THE POTENTIAL OF ATMOSPHERIC SOLID ANALYSIS PROBE (ASAP) IONISATION MASS SPECTROMETRY FOR RAPID IDENTIFICATION OF ANABOLIC STEROID ESTERS Mickael DOUE, Gaud DERVILLY-PINEL, Audrey GICQUIAU, Laure BEUCHER, Fabrice MONTEAU and Bruno LE BIZEC
LABoratoire d’Étude des Résidus et Contaminants dans les Aliments (LABERCA) USC INRA 1329, Oniris, LUNAM Université, BP 50707, 44307 Nantes Cedex 3, France Fax : +33 (0)2 40 68 78 78 - Tél : +33 (0)2 40 68 78 80 [email protected]
NATURAL HORMONES ABUSE IN MEAT PRODUCING ANIMALS INTRODUCTION
The use of anabolic substances for livestock fattening has been prohibited in the European Union (Directive 96/22/EC). Nevertheless, their uses are still suspected regarding potential benefits in terms of income. Anabolic steroid esters are the most commonly used due to their higher bioavailability and their longer lasting effect. In this context, the present study aims at providing a new rapid, efficient and multiresidue confirmatory strategy based on Atmospheric Solids Analysis Probe (ASAP) technology.
The potential of ASAP as new, efficient and rapid confirmatory strategy was assessed. Investigation of ionization and fragmentation both in positive and negative mode for 23 steroid esters was carried out. Rapid method to identify large set of steroid esters was developed. Isotopic dilution method in Selected Reaction Monitoring (SRM) mode was validated for the rapid quantification of these compounds.
ATMOSPHERIC SOLIDS ANALYSIS PROBE (ASAP) TARGETED STEROID ESTERS CH3 OR
PRINCIPLE CH3 OR
6 Testosterone Esters
8 Estradiol Esters
. + M
4 Boldenone Esters
Adapted from Nielen et al., TRAC , 30 (2011) 165
5 Nandrolone Esters
Ambient Mass Spectrometry
R : From C2 to C10 including cyclic esters
MS parameters Mode Polarity Corona Cone Source offset
WATERS XEVO TQS®
Values Current Positive 3 µA 45 V 70 V
MS parameters Probe Temperature Gas Desolvation Flow Cone Nebuliser
MS Optimized Tune Conditions O
RAPID IDENTIFICATION AND QUANTIFICATION OF ANABOLIC STEROID ESTERS
ASAP-MS/MS SCAN of Testosterone Enanthate ASAP POSITIF
Observed m/z: 401.3 [M+H]+ 289.2 [T+H]+ 271.2 [T+H-H2O]+ 253.2 [T+H-2H2O]+
253.2 271.2 289.2
NEUTRAL LOSS SCAN 96.1
271.2 > 175.1 TESTOSTERONE
269.2 > 173.1 BOLDENONE
257.2 > 161.1 NANDROLONE
255.2 > 159.1 ESTRADIOL STEROID IDENTIFICATION
PRECURSOR ION 271.2 m/z ASAP NEGATIF
Values 450 °C 500 L.H-1 150 L.H-1 4 bars
PRECURSOR ION 269.2 m/z
PRECURSOR ION 257.2 m/z
PRECURSOR ION 255.2 m/z
129.1 O H3C
[M+H]+ Observed m/z: 399.3 [M-H]269.2 [T-H-H2O]129.1 [EN]-
PRODUCT ION OF THE CORRESPONDING [M-H]-
59.1 => Acetate 121.1 => Benzoate 149.1 => Phenyl Propionate
VALIDATION – ASAP SRM POSITIVE MODE Labeled internal standard + Steroid Ester
RSD (1 ppm)
< 20 %
73.1 => Propionate 129.1 => Enanthate 171.1 => Decanoate 199.1 => Laurate
101.1 = > Valerate 141.1 => Cypionate 183.1 => Undecylenate ESTER IDENTIFICATION
RAPID IDENTIFICATION OF ANABOLIC STEROID ESTERS (3 min)
CONCLUSION TAKE HOME MESSAGE
Rapid, efficient and multiresidue method was developed in order to identify a large set of anabolic steroid esters in less than 5 minutes.
Extend the developed procedure to other anabolic steroid esters.
Thanks to labeled internal standards, quantification of the identified steroid esters was realized in oily commercial solution.
Develop an algorithm to identify both known and also unknown anabolic steroid esters.