Construction of Novel Yeast Strains from Candida tropicalis KBKTI 10.5.1 and Saccharomyces cerevisiae DBY1 to Improve the Performance of Ethanol Production Using Lignocellulosic Hydrolysate.

Increased consumption of xylose-glucose and yeast tolerance to lignocellulosic hydrolysate are the keys to the success of second-generation bioethanol production. Candida tropicalis KBKTI 10.5.1 is a new isolated strain that has the ability to ferment xylose. In contrast to Saccharomyces cerevisiae DBY1 which only can produce ethanol from glucose fermentation. The research objective is the application of the genome shuffling method to increase the performance of ethanol production using lignocellulosic hydrolysate. Mutants were selected on xylose and glucose substrates separately and using random amplified polymorphic DNA (RAPD) analysis. The ethanol production using lignocellulosic hydrolysate by parents and mutants was evaluated using a batch fermentation system. Concentrations of ethanol, residual sugars, and by-products such as glycerol, lactate and acetate were measured using HPLC machine equipped with Hiplex H for carbohydrate column and a refraction index detector (RID). Ethanol produced by Fcs1 and Fcs4 mutants on acid hydrolysate increased by 26.58% and 24.17% from parent DBY1, by 14.94% and 21.84% from parent KBKTI 10.5.1. In contrast to the increase in ethanol production on alkaline hydrolysate, Fcs1 and Fcs4 mutants only experienced an increase in ethanol production by 1.35% from the parent KBKTI 10.5.1. Ethanol productivity by Fcs1 and Fcs4 mutants on acid hydrolysate reached 0.042 g/L/h and 0.044 g/L/h. The recombination of the genomes of different yeast species resulted in novel yeast strains that improved resistance performance and ethanol production on lignocellulosic hydrolysates.

Bioprospecting and Diversity of Yeast Producing Ethanol Isolated from Indonesia.

Bioethanol is considered the most environmentally friendly as renewable fuels. Indonesia has abundant microbe diversity which is potential for bioprospecting such as fermenting agents using agriculture product as raw materials for producing bioethanol. This study aims to isolate, characterise and molecular identify of 15 isolates of bioethanol-producing yeasts from various sources. Characterisation based on ethanol production, cell morphology and various substrate utilisation has been carried out. Molecular characterisation of 15 yeast isolates using tree sets of primers had been carried out. Amplification in the internal area of transcribe spacers (ITS) was successfully carried out with an amplitude of 400 bp-900 bp. Amplifiers in the D1/D2 26s rDNA domain are 250 bp. Amplification with ScerF2 and ScerR2 specific primers was carried out successfully and proved that there were two isolates which were not Saccharomyces cerevisiae analysis of yeast genetic diversity showed 12 yeast isolates classified as S. cerevisiae and the rest belonged to the genus Clavispora, Candida and Kodamaea (Pichia).

Soursop leaves (Annona muricata L.) endophytic fungi anticancer activity against HeLa cells.

Cervical cancer causes many deaths in females worldwide, including in Indonesia. Several studies have reported that soursop (Annona muricata L.) leaves can be used to treat cervical cancer. This study aims to determine the use of endophytic fungi of A. muricata leaves extract as an ingredient that inhibits cervical cancer. The isolated endophytic fungi from various soursop leave accessions were grown in culture media, then extracted using ethyl acetate. The extract was then tested against anti-yeast, cervical cancer cells, and on normal cells as control using the MTT method. Five isolated fungi were selected based on the greatest inhibition in one concentration, and the inhibitory concentration 50 (IC50) value was determined. The soursop leaves endophytic fungi extracts showed cytotoxicity against cervical cancer cells by inhibiting the multiplication of HeLa cancer cells in vitro. The Sir-SM2 endophytic fungi crude ethyl acetate extract showed high cytotoxicity to cervical cancer cells (HeLa cells) but less harmful to the normal Chang cells; therefore can be a natural anticancer. Identification based on morphology shows that the isolated Sir-SM2 endophytic fungi belong to the Penicillium genus, and molecular identification based on Internal Transcribed Spacer shows high similarities with Penicillium crustosum.

Anticancer activity test of ethyl acetate extract of endophytic fungi isolated from soursop leaf (Annona muricata L.).

OBJECTIVE: To analyze anticancer activity of an ethyl acetate extract of endophytic fungi isolated from soursop leaf (Annona muricata L.). METHODS: Anticancer activity of fungal extracts was determined by observing its toxicity against MCF-7 (Michigan Cancer Foundation-7) cells in vitro by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay method. At an extract concentration of 100 µg/mL, 4 isolates out of 12 showed high activity against the cancer cell growth. The four isolates were then selected for further IC50 determination, by measuring the inhibition of cancer cell proliferation at extract concentration of 25 µg/mL, 50 µg/mL, 100 µg/mL, 200 µg/mL and 400 µg/mL. RESULTS: Results showed that isolate Sir-G5 had the highest anticancer activity with an IC50 of 19.20 µg/mL. The best isolates were screened again using a normal cell (Chang cells) to determine its toxicity against normal cells. Results indicated that the extracts do not affect the proliferation of normal cells. Molecular identification showed that the fungal isolate Sir-G5 has a close relationship with Phomopsis sp. CONCLUSIONS: The endophytic fungi isolated from soursop leaf has the potential to be used as a source of anticancer agents.

Challenges of non-flocculating Saccharomyces cerevisiae haploid strain against inhibitory chemical complex for ethanol production.

This study provides insight observation based on the gene expression and the metabolomic analysis of the natural robust yeast Saccharomyces cerevisiae NBRC849 during the fermentation in the medium containing inhibitory chemical complexes (ICC) at different concentrations. The tolerance mechanisms involved in the strain might have existed through the upregulation of genes involved in NAD(H)/NADP(H) cofactors generations (ALD6, ZWF1, GND1), membrane robustness for efflux pump (YOR1, PDR5, TPO3) and cation/polyamine transport (TPO3). The alteration of metabolic flux to the shikimic pathway was also found in this strain, resulted in the enhanced formation of aromatic amino acid required for cell survival. Enhanced expression of these genes as well as the increase of metabolic flux to shikimic pathway were suggested to result in the robustness of non-flocculating S. cerevisiae haploid strain.