QUANTIFICATION OF ETHYL CARBAMATE IN TOBACCO POSTER-15 Dinesh.T.K., Sathya Gourishankar and Sharad.K.Mehta* ITC Ltd, ITC R&D Centre, Peenya, Bangalore – 560 058, INDIA.

ABSTRACT SAMPLE PREPARATION SAMPLE CHROMATOGRAM REPRODUCIBILITY ETHYL 2.5g Sample CARBAMATE ANALYST TRIAL NO. SAMPLE (ng/g) 2012_TSRC15_Mehta.pdf A simple method is developed for quantification of Ethyl 1 Tobacco 371.0 carbamate in Tobacco. Ethyl carbamate in Tobacco is 2 Tobacco 368.7 extracted with water and is partitioned with Sonicate with 25ml water for 30 mins and 3 Tobacco 382.2 dichloromethane. The dichloromethane extract is Centrifuge at 10000 rpm for 10 minutes. 4 Tobacco 374.1 concentrated and injected in GC/MS-SIM mode. The 1 5 Tobacco 384.1 method has been validated using standard validation 6 Tobacco 389.0 protocols and there is excellent linearity over 7 Tobacco 368.3 concentration range from 100 to 4000 ng/g with Extract 10 ml of water extract with 30ml correlation coefficient of 0.9939. The recoveries are more dichloromethane twice. 8 Tobacco 367.1 than 90% for Ethyl carbamate with limit of quantification 9 Tobacco 364.5

100 ng/g and limit of detection 40ng/g respectively. carbamate Ethyl 2 10 Tobacco 372.2

Concentrate the dichloromethane extract STDEV 8.2 after passing through sodium sulphate and MEAN 374.1 inject in GC-MS. %RSD 2.2

INTRODUCTION GC-MS CONDITIONS LINEARITY RECOVERY STUDIES CALIBRATION FOR ETHYL CARBAMATE 400000 Ethyl carbamate (Urethane) is a genotoxic carcinogen found 350000 in fermented foods and beverages. Ethyl carbamate in y = 340.35x - 5869.2 SPIKED OBTAINED Tobacco is formed from Maleic hydrazide, which is a plant • Column : Zebron-ZB Wax plus 30mX0.25mmX0.25m. R² = 0.9939 TRIAL AMOUNT AMOUNT % 300000 growth regulator applied to Tobacco. Ethyl carbamate is the • Column temperature : 80°C(2 mins.) 2°C per min. to NO. SAMPLE (ng/g) (ng/g) RECOVERY 130°C(2 mins.) 20°C per min. R ethyl ester of carbamic acid. It is also formed from E 250000 hydrogen cyanide and ethanol or urea and ethanol. Although to 220°C(3 mins.). S P there are a number of analytical methods reported in the • Injector temperature: 200°C. 200000 O 1 Tobacco 403 459 114 literature, they all have two major drawbacks, namely, (1) • Transfer Line Temperature: 210°C. N Requires sophisticated instruments like UPLC/MS/MS and • Injection volume: 1.0 µl. S 150000 E (2) Need laborious clean up procedure to remove the matrix • Scan Range: 50 – 550 amu. effect of Tobacco. The aim of the method is to develop a • Quantification Ions: 62, 74 & 89. 100000 2 Tobacco 403 449 111 simple method with less clean up to remove matrix and use 50000 routine instrument like GCMS to quantify Ethyl carbamate in Tobacco. 0 0 200 400 600 800 1000 1200 CONC. IN PPM 3 Tobacco 403 389 96.5

LITERATURE STANDARD CHROMATOGRAM REPEATABILITY CONCLUSIONS METHOD REFERENCES LIMITATIONS ETHYL • Laborious extraction A Formation and Determination of CARBAMATE procedure. Ethyl carbamate in Tobacco and TRIAL NO. SAMPLE (ng/g) Tobacco Smoke, Schmeltz, Chiong • Matrix interference.

and Hoffmann. J Analytical Toxicology 2(1978) 265-268. 1 Tobacco 371.0 • A simple method for quantification B Ethyl carbamate levels in US and • Sophisticated Swedish smokeless tobacco products. instrumentation 2 Tobacco 368.7 of Ethyl carbamate in Tobacco is developed. Faizi A., Kimpton H., McAdam K., (LC-MS/MS). BAT, Group Research and Development, Southampton, UK. 3 Tobacco 382.2 • Recovery of greater than 95%. University of Louisville, Louisville, USA. SSPTPOST14, CORESTA 4 Tobacco 374.0 Conference, Edinburgh September 2010. 5 Tobacco 384.1 • Method for stone-fruit. C Application of tandem mass spectrometry combined with gas chromatography to the routine STDEV 6.8

analysis of ethyl carbamate in stone- Ethyl carbamate Ethyl fruit spirits. Dirik W.Lachenmeier*, MEAN 376.0 Willi Frank and Thomas Kuballa. Rapid Commun.Mass Spectrom.2005; 19: 108-112. %RSD 1.8 TSRC2012(66) - Document not peer-reviewed