Mon, 18 Dec 2017

A Sensitive UPLC Method Development and Validation with LC-MS Compatible for the Determination of 1-Deoxynojirimycin In Mulberry Leaves using Fluorescence Detection

J Rudraprasad Reddy1*, R Pramod Kumar2, K M Ch Appa rao1, K Ramakrishna1

1 Department of Chemistry, Gitam Institute of Science, GITAM University, Visakhapatnam-530 045, Andhra Pradesh, India

2 Department of Chemistry, JNT University, Kakinada- 533 003, Andhra Pradesh, India


ABSTRACT

A simple, rapid, precise LC-MS compatible method for the degradation impurities if any UHPLC method was developed for the determination of 1-deoxynojirimycin (DNJ) in Mulberry leaves. The DNJ in 200 mg dried leaves was extracted twice with 50 mL aqueous 0.05 mol/L HCl, then derivatized by 9 fluorenylmethyl chloroformate (9 FMOC-Cl) in Potassium Borate buffer, and analyzed using a high performance liquid chromatograph equipped with a fluorescence detector. Chromatographic separation of the same was performed by using a Shimpack XR ODS-III (150*3.0mm) 2.1µm as stationary phase with a mobile phase of 0.1% Acetic acid in water: Acetonitrile (60: 40, v/v) at the rate of 0.5 mL/min. Wavelengths used were 254 nm for excitation and 322 nm for emission, which were suitable for detecting the native fluorescence of all the pigments assayed. The derivatized DNJ was well dissolved from the hydrolysis products of 9 FMOC-Cl. The linearity ranged from 0.45 to 66 mg/L, and the detection limit was 0.2 mg/L (S/N = 3). The content of DNJ in Mulberry leaves was 0.09%, the recovery was 90.0%-110.0%.

Keywords:  UPLC, Method development, Method Validation, 1-deoxynojirimycin and Mulberry leave


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