KRA-01

FDA/ORA/ORS

LIB #4578 25 of 25

Sample Number Sample

Confirmation GC/MS

Quantitation UPLC

26 27 28 29 30

Liquid Extract

Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y

1.56% 1.86% 1.74% 0.495% 0.049% 0.076% 0.060% 0.101% 0.166% 0.173% 0.174% 0.056% 0.080% 0.038% 0.037%

Biotanical Extract powder gelatin capsule, powder

Suspension Oil

Spent Herbs - Solid Material

a Sub 1 # 009 b Sub 2 #003 c Sub2 # 004 d Sub 2 #010 e Sub 2 #011 f Sub 3 #005 g Sub 3 #006 h Sub 3 #012 i Sub 3 # 013 j Sub 4 #007 k Sub 4 # 014

31 32 33 34 35

Brown Liquid

84.95 ug/mL

Mitragynine Extract

34.65%

Brown Liquid Brown Liquid

198 ug/mL

1.32%

Liquid Supplemental Pack

398.1 ug/mL

CONCLUSION This LIB describes the method development and validation for the detection, quantification, and confirmation of identity for mitragynine in products containing Kratom ( Mitragyna speciosa) . The GC/MS is the primary technique for the initial qualitative determination (screening) of mitragynine in Kratom. Products resulting in a positive response to mitragynine are quantified via the UPLC/PDA. A secondary analytical technique, LC-MS/MS, was validated and used for qualitative identification and quantitation of mitragynine in Kratom products. REFERENCES 1. Asian leaf 'kratom' making presence felt in US emergency rooms, NBC News U.S. News Monday Mar 19, 2012 1:48 PM 2. Kikura-Hanajiri R., Kawamura M., et al Forensic Toxicoogy, 27: 2009, 67-74 3. Chan K.B., Pakiam C., Rahim R.A. Bulletin on Narcotics, vol. LVII, Nos. 1 and 2, 2005, 249-256 4. US FDA/ Center for Veterinary Medicine Mass Spectrometry for Confirmation of the Identity of Animal Drug Residues, Guidance for Industry #118; 2003 .