# Characterization of Novel Pullulanase Type I from Newly Isolated Bacillus cereus Strain NP9: Potential Additive for Laundry Detergent Formulations

## TL;DR

A new pullulanase from Bacillus cereus NP9 was characterized for its potential use in laundry detergents due to its stability and effectiveness in removing starchy stains.

## Contribution

The study identifies and characterizes a novel pullulanase type I from Bacillus cereus NP9 with detergent-compatible properties.

## Key findings

- NP9 pullulanase is stable across a wide pH and temperature range, making it suitable for detergent use.
- The enzyme effectively converts pullulan and starch into maltotriose, confirming it as a type I pullulanase.
- NP9 pullulanase improved starchy stain removal when combined with commercial detergent.

## Abstract

This study aimed to produce a pullulanase that can be utilized as an additive in detergent formulations. A newly isolated Bacillus cereus strain NP9 exhibited the highest pullulanase activity and was selected for production. The optimum conditions for crude NP9 pullulanase were a pH of 7.0 and a temperature of 40 °C. It maintained stability at high rates within the pH range of 5.0 to 11.0 and temperatures between 25 and 65 °C. The molecular weight of the enzyme was determined to be approximately 170 kDa via native-PAGE. Thin-layer chromatography and high-performance liquid chromatography analyses indicated that NP9 pullulanase converted pullulan and starch substrates into maltotriose units (pullulanase type I). The enzyme exhibited moderate activity with certain metal ions and was not Ca2+-dependent. The inhibition of the enzyme by EDTA, EGTA, and 1,10-phenanthroline indicated it is a metalloenzyme. The enzyme moderately retained activity when exposed to non-ionic detergents such as Triton X-100, Tween 20, and Tween 80. It demonstrated high compatibility (90%) with the commercial detergent “Peros.” Wash performance analyses showed that the NP9 pullulanase and commercial detergent mixture removed starchy stains more effectively than washing with commercial detergent alone. In conclusion, NP9 pullulanase exhibited favorable properties, making it a potential candidate for the laundry detergent industry.