Bioprospecting for Biopolymer-Producing Algae from Various Yemeni Habitats: Isolation, Characterization, Chemical Staining Screening, and FTIR Analysis

Microalgae are promising biological platforms for the sustainable production of biodegradable polymers, particularly polyhydroxybutyrate (PHB), a member of the polyhydroxyalkanoate (PHA) family. The environmental impacts of conventional plastics underscore the need for rapid, reliable screening methods to identify microalgal strains that accumulate PHB. Staining-based methods offer a cost-effective approach for the preliminary detection of intracellular polymer inclusions. This study, conducted in Yemen, compared three staining techniques—Nile Red, Nile Blue A, and Sudan Black B—for screening PHB accumulation in microalgal isolates. Nile Blue A was used to visualize putative PHB granules by fluorescence microscopy, Nile Red to detect intracellular hydrophobic inclusions, and Sudan Black B to provide a qualitative assessment by light microscopy. The findings indicate that the highest positive-reading rate was 100% for Nile Blue A dissolved in dimethyl sulfoxide (DMSO), followed by 98.20% for Nile Red in DMSO, 92.79% for Nile Blue A in acetone, and 88.29% for Nile Red in acetone. Sudan Black B in 70% ethanol yielded the lowest positive-reading rate (40.54%), with positive inclusions appearing blue-black. Negative reactions were characterized by pink or red cells or granules (Appendix A). Of the 37 samples examined in triplicate, 32 were classified as positive for intracellular biopolymer accumulation (86.49%). These samples may warrant further investigation for biotechnological applications. FTIR spectra of extracts from selected algal isolates showed absorption bands consistent with PHB, including an ester carbonyl (C=O) stretching band at approximately 1720–1724 cm⁻¹. These spectral features support the preliminary identification of PHB; additional confirmatory analyses are needed to establish polymer identity conclusively.