1. Nitrosamines and Plant Nutrition Dr. Gary Palmer University of Kentucky

2. What are Nitrosamine? Nitrogen containing compounds that are know to have a potential affect on cancer Nitrogen containing compounds that are know to have a potential affect on cancer Those occurring in tobacco are specific to tobacco Those occurring in tobacco are specific to tobacco TSNA ( Tobacco Specific NitrosAmines ) TSNA ( Tobacco Specific NitrosAmines ) Occur during curing Occur during curing Increase during storage of cured leaf Increase during storage of cured leaf

3. Major Factors that Influence TSNA Accumulation Curing Conditions Curing Conditions Related to rate of curing and moisture content Related to rate of curing and moisture content Storage Storage Related to moisture content and nitrogenous compounds Related to moisture content and nitrogenous compounds Nitrogen Fertilization Nitrogen Fertilization Rate? Rate? Source? Source? Varietal Differences Varietal Differences Nornicotine Nornicotine Nornicotine can Nornicotine can

4. Effect of 250 lb/a Nitrogen Fertilizer on Nicotine, Nitrate, Nitrite and TSNA Accumulation in Air-Cured Tobacco Lamina Fertilizer treatment nicotineNO3-NNO2-NTSNA %ppm 0 (control)1.7517103.52.49 transplant1.9959853.46.40 midseason2.0442652.76.68 topping1.8429453.25.40

5. Nitrogen Source Study Harrison Co. – Keith Smith Farm 112 kg/ha AN Broadcast Sidedressed with 168 kg/ha each, AN = Ammonium Nitrate, U= Urea, CN = Calcium Nitrate, SN = Sodium Nitrate

6. Nitrate & Nitrite levels from different N sources Harrison Co. – Keith Smith Farm 112 kg/ha AN Broadcast Sidedressed with 168 kg/ha each, AN = Ammonium Nitrate, U= Urea, CN = Calcium Nitrate, SN = Sodium Nitrate LaminaMid-rib

7. Nitrosamine levels from different N sources Harrison Co. – Keith Smith Farm 112 kg/ha AN Broadcast Sidedressed with 168 kg/ha each, AN = Ammonium Nitrate, U= Urea, CN = Calcium Nitrate, SN = Sodium Nitrate Lamina Mid-rib

8. Fertility Trial Jackson Co. – Mike Huff Farm Number = kg of product/ha, AN = Ammonium Nitrate, CN = Calcium Nitrate, SN = Sodium Nitrate Broadcast Sidedress

9. Nitrosamine levels from different N sources Jackson Co. – Mike Huff Farm Number = lbs of product, AN = Ammonium Nitrate, CN = Calcium Nitrate, SN = Sodium Nitrate Broadcast Sidedress LaminaMid-rib

10. Nitrosamine levels from different N sources Jackson Co. – Mike Huff Farm Number = lbs of product, AN = Ammonium Nitrate, CN = Calcium Nitrate, SN = Sodium Nitrate Broadcast Sidedress Lamina Mid-rib

11. Effects of Nitrogen Rate on Burley Yield Madison Co. – Gregg Harris Farm All plots received 450 lb/a Ammonium Nitrate Broadcast followed by the remainder of the nitrogen shortly after transplanting. AN = Ammonium Nitrate. 14-0-14 is a calcium/potassium nitrate mix 161

12. Effects of Nitrogen Rate on Burley Yield Shelby Co. – Gilbert Ray Tucker, Jr. Farm All plots received 450 lb/a Ammonium Nitrate Broadcast followed by the remainder of the nitrogen shortly after transplanting. AN = Ammonium Nitrate, MS=mid-season, LS=late season. Adding 714 lb/a 14-0-14 significantly improved yield over 450 lb/a alone

13. Influence of Seed Source on TSNA Levels

14. Screening Air-cured Varieties for Nornicotine Conversion High levels of the nornicotine increases chance for high TSNA levels A variety may have a high number of converters in a population Conversion is Chemical Change of Nicotine to Nornicotine By selection of non-converters, a “clean” line can be established

15. Ethephon Treatment

21. Screening (3%)

22. Effect of Screening Lowers mean conversion does not necessarily eliminate all converter plants most dramatic with high converters reduced effect with low converters

23. Effect of screening 82 Unscreened Screened TN 90 VA 509 57 5

24. Bottom Line Screened seed lower nornicotine will HELP reduce TSNAs – no guarantee enable growers to meet manufacturers needs Need to identify screened seedlots can only do this by renaming screened varieties

25. “ “LC” Varieties Screened varieties Re-released as “LC” varieties “LC” varieties available to growers in 2005 TN 90LC, TN 86LC, TN 97LC, KT 200LC, KY 907LC NO difference except for conversion Same for yield, quality, disease resistance TN 90 “LC” is still TN 90

26. Accumulation of TSNA during Storage

27. Air-curing experiments have shown that a substantial nitrite production during air- curing does not necessarily lead to high levels of TSNA in cured tobacco. pH of the cured tobacco = 6 Not an effective pH for reaction between alkaloids and nitrite Other Nitrogenous compound may react with alkaloids even under dry conditions

28. Avoiding TSNA accumulation during storage of cured tobacco Good curing process with as low nitrite formation as possible Tobacco in the bales should not be packed too tightly Allows ventilation of nitrogenous compounds into atmosphere Instead of reacting with alkaloids Moisture content in the bales should be as low as possible Prevent any chemical reactions and microbial growth