Analysis of Natural Toxins by Liquid Chromatography-Chemiluminescence Nitrogen Detection and Application to the Preparation of Certified Reference Materials

Abstract

The implementation of instrumental analytical methods such as LC-MS for routine monitoring of toxins requires the availability of accurate calibration standards. This is a challenge because many toxins are rare, expensive, dangerous to handle, and/or unstable, and simple gravimetric procedures are not reliable for establishing accurate concentrations in solution. NMR has served as one method of qualitative and quantitative characterization of toxin calibration solution Certified Reference Materials (CRMs). LC with chemiluminescence N detection (LC-CLND) was selected as a complementary method for comprehensive characterization of CRMs because it provides a molar response to N. Here we report on our investigation of LC-CLND as a method suitable for quantitative analysis of nitrogenous toxins. It was demonstrated that a wide range of toxins could be analyzed quantitatively by LC-CLND. Furthermore, equimolar responses among diverse structures were established and it was shown that a single high-purity standard such as caffeine could be used for instrument calibration. The limit of detection was approximately 0.6 ng N. Measurement of several of Canada's National Research Council toxin CRMs with caffeine as the calibrant showed precision averaging 2% RSD and accuracy ranging from 97 to 102%. Application of LC-CLND to the production of calibration solution CRMs and the establishment of traceability of measurement results are presented.

Isolation and Characterization of [D-Leu1]microcystin-LY from Microcystis aeruginosa CPCC-464

“…The LC-MS/MS and LC-UV results are therefore entirely consistent with 1 being [D-Leu 1 ]MC-LY.A portion of the purified 1 was used to prepare a stock solution. This was quantitated using qNMR [18] and LC with chemiluminescence nitrogen detection (CLND) [19], then accurately diluted with 1:1 MeOH-H 2 O to prepare a reference material (RM) (~7.7 µM). LC-UV analysis of this RM showed the relative concentration of [D-Leu 1 ,D-Glu(OMe) 6 ]MC-LY to be 3.1%.…”

“…This stock solution was quantitated directly by 1 H NMR using high purity caffeine as the external calibrant as described previously [18]. A dilution of the stock solution was prepared with 50% MeOH-H 2 O for analysis by LC-UV-CLND [19] using an Agilent 1100 HPLC system with a 1050 UV detector connected to a model 8060 CLND (Antek PAC, Houston, TX, USA). Separations were performed on an Agilent 3.5 µm Poroshell SB-C8 (2.1 × 150 mm) maintained at 40 • C. Isocratic elution was at 0.2 mL/min, using 65% MeOH-H 2 O (0.2% HCOOH) for 1.…”

Rapid quantitative screening of cyanobacteria for production of anatoxins using direct analysis in real time high‐resolution mass spectrometry

“…A calibration solution for hATX was prepared using toxin isolated in‐house from Oscillatoria sp. strain PCC 6407 and quantitated by 1 H nuclear magnetic resonance spectroscopy and LC chemiluminescence nitrogen detection, as described previously 37–39 …”

Disposable Electrochemical Aptasensor Based on Graphene Oxide-DNA Complex as Signal Amplifier towards Ultrasensitive Detection of Ochratoxin A

“…Currently, the most commonly used methods are the enzyme-linked immunosorbent assay (ELISA) and high-performance liquid chromatography (HPLC) [5,6], which possess high sensitivity and accuracy. However, they require expensive reagents or equipment, and highly trained operators, limiting their wide application in point-of-care (POC) testing, especially in less developed or remote areas.…”