Synthesis of allitol from D-psicose using ribitol dehydrogenase and formate dehydrogenase

Hinawi A.M Hassanin,Wanmeng Mu,Ammar Elferga,Rebaone Letsididi,Marwa Y.F Koko,Bo Jiang

doi:10.4314/tjpr.v15i12.23

Abstract

Purpose: To synthesize allitol from D-psicose by a combination of novel ribitol dehydrogenase (RDH) and formate dehydrogenase (FDH) under optimised production conditions. Methods: RDH and FDH genes were cloned and introduced into pET-22b(+) vectors for expression in Escherichia coli to produce the corresponding enzymes. The effects of temperature, pH, shaking velocity (75, 100, 125, and 150 rpm), and shaking type (horizontal and vortex) were optimised to maximise the production yield of allitol. The final product was purified and subjected to nuclear magnetic resonance (NMR) spectroscopy, infrared (IR) spectrometry, and liquid chromatography-mass spectrometry (LC-MS) to confirm its structure. Results: The optimal pH and temperature for the reaction were 7.5 and 40 °C, respectively. The results revealed that allitol yield significantly increased with increase in reaction shaking velocity and reached a maximum yield of 95.60 ± 0.54 % at 150 rpm shaking velocity after 6 h of reaction. When the reaction was run under horizontal shaking, allitol yield increased from 100.00 ± 6.05 (without shaking) to 124.20 ± 9.70 %. Twenty milligrams of D-psicose were successfully reduced to allitol under optimum conditions with a high production yield of 16.7 ± 0.62 mg after 6 h. No by-products were formed during or after the reaction. The produced allitol had a purity of 95 %, and its structure was confirmed by HPLC, IR, LCMS, and NMR spectral analyses. Conclusion: Using D-psicose as a substrate, allitol with 95 % purity was successfully produced by the combination of novel RDH and FDH. Keywords: Allitol, Ribitol dehydrogenase, Formate dehydrogenase, D-Psicose, Providencia alcalifaciens

Highlights

Rare sugars are monosaccharides that are less abundant in nature than common sugars such as D-fructose and D-glucose
Previous studies confirmed the validity of this approach; for example, L-xylulose, D-tagatose, D-sorbose, Dpsicose, D-talitol, and allitol were successfully produced using microorganisms [7,8,9,10,11]
We report for the first time the use of new ribitol dehydrogenase (RDH) from P. alcalifaciens RIMD 1656011 and formate dehydrogenase (FDH) from the methanolassimilating yeast O. parapolymorpha DL-1 for the production of allitol

Summary

Introduction

Rare sugars are monosaccharides that are less abundant in nature than common sugars such as D-fructose and D-glucose. Their rarity limits their large-scale extraction and separation from natural sources. These compounds, which are classified as ‘sugars’ and ‘sugar alcohols,’ have interesting characteristics, and are used in both food production and pharmaceuticals [1,2,3,4,5,6]. Izumori [10] reported a useful strategy for processing most hexoses, called the Izumoring map, in which allitol and galactitol are placed at symmetric points of D and L in the production zone of hexose sugars

Results

Discussion

Conclusion