The Simultaneous HPLC Determination of Aflatoxins G2, G1, B2, B1, Zearalenone and
Ochratoxin A by Post-
by Henry Joshua
Aura Industries Inc., 545 Eighth Ave., Suite 5W, New York, NY 10018
Phone: 212-290-9190, e-mail: info@aura-inc.com, Web site: aura-inc.com
Aura Industries Inc., 545 Eighth Ave., Suite 5W, New York, NY 10018
Phone: 212-
Aflatoxins G2, G1, B2 and B1, can conveniently be detected by HPLC and fluorescence detection using post-column photochemical derivatization (1, 2, 3). This demonstration extends the analysis of the standard aflatoxins to include the mycotoxins zearalenone and ochratoxin A. To this end a gradient elution is required and the excitation wavelength of the fluorescence detector is changed to 315 nm.
Apparatus, chemicals and chromatographic conditions
HPLC Apparatus:
An HPLC apparatus with a fluorescence detector, injector, gradient and data handling capability is required. The fluorescence detector settings: excitation 315 nm, emission >415 nm.
An HPLC apparatus with a fluorescence detector, injector, gradient and data handling capability is required. The fluorescence detector settings: excitation 315 nm, emission >415 nm.
GRADIENT COMPOSITION
|
Time (min.) |
0.01 M H3PO4 % |
Acetonitrile % |
Methanol % |
|
0 |
60 |
20 |
20 |
|
7 |
60 |
20 |
20 |
|
10 |
40 |
30 |
30 |
|
20 |
40 |
30 |
30 |
|
21 |
60 |
20 |
20 |
The photochemical reactor is inserted between the column exit and the detector inlet. The specific knitted reactor used for this determination is the KRC 25-25 (Aura Industries Inc.) consisting of 25 meter of PTFE 1/16 inch OD x 0.25 mm ID tubing.
Column:
A Supelcosil LC-18, 15 cm x 4.7 mm ID. The column was kept at 40°C.
A Supelcosil LC-
Chemicals:
The eluent consists of HPLC grade methanol, acetonitrile and water and is pumped at 1 ml/min.-
The eluent consists of HPLC grade methanol, acetonitrile and water and is pumped at 1 ml/min.-
Standards: Aflatoxin Mix Kit-M (Supelco) containing 1µg B1, 1µg G1, 0.3 µg B2 and 0.3 µg G2 in one ml of methanol in each ampul. The contents of one ampul was diluted with 4.20 ml water and 1.47 ml acetonitrile. Zearalenone and Ochratoxin A were added to the aflatoxin mix so that the concentrations of the analytes in the injection solution were: Aflatoxins B1, and G1, 150 ng per ml, B2 and G2, 45 ng per ml, Zearalenone, 1000 ng per ml and Ochratoxin A, 50 ng per ml.- Injection size: 20 µl.
References:
1) Henry Joshua, Determination of Aflatoxins by Reversed-Phase High Performance Liquid Chromatography with Post-Column In-Line Photochemical Derivatization and Fluorescence Detection, Journal of Chromatography, 654, 247-254 (1993).
1) Henry Joshua, Determination of Aflatoxins by Reversed-
2) H. Joshua, Analysis of Aflatoxins in Naturally Contaminated Corn by HPLC, Post-Column Photochemical Derivatization and Fluorescence Detection, Pittsburgh Conference and Exhibition, Chicago, IL, March 1994. Published in American Laboratory, p 36J-36M, April 1995.
3) B. W. Horn, R. L. Greene, V. S. Sobolev, J. W. Dorner, J. H. Powell and R. C. Layton, Association of morphology and mycotoxin production with vegetative compatibility groups in Asperagillus flavus, A. parasiticus and A. tamarii, Mycologia 88, 574-587 (1996).
* Presented at the Satellite Workshop to the Gordon Conference on Mycotoxins and Phycotoxins "Advances in Detection Methods for Fungal and Algal Toxins." June 17-19, 1999.
For a Bibliography of selected review and application articles related to Aura Industries’ Photochemical Reactor for Enhanced Detection PHRED™ please click here.
For more information about the PHRED™ please click here.
Aura Industries, Inc. • Email: info@aura-inc.com
Ph: 212-290-9190 • Fax: 212-290-9191
8895 Towne Centre Dr Ste 105-330 • San Diego, CA 92122 USA