Choon-Sheen Lai

Typhonium flagelliforme inhibits cancer cell growth in vitro and induces apoptosis: An evaluation by the bioactivity guided approach

AIM OF THE STUDY Typhonium flagelliforme (Lodd.) Blume (Araceae) is a Malaysian plant used locally to combat cancer. In order to evaluate its antiproliferative activity in vitro and to possibly identify the active chemical constituents, a bioactivity guided study was conducted on the extracts of this plant. MATERIALS AND METHODS The active extracts of Typhonium flagelliforme were fractionated by flash column chromatography and each fraction was evaluated for antiproliferative activity using MTT assay. The apoptotic effect of the active fraction was determined microscopically and by using TUNEL colorimetric assay. GC-MS and NMR were used to determine the chemical constituents of this active fraction. RESULTS Several fractions of the hexane and dichloromethane extracts were found to inhibit the growth of NCI-H23 non-small cell lung carcinoma cell line significantly, with IC(50)<15 microg/ml. However, most of these active fractions were also found to inhibit the growth of non-tumorigenic BALB/c 3T3 mouse fibroblast cell line except for fraction 21 of the dichloromethane extract (D/F21). This particular fraction was not only less cytotoxic to the non-tumorigenic cells, where the IC(50) was 48.6 microg/ml compared to IC(50) 7.5 microg/ml for NCI-H23, but it was also found to induce apoptosis in the cancer cell line. GC-MS analysis revealed that D/F21 contains hexadecanoic acid, 1-hexadecene, phytol and a derivative of phytol. The presence of non-saturated fatty acids in this fraction was confirmed by nuclear magnetic resonance spectroscopy. CONCLUSIONS D/F21 was found to be the active and cancer cell line specific fraction of Typhonium flagelliforme. Its major chemical constituents had been determined spectroscopically.

Validation of high performance liquid chromatography-electrochemical detection methods with simultaneous extraction procedure for the determination of artesunate, dihydroartemisinin, amodiaquine and desethylamodiaquine in human plasma for application in clinical pharmacological studies of artesunate-amodiaquine drug combination.

With the expanded use of the combination of artesunate (AS) and amodiaquine (AQ) for the treatment of falciparum malaria and the abundance of products on the market, comes the need for rapid and reliable bioanalytical methods for the determination of the parent compounds and their metabolites. While the existing methods were developed for the determination of either AS or AQ in biological fluids, the current validated method allows simultaneous extraction and determination of AS and AQ in human plasma. Extraction is carried out on Supelclean LC-18 extraction cartridges where AS, its metabolite dihydroartemisinin (DHA) and the internal standard artemisinin (QHS) are separated from AQ, its metabolite desethylamodiaquine (DeAQ) and the internal standard, an isobutyl analogue of desethylamodiaquine (IB-DeAQ). AS, DHA and QHS are then analysed using Hypersil C4 column with acetonitrile-acetic acid (0.05M adjusted to pH 5.2 with 1.00M NaOH) (42:58, v/v) as mobile phase at flow rate 1.50ml/min. The analytes are detected with an electrochemical detector operating in the reductive mode. Chromatography of AQ, DeAQ and IB-DeAQ is carried out on an Inertsil C4 column with acetonitrile-KH(2)PO(4) (pH 4.0, 0.05M) (11:89, v/v) as mobile phase at flow rate 1.00ml/min. The analytes are detected by an electrochemical detector operating in the oxidative mode. The recoveries of AS, DHA, AQ and DeAQ vary between 79.1% and 104.0% over the concentration range of 50-1400ng/ml plasma. The accuracies of the determination of all the analytes are 96.8-103.9%, while the variation for within-day and day-to-day analysis are <15%. The lower limit of quantification for all the analytes is 20ng/ml and limit of detection is 8ng/ml. The method is sensitive, selective, accurate, reproducible and suited particularly for pharmacokinetic study of AS-AQ drug combination and can also be used to compare the bioavailability of different formulations, including a fixed-dose AS-AQ co-formulation.

Cholinesterase inhibitory activity and chemical constituents of Stenochlaena palustris fronds at two different stages of maturity

Stenochlaena palustris fronds are popular as a vegetable in Southeast Asia. The objectives of this study were to evaluate the anticholinesterase properties and phytochemical profiles of the young and mature fronds of this plant. Both types of fronds were found to have selective inhibitory effect against butyrylcholinesterase compared with acetylcholinesterase. However, different sets of compounds were responsible for their activity. In young fronds, an antibutyrylcholinesterase effect was observed in the hexane extract, which was comprised of a variety of aliphatic hydrocarbons, fatty acids, and phytosterols. In the mature fronds, inhibitory activity was observed in the methanol extract, which contained a series of kaempferol glycosides. Our results provided novel information concerning the ability of S. palustris to inhibit cholinesterase and its phytochemical profile. Further research to investigate the potential use of this plant against Alzheimers disease is warranted, however, young and mature fronds should be distinguished due to their phytochemical differences.

Polyphenols rich fraction of Dicranopteris linearis promotes fibroblast cell migration and proliferation in vitro

ETHNOPHARMACOLOGICAL RELEVANCE Dicranopteris linearis is a fern used traditionally for the treatment of skin afflictions such as external wounds, boils and ulcers. However, there are no scientific studies to date to demonstrate its ability to induce wound recovery. The objective of the present study was to explore the wound healing properties of an active fraction of D. linearis through several in vitro assays and to determine its chemical profile. MATERIALS AND METHODS The antioxidant activities were evaluated by DPPH and FRAP assays. The ability of the active fraction to induce fibroblast cell proliferation in serum deprived medium or under the challenge of exogenous hydrogen peroxide was evaluated by MTT assay. Cell migration was investigated using the scratch-wound assay. Chemical constituents of the active fraction were isolated and their structures were determined by NMR and MS spectroscopic techniques. Chemical profiling was carried out by the HPLC-UV method. RESULTS The fifth subfraction of the methanol extract of D. linearis (sF5) showed prominent antioxidant activities. It was non-cytotoxic at the concentration tested and was able to induce cell proliferation of selected fibroblast cells under serum deprived conditions. In addition, sF5 was found to enhance cell migration and promote cellular repair following oxidative damage caused by hydrogen peroxide through a mechanism which was independent of its hydroxyl radical scavenging capability. Six phenolic glycosides were identified in sF5 where three were known, two were new and one phenolic glycoside first time reported from the plant. CONCLUSION The active fraction of D. linearis which was rich in polyphenolic substances exhibited substantial antioxidant activities, induced cellular repair and contributed positively to fibroblasts proliferation and migration in vitro. Hence, it may have potential application in inducing wound recovery and supports its ethnopharmacological use for treating skin problems.

Anti-Tumor Action, Clinical Biochemistry Profile and Phytochemical Constituents of a Pharmacologically Active Fraction of S. crispus in NMU-Induced Rat Mammary Tumour Model

Cancer patients seek alternative remedies such as traditional medicinal plants for safe and effective treatment and help overcome the side effects of conventional therapy. Current knowledge indicates that extracts of Strobilanthes crispus of the Acanthaceae family exhibit potent anticancer properties in vitro and are non-toxic in vivo. S. crispus was also reported to be protective against chemical hepatocarcinogenesis. We previously showed that a bioactive fraction of S. crispus leaves also synergized with tamoxifen to cause apoptosis of human breast cancer cell lines without damaging non-malignant epithelial cells. The present study aimed to evaluate the antitumor effect of S. crispus dichloromethane fraction (F3) using N-methyl-N-Nitrosourea (NMU)-induced rat mammary tumor model. Tumor regression was observed in 75% of the rats following 8-week oral administration of F3 with no secondary tumour formation and no signs of anemia or infection. However, no improvement in the liver and renal function profiles was observed. Major constituents of F3 were identified as lutein, 131-hydroxy-132-oxo-pheophytin a, campesterol, stigmasterol, β-sitosterol, pheophytin a and 132-hydroxy-pheophytin a. These compounds however, may not significantly contribute to the antitumor effect of F3.

A Comparative Study of the Volatile Constituents of Southeast Asian Coffea arabica, Coffea liberica and Coffea robusta Green Beans and their Antioxidant Activities

Abstract A comparative study on the volatile constituents of green beans of three commercially grown coffees in Southeast Asia, namely Coffea arabica, Coffea robusta and Coffea liberica, and their antioxidant properties was carried out. Volatile constituents of the green beans were isolated by hydrodistillation followed by extraction of the distillates with dichloromethane. These compounds were analysed using capillary gas chromatography and gas chromatography-mass spectrometry. The volatile compositions of the green beans of these three species were remarkably similar, being characterized by high levels of phenolic compounds, principally p-vinylguaiacol, and carboxylic acids. The volatile extracts were examined for antioxidant activity using total phenols, DPPH radical scavenging and FRAP assays which showed that the volatile extract from C. liberica possessed the highest antioxidant capacity, followed by those of C. arabica and C. robusta, respectively.