Single step aqueous two-phase extraction for downstream processing of C-phycocyanin from Spirulina platensis.

C-phycocyanin, a natural food colorant, is gaining importance worldwide due to its several medical and pharmaceutical applications. In the present study, aqueous two-phase extraction was shown to be an attractive alternative for the downstream processing of C-phycocyanin from Spirulina platensis. By employing differential partitioning, C-phycocyanin selectively partitioned to the polymer rich (top) phase in concentrated form and contaminant proteins to the salt rich (bottom) phase. This resulted in an increase in the product purity (without losing much of the yield) in a single step without the need of multiple processing steps. Effect of process parameters such as molecular weight, tie line length, phase volume ratio, concentration of phase components on the partitioning behavior of C-phycocyanin was studied. The results were explained based on relative free volume of the phase systems. C-phycocyanin with a purity of 4.32 and yield of about 79 % was obtained at the standardized conditions.

Identification of Taxol-producing endophytic fungi isolated from Salacia oblonga through genomic mining approach.

The most promising anti-tumor agent developed in the past three decades is Taxol. It is proven to be effective against many cancers. It is necessary to isolate pharmacologically potent endophytic microbial strains from medicinal plants with special reference to Taxol production. In the current study, endophytic fungi were isolated from the bark of the medicinal plant, Salacia oblonga. The isolated endophytes were identified morphologically, and further characterized by ITS-PCR using genomic DNA samples, later the products were sequenced for identification and phylogenetic linkage mapping. The samples were screened for the potential to produce Taxol or taxanes, employing PCR. The resulted data have been sequenced to confirm the presence of the two genes implicated in Taxol biosynthesis, 10-deacetylbaccatin III-10-O-acetyl transferase (DBAT) and C-13 phenylpropanoid side chain-CoA acyltransferase (BAPT). Seven samples showed the amplicons of DBAT gene and one showed the amplicons of BAPT gene. Sequencing of these products was carried out, of which one sample has revealed sequence homology to the original DBAT gene from Taxus. The present work confirms and substantiates the potential of genomic mining approach to discover novel Taxol-producing endophytic fungi.

Extraction and purification of Ipomoea peroxidase employing three-phase partitioning.

Three-phase partitioning (TPP) is a novel separation process used for the extraction and purification of biomolecules. The biomolecules are recovered in a purified form at the interface (precipitate), while the contaminants partition in t-butanol and aqueous phases. Peroxidase from the leaves of Ipomoea palmata was purified by using TPP. The ratio of the crude extract to t-butanol of 1:1 and 30% ammonium sulfate at 37 degrees C resulted in about 160% activity recovery and twofold purification in the aqueous phase of the first cycle of TPP. On subjecting the aqueous phase to the second cycle of TPP, a purification of 18-fold was achieved with about 81% activity recovery. The sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis showed substantial purification, and the molecular weight of peroxidase was found to be 20.1 KDa. The present study shows a higher degree of purification and activity yield as a primary purification process in comparison with existing literature values, thus demonstrating TPP as an attractive downstream process for the purification of peroxidase.

Fractionation and purification of the phycobiliproteins from Spirulina platensis.

C-Phycocyanin and allophycocyanin of Spirulina platensis are fractionated and purified using a non-chromatographic method namely, aqueous two phase extraction for the first time. Optimized process parameters of aqueous two phase extraction (PEG 4000/potassium phosphate of tie line length 18.64% with a phase volume ratio 1.45) resulted in pure C-phycocyanin and allophycocyanin with a purity of 3.23 and 0.74, respectively, in a single extraction. Multiple extractions (two) improved the purity of C-phycocyanin from 3.23 to 4.02. Integration of aqueous two phase extraction with membrane process not only facilitated the separation of phase forming components from the products and also increased the purity of allophycocyanin from 0.74 to 1.5.