INRA UMR SPO Montpellier/ Centre de Recherches de Biochimie Macromoléculaire (CRBM) - Centre National de la Recherche Scientifique (CNRS)
Faculty of Pharmaceutical Science, Khon Kaen University, Khon Kaen 40002, Thailand
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Samenvatting 1 eindwerk 2013-2014: STUDY OF ANTICANCER ACTIVIY OF THE OPILACEAE EXTRACTS AGAINST CANCER CELL LINES
In this experiment, two types of Thai plants on Opillaceae plants were used which are Melientha suavis Pierre and the Urobotrya siamensis Hiepko. Those two plants are cultivated in Thailand and traditionally used as Thai medicinal plant. The plants were harvested in 3 provinces in Thailand. Urobotrya siamensis Hiepko was harvested from one province and the extract was coded as KB. Melientha suavis Pierre was harvested from two other provinces and was coded as SB and UT followed by their origin. The sample preparation and extractions were already done in the lab by using solvent extraction method by using 4 different solvents with the different in their polarity index. The solvents were water, methanol, hexane and ethyl acetate. The crude extracts of each sample extracted in each solvent was evaporated into dried crude extract. So in this study, there were 12 extracts from 3 plant samples extracted with 4 extraction solvents each. The purpose of this study is to test the anticancer activity of the extracts against cancerous cell by using neutral red cytotoxicity assay. The morphological changes effected by the extract was determined by a DAPI staining and the imaging with fluorescent microscopy.The different types of cell lines colorectal cancer cell line (HCT 116), hepatocarcinoma cell line (HepG2), a leukemia cancer cell line (Jurkat) and a normal vero cell line isolated from the epithelial of the kidney of African green monkey was used as reference. The cell lines were treated by extracts in different concentrations. With these cytotoxicity results the IC50 was calculated.
From the results of the study, the hexane extracts coded as KB and SB gave best results and amongst all tested cancer cell lines, they were found to have the highest sensitivity against colon cancer cell line. As the basic knowledge of cancer treatment, there are two different cell death pathways, necrosis and apoptosis. The programmed cell death, also called apoptosis pathways is commonly known as the prefer cell death pathway with no harmful to healthy neighboring cells. The confirmation of the preferred apoptosis pathway was determined in the point of morphological change. The DAPI nuclear staining is employed to distinguish the cell morphology after treated by the extracts. Further research is necessary to provide more information about the specific bioactive compounds in plant and also the specific mechanism of cell death by the plant extract against cancer cell lines.
Samenvatting 2 eindwerk 2013-2014: Near InfraRed Spectroscopy (NIRS) as an alternative method for 'wet analysis' for the quantitative analysis of benzoic acid in a gel matrix
The goal of this project is to find out whether Near InfraRed Spectroscopy (NIRS) is an alternative method for the classic ‘wet analysis’ for the quantitative analysis of benzoic acid in a gel matrix or not. Gels with different formulations were used to test the alternative method with. This has the purpose to obtain a robust prediction model, so a general PLS model could be built to use for quantitative prediction of benzoic acid.
First, a general comprehension of the NIRS instrument is needed to understand the principles of the vibrational spectroscopy and its absorption phenomenon. Also, a study of Chemometrics is needed to comprehend how obtained data can be analyzed. The quantitative NIRS model is a relative analytical method and therefore this method demands a reference analysis method, whereas HPLC (High Pressure Liquid Chromatography) was chosen as the reference method.
The first step is to determine the solubility of benzoic acid in propylene glycol and make a stock solution of benzoic acid in propylene glycol with optimal solubility, because this solved benzoic acid is added to the placebo samples of the gel in order to create a standard series where the concentration of interest is included into. Once the placebo sample series are prepared, they can be analyzed with NIRS. Each sample is measured for 30 times and the sample is stirred between every measurement. The reference should be scanned between every 10 measurements in order to obtain more stable results. The obtained data is then analyzed with Chemometrics, using the Unscrambler 10.2 software. PLS models with different pre-treatments are built, using PCA models to detect outliers. The optimal PLS model is used to test the batch samples with. The analyzing method of the batch samples is the same as for the placebo samples.
Prediction models were built for gels with four different formulations in order to create one general prediction model afterwards, suitable to predict the amount of benzoic acid in different formulations. The recovery of the NIRS method compared to the HPLC was too low to be accepted. A general prediction model therefore, could not be built.
According to the results, Near InfraRed Spectroscopy cannot be accepted as an alternative method for the ‘wet’ analysis for quantitative determination of benzoic acid in a gel matrix. Additional investigation with a more robust prediction model, can give a better conclusion whether NIRS can be considered as an alternative method or not. The optimal situation is to obtain a robust PLS model, containing al the variability’s that can be faced in real life situation.
123 Moo 16 Mittapap Rd., Nai-Muang, Muang District
40002 Khon Kaen
Dr. Jomjai Peerapattana