An Alternative Approach to Quantitative Methods: Limit test for Analysis of Deeming Analytes in Electronic Cigarettes

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1 An Alternative Approach to Quantitative Methods: Limit test for Analysis of Deeming Analytes in Electronic Cigarettes Carl Adams, Salem Chouchane and Darius Grissom

2 Questions 1000 or more samples of different electronic cigarettes are being analyzed Many constituents (deeming analytes), are reported as below detections, ND, below LOQ, BDL, BQL, NQ etc When quantitated, levels are extremely low, in ppb, or even ppt Aromatic amines, ammonia, BaP, etc Is it efficient, cost effective, and common sense to continue quantitating, in E-liquid or aerosol, constituents that are consistently either not present in the matrices or at extremely low level?. Lower than what you can find in environment or food. 2

3 Questions Can constituents in E-liquid or aerosol be considered impurities? Is Limit Test an acceptable analytical methodology to monitor such constituents in electronic cigarettes, or ENDs products? 3

4 Background It is well known in industry that regulating agencies will require a product manufacturer to identify and control the level of impurities in a product, such that the integrity of the product is maintained with respect to identity, strength, quality, and purity for use by the consumer or patient. As part of the purity analysis of a product during the development lifecycle, it is typical for product manufacturers to characterize impurities that may arise from the synthesis, manufacturing and packaging of those products and also the degradation products that occur over the shelf life of that product. 4

5 Background While it is common for many companies to quantitate and report specific levels of impurities in their developmental or commercial products, the specific quantitation of each impurity is not required provided that the overall total of impurity is below a limit of significance. The significance of a limit can be based upon, but is not limited too: Toxicological Assessment (i.e. Lowest Concentration Resulting in a Toxic Effect or (TCLo), Acceptable Daily exposure (ADE) or Occupational Exposure Limit (OEL) Regulatory requirement (i.e. NNN in Smokeless Products) Best Practice Process Capability Equipment Capability Economic Factors (industry/region dependent) 5

6 The LIMIT Test Limit Test A quantitative or semi-quantitative test designed and validated to allow an organization to assess the overall purity of their product with respect to a specific level of known or theoretical impurity. 6

7 Regulatory, Compendia, and Guidelines Are Limit Tests allowed according to Regulatory Agencies, compendia, and guidelines? Answer: Yes Food and Drug Administration: A review of the FDA website allows the user to pull up the guidance for industries whereby Q2A states: Testing for impurities can be either a quantitative test or a limit test for the impurity in a sample. Either test is intended to accurately reflect the purity characteristics of the sample. Different validation characteristics are required for a quantitative test than for a limit test. FDA Guidance for industry ANDAs: Impurities in Drug Substances International Conference on Harmonisation, ICH (2003) Q1A (R2) Stability testing of new drug substances and products, USA International Conference on Harmonisation, ICH (1995) Q2A,Validation of analytical procedures(definitions and terminology), USA12. International Conference on Harmonisation, ICH (1996) Q2B,Validation of analytical procedures:methodology, USA 7

8 Regulatory, Compendia, and Guidelines United States Pharmacopeia: Category II Analytical procedures for the determination of impurities in bulk drug substances or degradation compounds in finished pharmaceutical products. These procedures include quantitative assays and limit tests. International Conference on Harmonization: The discussion of the validation of analytical procedures is directed to the four most common types of analytical procedures: - Identification tests; - Quantitative tests for impurities' content; - Limit tests for the control of impurities; - Quantitative tests of the active moiety in samples of drug substance or drug product or other selected component(s) in the drug product. 8

9 What are the benefits of Limits test? The benefits of limit tests come in the form of: Laboratory Efficiency Faster development and validation Fewer standard preparations Reduced # of injections per sequence Simplified data processing, reviewing and reporting Higher throughput. Cost Savings Reduced consumption of reference standards Reduced consumption of mobile phase Reduced labor hours for analysis and data processing and review 9

10 Example Efficiency Gain Efficiency Gain Typical Sequence Quantitative Limit Solvent Solvent Calibration Blank Calibration Blank L1 L1 L2 FMCS Background L3 FMCS Level 1 L4 Rep 1 L5 Rep 2 Blank Rep 3 ICV Rep 4 Rep 1 Rep 5 Rep 2 Rep 3 40 Minute efficiency gain Rep 4 on instrument Rep 5 CCV 10

11 Regulatory, Compendia, and Guidelines Are the ICH, USP, and FDA in agreement with respect to what validation parameters are required? Answer: YES Validation parameter required per ICH, USP, and FDA for different type of tests testing for Impurities Validation parameters Assay Quantitative Limit Identification Accuracy YES YES NO NO Precision - Repeatability YES YES NO NO Precision - Intermediate Precision YES (1) YES * NO NO Specificity YES YES YES YES Detection Limit NO NO YES NO Quantitation Limit NO YES NO NO Linearity YES YES NO NO Range YES YES NO NO Robustness YES YES NO NO 11

12 Validation Strategies We identified two ways in which limit test methods can be validated. 1. Validate as Limit Test; Implement at Limit Test Confidence that analytes will not be present at significant levels in Vapor products (i.e. PAAs, BaP, Ammonia, SVO). 2. Validate as Quantitative Test; Implement as Limit Test with option to Quantitate. Confidence that most formulations will result in trace level impurities, but new product formulations may vary (i.e. Diacetyl; Acetyl Propionyl). 12

13 Preparing Your Limit Standard How to prepare the limit standard: External Standard diluted in solvent (clean). External Standard spiked/extracted/diluted in matrix (dirty, builds in comparative recovery). Deuterated Internal Standard for direct comparison. Surrogate Internal Standard with defined relative response factor Blank Corrected Response We used external Standard in Matrix, blank corrected where applicable. 13

14 Some examples Deeming Method Validations in Electronic Cigarette Aerosol and Liquid: Benzo-a-pyrene (Limit) Select Volatile Organics (Limit) Ammonia (Limit with Option to Quant) Primary Aromatic Amine (Limit with Option to Quant) Diacetyl; Acetyl Propionyl (Limit with Option to Quant) Leachables Method Validations in Electronic Cigarette Aerosol: Organic Leachables by LCMSMS Quantitative with level 1 standard at or below Analytical Evaluation Threshold (AET). Organic Leachables by GCMS Quantitative with level 1 standard at or below AET. 14

15 Some examples Vapor Analysis: External Limit Standards in Matrix 50 ng/ml Ammonia Standard vs. Aerosol (IC) ng/mL PAA Standards vs. Aerosol (GCMS) 0.1 ng/ml BaP Standard vs. Aerosol(UPLC/Fluorescence) ng/ml SVO Standards vs. Aerosol (GCMS) Note: Specifics about each method will be available during the Poster session. 15

16 Example Results Example Results for PAAs in Electronic Cigarette Aerosol Sequence #1 Sample Reps 1-Amino 2-Amino 4-Aminobi NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004ng/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004ng/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004ng/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004ng/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004g/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004g/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004g/puff NMT 0.004ng/puff NMT 0.005ng/puff NMT 0.004g/puff 16

17 Example Results Example Results for Ammonia in Electronic Cigarette Aerosol Samples (µg NH3/E-Cig) _1 NMT _2 NMT _3 NMT _4 NMT _5 NMT _1 NMT _2 NMT _3 NMT _4 NMT _5 NMT _1 NMT _2 NMT _3 NMT _4 NMT _5 NMT 1.02 Sample ID Calculated Concentration (µg NH3/E-Cig) _1 NMT _2 NMT _3 NMT _4 NMT _5 NMT 1.02 Limit 1.02 µg/e-cig 17

18 Example Results Final concentration pg/puff Sample ID NNN NNK <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 <9.4 Final concentration pg/puff Sample ID NNN NNK <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 < <9.6 <9.4 18

19 Take Aways Limits Tests are acceptable for use in the industry Economically beneficial Improve laboratory efficiency/throughput Simplified laboratory operations For selected constituents only 19

20 Questions 20