The utilization of methanolic Bixa orellana (Annatto) seed extract as substitute for safranin in Gram staining

SAM DANIEL T. VENTURINA, JOSH ZAVION A. COMUELO, WAYNE ROCQ A. SAMANIEGO, and FERNANDO CHRISTIAN JOLITO III

Philippine Science High School – Western Visayas Campus, Brgy. Bito-on, Jaro, Iloilo City 5000, Department of Science and Technology, Philippines

Abstract

The routine use of safranin, a synthetic dye, in Gram staining raises a number of environmental and safety concerns. This study evaluates the utility of the methanolic extract of Bixa orellana (Annatto) seeds as replacement for safranin as counterstain in Gram staining. Pure and mixed bacterial smears of E.coli and S.aureus were stained with Annatto and safranin. The staining showed that the Bixa orellana extract did not stain the bacteria in any of the slides. Results show that the pH of the extract was found to be acidic at 5.92 in comparison to safranin, which was basic at 7.44. The inability of the extract to stain the bacteria may be attributed to its acidic pH which makes it unable to bind to the acidic cell wall of the bacteria.

Keywords: annatto, bixin, safranin, Gram staining, methanol