MG) show distinct responses to acetylcholinesterase inhibitors (AChEIs). Although clinical responses to AChEIs in MuSK-Ab MG are reasonably well known, little is known about the electrophysiologic responses to AChEIs. We therefore investigated the clinical and electrophysiologic responses to AChEIs in MuSK-Ab-positive MG patients. METHODS: We retrospectively reviewed the medical records and electrodiagnostic findings of 17 MG patients (10 MuSK-Ab-positive and 7 MuSK-Ab-negative patients) who underwent electrodiagnostic testing before and after a neostigmine test (NT). RESULTS: The frequency of intolerance to pyridostigmine bromide (PB) was higher in MuSK-Ab-positive patients than in MuSK-Ab-negative patients (50% vs. 0%, respectively; p=0.044), while the maximum tolerable dose of PB was lower in the former (90 mg/day vs. 480 mg/day, p=0.023). The frequency of positive NT results was significantly lower in MuSK-Ab-positive patients than in MuSK-Ab-negative patients (40% vs. 100%, p=0.035), while the nicotinic side effects of neostigmine were more frequent in the former (80% vs. 14.3%, p=0.015). Repetitive compound muscle action potentials (R-CMAPs) developed more frequently after NT in MuSK-Ab-positive patients than in MuSK-Ab-negative patients (90% vs. 14.3%, p=0.004). The frequency of a high-frequency-stimulation-induced decrement-increment pattern (DIP) was higher in MuSK-Ab-positive patients than in MuSK-Ab-negative patients (100% vs. 17.7%, p=0.003). CONCLUSIONS: These results suggest that MuSK-Ab-positive MG patients exhibit unique and hyperactive responses to AChEIs. Furthermore, R-CMAP and DIP development on a standard AChEI dose may be a distinct neurophysiologic feature indicative of MuSK-Ab-positive MG.

Design, synthesis and evaluation of thiochroman-4-ones as acetylcholinesterase inhibitors / 中国药学杂志

OBJECTIVE: To research the effect of thiochroman-4-one derivatives on acetylcholine esterase activity. METHODS: Substituted thiophenols as the starting materials were transformed into 6-hydroxythiochroman-4-ones by substitution reaction and cycliza-tion reaction with concentrated sulfuric acid, and the resultant compounds were reacted with benzylpiperazins to prepare 3-(4-benzylpiperazine-1-methyl) thiochroman-4-ones hydrochloride, and their bioactivities as AChE inhibitors were measured in vitro. RESULTS: The target compounds had AChE inhibitive activity in micromolar range, and compound 8a was better than rivastigmine with IC50=0.96 μmol · L-1. CONCLUSION: 3-(4-Benzylpiperazine-1-methyl) thiochroman-4-ones show anti-acetylcholinesterase activity in vitro, which are worth further research.

Isolation, Identification and Characterization of a Antidementia Acetylcholinesterase Inhibitor-Producing Yarrowia lipolytica S-3

This report describes the isolation and identification of a potent acetylcholinesterase (AChE) inhibitor-producing yeasts. Of 731 species of yeast strain, the S-3 strain was selected as a potent producer of AChE inhibitor. The selected S-3 strain was investigated for its microbiological characteristics. The S-3 strain was found to be short-oval yeast that did not form an ascospore. The strain formed a pseudomycelium and grew in yeast malt medium containing 50% glucose and 10% ethanol. Finally, the S-3 strain was identified by its physiological characteristics and 26S ribosomal DNA sequences as Yarrowia lipolytica S-3.

Reversible acetylcholinesterase inhibitory effect of Tabernaemontana divaricata extract on synaptic transmission in rat CA1 hippocampus.

Background & objectives: Acetylcholinesterase inhibitors (AChE-Is) are used for the treatment of Alzheimer’s disease (AD). These drugs including galanthamine have been shown to modulate synaptic activity in hippocampus and improve memory processes. Although Tabernaemontana divaricata extract (TDE) has been used as traditional medicine for various pharmacological effects, its effect in enhancing cholinergic activity provides additional benefit to its known effects. We investigated whether TDE can modulate the synaptic function in hippocampus and compared its effects to those of galanthamine. Methods: Hippocampal slices were prepared from male wWistar rats, functional effects of TDE were characterized by using pharmacological tools and extracellular recordings of field excitatory postsynaptic potentials (fEPSPs). Results: TDE significantly reduced fEPSPs. The fEPSPs reduction was prevented by atropine, but not pancuronium. These TDE effects were similar to those of galanthamine. Interpretation & conclusions: Our findings indicate that TDE can effectively modulate synaptic responses in the hippocampus similar to galanthamine, suggesting that this traditional medicine could be beneficial in ageing with ACh deprivation in the brain.

Nutritional and Physicochemical Characteristics of the Antidementia Acetylcholinesterase-Inhibiting Methanol Extracts from Umbilicaria esculenta

To develop new antidementia nutraceuticals, a potent acetylcholinesterase (AChE)-inhibiting extract was screened from various extracts of nutritional mushrooms and lichens nutritional and its physicochemical properties were investigated. Among the several extracts tested, methanol extracts of Umbilicaria esculenta fruiting body showed the highest AChE inhibitory activity of 22.4%. U. esculenta AChE inhibitor was maximally extracted when fruiting bodies were treated with 80% methanol at 40degrees C for 18 h. The methanol extracts contained 18.9% crude lipid, 18.8% crude protein, and 11.6% total sugar. In addition, they contained 444 mg/g glutamic acid, 44 mg/g histidine, and 41 mg/g aspartic acid. The methanol extracts were soluble in a solution of methanol and 20% dimethylsulfoxide, insoluble in n-hexane, chloroform, and water, and were stable at 20~60degrees C and pH 1.0~5.0 for 1 h.

Construction of the pharmacophore model of acetylcholinesterase inhibitor / 药学学报

Based on ninety three acetylcholinesterase inhibitors (AChEIs) which have the same mechanism of action but are different in structural characteristics, the pharmacophore model for acetylcholinesterase inhibitor was constructed by the CATALYST system. The optimal pharmacophore model with three hydrophobic units, a ring aromatic unit and a hydrogen-bond acceptor unit were confirmed (Weight=3.29, RMS=0.53, total cost-null cost=62.75, Correl=0.93, Config=19.05). This pharmacophore model will act on the double active site of acetylcholinesterase and is able to predict the activity of known acetylcholinesterase inhibitors that are used for clinical treatment of Alzheimer's disease (AD), and can be further used to identify structurally diverse compounds that have higher activity treating with Alzheimer's disease (AD) by virtual screening.

Production of the Acetylcholinesterase Inhibitor from Yarrowia lipolytica S-3

The acetylcholinesterase (AChE) inhibitor of Yarrowia lipolytica S-3 was maximally produced when it was incubated at 30degrees C for 36 h in an optimal medium containing 1% yeast extract, 2% peptone and 2% glucose, with an initial pH 6.0. The final AChE inhibitory activity under these conditions was an IC50 value of 64 mg/ml. After partial purification of the AChE inhibitor by means of systematic solvent extraction, the final IC50 value of the partially purified AChE inhibitor was 0.75 mg/ml. We prepared a test product by using the partially purified AChE inhibitor and then determined its stability for the development of a new antidementia commercial product. The test product was stable at room temperature for 15 weeks.

Effect of Itopride Hydrochloride on the Ileal and Colonic Motility in Guinea Pig In Vitro

PURPOSE: Itopride hydrochloride (itopride) inhibits acetylcholinesterase (AChE) and antagonizes dopamine D(2) receptor, and has been used as a gastroprokinetic agent. However, its prokinetic effect on the small bowel or colon has not yet been thoroughly investigated. The aim of this study was to investigate the effects of itopride on motor functions of the ileum and colon in guinea pigs. MATERIALS AND METHODS: The distal ileum was excised and the activity of peristaltic contraction was determined by measuring the amplitude and propagation velocity of peristaltic contraction. The distal colon was removed and connected to the chamber containing Krebs-Henseleit solution (K-H solution). Artificial fecal matter was inserted into the oral side of the lumen, and moved toward the anal side by intraluminal perfusion via peristaltic pump. Colonic transit times were measured by the time required for the artificial feces to move a total length of 10cm with 2-cm intervals. RESULTS: In the ileum, itopride accelerated peristaltic velocity at higher dosage (10(-10)-10(-6)M) whereas neostigmine accelerated it only with a lower dosage (10(-10)-10(-9)M). Dopamine (10(-8)M) decelerated the velocity that was recovered by itopride infusion. Itopride and neostigmine significantly shortened colonic transit at a higher dosage (10(-10)-10(-6)M). Dopamine (10(-8)M) delayed colonic transit time that was also recovered after infusion of itopride. CONCLUSION: Itopride has prokinetic effects on both the ileum and colon, which are regulated through inhibitory effects on AChE and antagonistic effects on dopamine D(2) receptor.

Comparative study on microassays for screening acetylcholinesterase inhibitors / 中国药理学通报

Aim To provide practical microassays for screening acetylcholinesterase (AChE) inhibitors in drug discovery.Methods The optimal conditions of assaying the activity of AChEs from electric eel,rat brain homogenate and cobra venom were determined in 96-well plates under physiological pH value and temperature by orthogonal matrix method.The concentrations of AChE,substrate and DTNB,and reaction time were optimized.After the effects of sample solvent (DMSO) used in the assay and stopping reagent on enzyme activity were assessed,the assay conditions were finally selected,and 492 kinds of extracts from Guizhou ethno-drugs were screened.Results Practical microassays for screening AChE inhibitors were successfully constituted by using AChEs mentioned above.The data analysis of screening results revealed that electric eel AChE possessed a high sentivity to inhibitors,and cobra venom AChE shared high similarity with rat brain homogenate in positive results.Conclusion Microassays constituted in this work possessed advantages of being easy,rapid,reliable,cost saving and flexible.AChE from electric eel was especially suitable for screening AChE inhibitors from extracts,and AChE from cobra venom was more suitable to be used in screening AChE inhibitors from large numbers of compounds.