Validated HPAEC-PAD Method for the Determination of Fully Deacetylated Chitooligosaccharides.

Lidong Cao,Xiuhuan Li,Jinlong Wu,Pingping Liu,Qiliang Huang,Miaomiao Wu,Li Zheng

doi:10.3390/ijms17101699

Abstract

An efficient and sensitive analytical method based on high-performance anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD) was established for the simultaneous separation and determination of glucosamine (GlcN)1 and chitooligosaccharides (COS) ranging from (GlcN)2 to (GlcN)6 without prior derivatization. Detection limits were 0.003 to 0.016 mg/L (corresponding to 0.4–0.6 pmol), and the linear range was 0.2 to 10 mg/L. The optimized analysis was carried out on a CarboPac-PA100 analytical column (4 × 250 mm) using isocratic elution with 0.2 M aqueous sodium hydroxide-water mixture (10:90, v/v) as the mobile phase at a 0.4 mL/min flow rate. Regression equations revealed a good linear relationship (R2 = 0.9979–0.9995, n = 7) within the test ranges. Quality parameters, including precision and accuracy, were fully validated and found to be satisfactory. The fully validated HPAEC-PAD method was readily applied for the quantification of (GlcN)1–6 in a commercial COS technical concentrate. The established method was also used to monitor the acid hydrolysis of a COS technical concentrate to ensure optimization of reaction conditions and minimization of (GlcN)1 degradation.

Highlights

Chitin is the second most abundant naturally occurring homopolysaccharide extracted from the exoskeleton of crustaceans and insects, and from fungal cell walls, etc
Good separation depends primarily on the column, mobile phase composition, and flow rate. These three variables were systematically screened during optimization of chromatographic conditions, which was carried out using mixed (GlcN)1–6 standard solutions
The coefficient of variations of seven replicate determinations of the same batch of COS technical concentrate are compared with the Horwitz value (%RSDr) [46]

Summary

Introduction

Chitin is the second most abundant naturally occurring homopolysaccharide extracted from the exoskeleton of crustaceans and insects, and from fungal cell walls, etc. [1]. Chitin is the second most abundant naturally occurring homopolysaccharide extracted from the exoskeleton of crustaceans and insects, and from fungal cell walls, etc. Chitosan is known to have attractive properties and has been used in many fields, such as agriculture, food, cosmetics, and medicine [2]. The study on chitosan has attracted interest in converting it into more soluble chitooligosaccharides (COS). These hydrolyzed oligomers of chitin and chitosan exhibit remarkable biological activities, such as antibacterial, antifungal, antioxidant, antitumor, and immunostimulant properties [3,4]. COS have been shown to induce various plant defense-related cellular responses and are widely used in agriculture [4]

Methods

Results

Conclusion