Blood Drug Analysis Test No Summary Report

Transcription

1 Collaborative Testing Services, Inc FORENSIC TESTING PROGRAM Blood Drug Analysis Test No Summary Report The sample sets contained blood samples from three cases, each with an individual case scenario. Each case sample consisted of two grey-topped vials containing human blood. Participants were requested to examine these items and report their findings. Data were returned from 119 participants and are compiled into the following tables: Page Manufacturer's Information Summary Comments Table 1: Item 1 Results Table 2: Item 2 Results Table 3: Item 3 Results Table 4: Additional Comments Appendix: Data Sheet This report contains the data received from the participants in this test. Since these participants are located in many countries around the world, and it is their option how the samples are to be used (e.g., training exercise, known or blind proficiency testing, research and development of new techniques, etc.), the results compiled in the Summary Report are not intended to be an overview of the quality of work performed in the profession and cannot be interpreted as such. The Summary Comments are included for the benefit of participants to assist with maintaining or enhancing the quality of their results. These comments are not intended to reflect the general state of the art within the profession. Participant results are reported using a randomly assigned "WebCode". This code maintains participant's anonymity, provides linking of the various report sections, and will change with every report.

2 Manufacturer's Information The sample sets contained blood samples from three cases, each with an individual case scenario. Each case sample consisted of two grey-topped vials containing human blood. Participants were asked to analyze the blood samples and report the presence of any drugs/metabolites and quantitative data obtained (including uncertainty). SAMPLE PREPARATION- The human blood used in this test was from the same lot, which tested negative for a variety of common controlled substances prior to being obtained from a commercial supplier. A stock solution of each drug was used to spike specific items. These solutions were obtained in sealed ampoules and were not opened until needed for production. Items were prepared at separate times using the following procedure, and different glassware was used for each item. ITEM 1 (PREPARATION): This item was prepared without the addition of any controlled substances. The blood was pipetted into each of the prelabeled vials which contained Potassium Oxalate and Sodium Fluoride. The vials were sealed and inverted multiple times to mix the chemicals in the vials with the blood. All vials were placed in a refrigerator immediately after production and stored there until the sample sets were prepared. ITEMS 2 and 3 (PREPARATION): Item preparation consisted of adding a predetermined amount of drug stock solution to human whole blood. It was stirred before pipetting the mixture into each of the pre-labeled vials, which contained Potassium Oxalate and Sodium Fluoride. The vials were sealed and inverted multiple times to mix the chemicals in the vials with the blood solution. All vials were placed in a refrigerator immediately after production and stored there until the sample sets were prepared. SAMPLE SET ASSEMBLY: Each sample set contained two vials of each of the three Items and placed into a Department of Transportation regulated shipping container. The sample packs were then returned to the refrigerator until shipment. VERIFICATION- Laboratories that conducted predistribution analysis of samples reported consistent drug/metabolite identifications with one exception, only one participant reported the presence of EDDP. The reported concentration results were proportionally comparable to the preparation drug concentrations. This information is representative of a small number of laboratories, evaluation of results should be deferred until the Summary Report is published. Item 1 Drug ( Concentration ) No drugs or metabolites added Item 2 Drug ( Concentration ) Item 3 Drug ( Concentration ) (260 ng/ml ng/ml) (65 ng/ml ng/ml) EDDP (70 ng/ml ng/ml) Alprazolam (100 ng/ml ng/ml) Please note that the preparation concentration is the value used for calculations during the test preparation phase and may not necessarily represent the final concentration of the samples. It is advised to wait for the Grand Mean statistics available in the Summary and Individual Reports before evaluating performance. ( 2 )

3 Summary Comments This test was designed to allow participants to assess their proficiency in the examination for the presence and concentration of drugs and/or metabolites in blood. Each participant was supplied with two vials containing human blood spiked with differing drugs and/or metabolites for each of three case scenarios. Participants were asked to report the presence of any drugs/metabolites, quantitative data obtained (including uncertainty), and methods used. (Refer to the Manufacturer's Information for preparation details.) Of the 117 participants who reported screening results for Item 1, 106 (90.6%) reported No drugs/metabolites detected. Another 11 participants (9.4%) reported Drug(s) detected utilizing screening methods, five listing over-the-counter substances, five reporting possible controlled substances and one reporting no immunoassay was performed. Of the 90 participants that reported results for confirmatory analysis for Item 1, 83 participants (92.2%) confirmed the absence of any drugs/metabolites while seven participants (7.8%) reported detecting substances. These seven participants had also reported the detection of some type of substance during screening, five confirming the presence of over-the-counter substances and two confirming controlled drugs. Of the 115 participants who responded to the Item 2 screening portion of the test, 114 (99.1%) reported Drug(s) detected while one participant reported No drugs detected utilizing screening methods however a confirmatory analysis was performed by this participant. There were 112 (98.2%) participants that reported the presence of Alprazolam and/or Benzodiazepines, 87 (76.3%) participants that reported the presence of and/or Opiates and of these, 19 (16.7%) also reported the presence of EDDP. One participant reported that they did not screen the sample by immunoassay. Of the 111 participants that reported results for confirmatory analysis for Item 2, 109 (98.2%) reported the presence of, 107 participants (96.4%) reported the presence of Alprazolam and 26 (23.4%) reported the presence of EDDP. Only 25 of the 111 participants reported finding all three substances. There were three participants (2.7%) that reported additional substances in Item 2 that included over-the-counter medications. Of the 113 participants who reported screening results for Item 3, 82 participants (72.6%) reported Drug(s) detected and 79 of these participants reported the presence of (PCP). There were 31 participants (27.4%) that reported No drugs detected utilizing screening methods, 25 of which went on to perform a confirmatory analysis. There were three participants that did not report screening results for Item 3, however did perform a confirmatory analysis. Of the 109 participants that reported results for confirmatory analysis for Item 3, 105 participants (96.3%) reported PCP including six participants who also found other substances in addition to PCP. Three participants (2.8%) reported and one participant reported a combination of over-the-counter and prescription drugs only. If a participant indicated that the confirmatory quantitative result was a single determination and reported it in ng/ml, the conclusive quantitative result was included in the raw data table. The raw data was used to calculate the grand mean and standard deviation for each item and are supplied to assist the participants and accrediting bodies in determining the acceptability of results. For Item 1, statistical analysis was not performed. For Item 2, there were no participants determined to have "extreme" data (±3 STD from grand mean) for any of the three drugs present in the sample. For Item 3, two participants were determined to have "extreme" data for PCP. ( 3 )

4 Screening Results - Item 1 TABLE 1A Item 1 Item Scenario: A 55 year old male driver was killed in a single-vehicle crash. Witnesses described the car drifting between lanes before crashing into the median. There were no other occupants in the vehicle. Item Contents and Preparation Concentration: No drugs or metabolites added Webcode 2ABGZF 2ATE97 2B8WAQ 2BQRRT 2RALHH 2RCBME 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4L4FBH 4LXETP 4VKDKU 69BD8G 6GZC6F 6NK9FG 6VFHDT 6VH8HQ 78TYLQ 7GE4VB 82LJZA 84NQUL 84TRBE 87VYAX 89ZEUH 9CYZ87 9LJJFJ 9RPCAE 9RYXEB Screening Results Pregabalin ( 4 )

5 TABLE 1A Item 1 Webcode A2YURD ADNQTB AHZW7W B3JDYN B9MCAD DAEXB9 DAUDAH DERKZ8 DGY6N8 DLPFY6 DRD8Z7 E2JFA6 EAPBY4 EP9RC2 ERRTBU ERUNPX EYBZ3Y FK2ZBE FVM4KY FZ76WX G8BQ3Y GGLV93 GUUGV2 H3EERY HA4KU2 HGNKCZ HGZDWF HL9YKD HZRJ48 J3T68X JCQLDU JM2ZA2 Screening Results Caffeine Acetaminophen, Chlorpheniramine, Gabapentin. Atenolol, Ibuprofen, Salicylic Acid, Naproxen [No screening results reported.] Screening: possible gabapentin possible promethazine possible lamotrigine possible cannabinol possible dehydroisoandrosterone possible chlorpheniramine Naproxen, Ibuprofen, Omeprazole, Salicylic Acid ( 5 )

6 TABLE 1A Item 1 Webcode JPTCUT K9GZUM KGBA3W KKRJ63 KWKWQT LMXB4V LQWNPZ LQWTXG LRBZLJ LVBCFX M2NQCZ MRPWBN MWG4TW NC99QT NTCXQW NXQKGU PJKNXT PUJKWN PYA4QQ QP9NRE QTMLWF RB4UVV RD2H36 RK7GEF TVG9HF TVLCMR U26DNN U6RN9N U8CMFQ UHBEMW UPQ9NK UVYG6N UX3CQD VE3L63 VH6KFV VNU3YD Screening Results Acetyl Fentanyl Caffeine [No screening results reported.] ( 6 )

7 TABLE 1A Item 1 Webcode VUY26W VVHQNH W82PYC WF4M2M WFYJXA WL9JLM WXWVPJ XANPDY XLZXGQ XVM6YF XVNN2X XY9RWA YFPCRK YLZJDH YN3D3M YUQZXR YWWCZM ZEB9V8 ZFMUK9 ZGHQT3 Screening Results Acetaminophen No immunoassay was performed Acetyl Fentanyl Item 1 - Response Summary Participants Reporting Screening Results: 117 : 106 *Other: 11 Participants can report multiple drugs/metabolites therefore the sum of the values here may be greater than the total number of participants responding for this item. *This category represents the total number of participants that reported a response other than that which is listed above. ( 7 )

8 Confirmatory Results - Item 1 What drugs/metabolites were detected in Item 1? TABLE 1B Item 1 Item Scenario: A 55 year old male driver was killed in a single-vehicle crash. Witnesses described the car drifting between lanes before crashing into the median. There were no other occupants in the vehicle. Item Contents and Preparation Concentration: No drugs or metabolites added Webcode 2ABGZF Qualitative Analyte Reported Only Reported Concentration Uncertainty Units 2BQRRT 2RALHH 2RCBME 2YBZ4J 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4LXETP 6GZC6F 6NK9FG 6VFHDT 6VH8HQ 78TYLQ 82LJZA 84NQUL 89ZEUH Meprobamate Methamphetamine Pregabalin 9LJJFJ 9RPCAE 9RYXEB A2YURD ADNQTB AHZW7W B3JDYN ( 8 )

9 TABLE 1B Item 1 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units B9MCAD DAEXB9 DAUDAH DERKZ8 DGY6N8 DLPFY6 E2JFA6 EAPBY4 EP9RC2 ERRTBU ERUNPX EYBZ3Y FK2ZBE FVM4KY Caffeine FZ76WX G8BQ3Y GGLV93 GUUGV2 HA4KU2 HGNKCZ HGZDWF Atenolol <50 ng/ml Ibuprofen <5000 ng/ml Naproxen <5000 ng/ml Salicylic Acid <5000 ng/ml HL9YKD J3T68X Gabapentin JCQLDU JM2ZA2 K9GZUM KGBA3W Ibuprofen % mg/l Naproxen % mg/l Omeprazole Salicylic Acid ( 9 )

10 TABLE 1B Item 1 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units KKRJ63 LQWTXG LVBCFX M2NQCZ MRPWBN Caffeine NC99QT NXQKGU PJKNXT PYA4QQ QP9NRE QTMLWF RB4UVV RD2H36 TVG9HF TVLCMR U26DNN U6RN9N U8CMFQ UVYG6N VE3L63 VH6KFV VNU3YD VUY26W WF4M2M WFYJXA WXWVPJ XANPDY XLZXGQ XVM6YF XVNN2X XY9RWA YFPCRK YLZJDH Acetaminophen <250ng/ml ( 10 )

11 TABLE 1B Item 1 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units YN3D3M YUQZXR YWWCZM ZEB9V8 ZGHQT3 Item 1 - Response Summary Participants Reporting Confirmatory Results: 90 : 83 *Other: 7 Participants can report multiple drugs/metabolites therefore the sum of the values here may be greater than the total number of participants responding for this item. *This category represents the total number of participants that reported a response other than that which is listed above. ( 11 )

12 Raw Data - Item 1 List of raw data determinations in ng/ml. TABLE 1C Item 1 Item 1 Raw Data - Other Webcode Analyte Raw Data (ng/ml) FZ76WX Atenolol Ibuprofen 1, Naproxen Salicylic Acid JCQLDU Ibuprofen Naproxen Statistical Analysis for Item 1 - Other Please note statistical analysis has not been provided due to the low number of raw data responses. ( 12 )

13 Reporting Procedures - Item 1 If quantitative analysis was performed, the reported concentrations are: TABLE 1D Item 1 Webcode 6GZC6F FZ76WX JCQLDU M2NQCZ U8CMFQ WFYJXA XY9RWA Quantitative Reporting Procedures The mean of duplicate/several determinations. No drugs were detected. Response Summary for Item 1 Participants: 7 A single determination: The mean of duplicate/several determinations: Other: 5 (71.4%) 1 (14.3%) 1 (14.3%) ( 13 )

14 Methods of Analysis - Item 1 TABLE 1E Item 1 Webcode Method Screening Confirmatory Quantitation 2ABGZF 2ATE97 2B8WAQ 2BQRRT 2RALHH 2RCBME 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG /FID 4DDRFF 4DNZXE GC/FID 4L4FBH 4LXETP 4VKDKU 69BD8G 6GZC6F 6NK9FG 6VFHDT 6VH8HQ 78TYLQ LC/MS 7GE4VB ( 14 )

15 TABLE 1E Item 1 Webcode Method Screening Confirmatory 82LJZA 84NQUL 84TRBE 87VYAX 89ZEUH 9LJJFJ 9RPCAE 9RYXEB A2YURD GC/FID ADNQTB AHZW7W GC/FID B3JDYN B9MCAD GC/NPD DAEXB9 LC-QTOF DAUDAH DERKZ8 GC/FID DGY6N8 DLPFY6 GC/FID DRD8Z7 E2JFA6 Quantitation ( 15 )

16 TABLE 1E Item 1 Webcode Method Screening Confirmatory EAPBY4 EP9RC2 ERRTBU ERUNPX EYBZ3Y FK2ZBE FVM4KY FZ76WX G8BQ3Y GGLV93 GUUGV2 H3EERY HA4KU2 HGNKCZ HGZDWF HL9YKD HZRJ48 J3T68X JCQLDU JM2ZA2 JPTCUT Quantitation LC-QTOF MS UPLC-QTOF-MS HPLC-DAD LC/MS ( 16 )

17 TABLE 1E Item 1 Webcode Method Screening Confirmatory K9GZUM GC/FID KGBA3W KKRJ63 KWKWQT LMXB4V LQWNPZ LQWTXG LRBZLJ LVBCFX /FID MRPWBN LC-QTOF MWG4TW NC99QT NTCXQW NXQKGU GC-FID PJKNXT PUJKWN PYA4QQ QP9NRE QTMLWF GC/FID RB4UVV GC/FID RD2H36 RK7GEF TVG9HF HPLC/UV-DAD Quantitation ( 17 )

18 TABLE 1E Item 1 Webcode Method Screening Confirmatory Quantitation TVLCMR U26DNN U6RN9N U8CMFQ UHBEMW UPQ9NK UVYG6N UX3CQD VE3L63 LC/MS VH6KFV VNU3YD VUY26W VVHQNH W82PYC WF4M2M WFYJXA LCMSQTOF WL9JLM WXWVPJ XANPDY XLZXGQ XVM6YF LC/QTOF XVNN2X XY9RWA GC/FID ( 18 )

19 TABLE 1E Item 1 Webcode Method Screening Confirmatory Quantitation YFPCRK /FID YLZJDH YN3D3M YWWCZM GC/FID ZEB9V8 GC/FID ZFMUK9 ZGHQT3 Response Summary for Item 1 Participants: 116 Screening Confirmatory Quantitation : : LC/MS: : Other: ( 19 )

20 Additional Comments for Item 1 TABLE 1F Item 1 Webcode 2RALHH Butyl Acetate- Promazine ISTD Item 1 - Comments 3L6ZEK 3LJJYH 4DNZXE 69BD8G 6VFHDT 6VH8HQ 78TYLQ 82LJZA 84TRBE 89ZEUH DAUDAH E2JFA6 EAPBY4 EP9RC2 ERUNPX FK2ZBE FVM4KY FZ76WX G8BQ3Y GGLV93 GUUGV2 HA4KU2 HGNKCZ ELISA screening panel includes: amphetamine, benzodiazepines, cannabinoids, carisoprodol, cocaine and metabolites, methadone, opiates, oxycodone/oxymorphone, phencyclidine and zolpidem. Laboratory does not routinely analyze postmortem samples (outside scope of testing). Negative Basic-GC/FID is used for quantitation, but not needed. No drugs present. No Basic Drugs Detected. ELISA was used to screen the sample. The sample was screened using Immunalysis kits for Opiates, Benzodiazepines, PCP, Cocaine/BE, Cannabinoids and Methamphetamine. Internal Standard flurazepam / Nalorphine Internal Standards used: phenyltoloxamine and heptabarbital Gabapentin was detected in all items, which is considered a component of the biological matrix and does not correlate with the context of case. Indications of promethazine. The sample was screened for the following type / class of drugs: Amphetamines, Benzodiazepines, Cannabinoids, Cocaine, Opiates, PCP Two internal standards were used: Methamphetamine-D9 and Codeine-D3 Hexobarbital and Phenyltoloxamine were the internal standards used. For the base extract fraction: possible chlorpheniramine noted Internal standard: mepivacaine Phenyltoloxamine - Internal Standard Chlorpheniramine and Promethazine were indicated Confirmation of acetaminophen done by UPLC. Butyl acetate screen used promazine as the internal standard. A peak with a mass spectrum similar to caffeine was detected by GC-MS. Further confirmation or quantitation is not routinely performed for this analyte at our lab. A peak with a mass spectrum similar to the synthetic cannabinoid JWH 018 2'-naphthyl-N-(3-methylbutyl) isomer was detected by GC-MS. Further confirmation for this analyte is not performed at our lab, due to the lack of certified standard for this cannabinoid. Screening & Quantitative analysis for Atenolol, Ibuprofen, Salicylic Acid, Naproxen. Instrument: UPLC-QTOF MS (Waters) (which is ). Salting-out assisted extraction. Internal Standards: Cyclobarbitone, Prazepam & D3-. Limits of detection: 5ng/mL Methaqualone was used as internal standard (ISTD). mepivacaine was used as internal standard Mepivacaine=internal standard I.S. mepivacaine internal standards used - mepivacaine, nalorphine, amphetamine-d11, methamphetamine-d11 ( 20 )

21 TABLE 1F Item 1 Webcode HGZDWF HL9YKD J3T68X JCQLDU K9GZUM KKRJ63 KWKWQT LQWNPZ MWG4TW NC99QT NTCXQW PUJKWN PYA4QQ RD2H36 TVG9HF U26DNN UX3CQD VE3L63 WF4M2M WFYJXA WL9JLM XANPDY XLZXGQ Item 1 - Comments Screening was not performed. STI used was tetracosane, limit of detection is 300 ng/ ml Other possible drugs in sample include: possible promethazine, possible lamotrigine, possible cannabinol, possible dehydroisoandrosterone, possible chlorpheniramine. Internal Reference Materials Used : Phenyltoloxamine, Hexobarbital, and 11-Hydroxy-Delta 9-THC Internal standard: Mepivacaine Indications of chlorpheniramine and promethazine. entry is actually a LC-QTOF-MS, online software would not allow addition of two "Other" methods. ISTD used for quantitation of Ibuprofen and Naproxen was Meclofenamic Acid. LOD Ibuprofen 0.5 mg/l, LOD Naproxen 0.1 mg/l. Omeprazole and Salicylic acid not routinely quantitated. No Basic Drugs Detected IS - Mepivacaine ELISA screening panel includes: amphetamine, benzodiazepines, cannabinoids, carisoprodol, cocaine and metabolites, methadone, opiates, oxycodone/oxymorphone, phencyclidine, and zolpidem. No further testing is performed following a none detected screen. Laboratory does not routinely analyze postmortem samples. This specimen screened negative for the following assays: benzodiazepines, cannabinoids, cocaine/metabolites, amphetamines, opiates, and phencyclidine. Screening Cut-Off's: Amphetamine: 20 ng/ml. Benzoylecgonine: 50 ng/ml. Carboxy-THC: 20 ng/ml. Methamphetamine: 20 ng/ml. Morphine: 20 ng/ml. Oxazepam: 50 ng/ml Mepivacaine used as internal standard. Indications of promethazine were observed, but did not meet criteria to report. Amphetamine/Methamphetamine/Morphine/C-THC: 20ng/mL, BE/Oxazepam: 50ng/mL ELISA screening panel includes: amphetamine, benzodiazepines, cannabinoids, carisoprodol, cocaine and metabolites, methadone, opiates, oxycodone/oxymorphone, phencyclidine, and zolpidem. The laboratory does not routinely analyze postmortem samples (outside scope of testing). Internal standard: Mepivacaine Indications of Chlorpheniramine and Lamotrigine Not detected Flurazepam as Internal Standard INTERNAL STANDARD USED FOR ANALYSIS : phenobarbital D5. INTERNAL STANDARD USED FOR ANALYSIS : CODEINE D3. INTERNAL STANDARDS USED FOR HPLC/UV-DAD: DEXTROPROPOXYPHENE. Butyl Acetate - Promazine (ISTD) Screening test only performed. Estazolam was used as internal standard. LCMSMS internal standard - mepivacaine. GCMS internal standard - mepivacaine The Laboratory Lowest Caliration point for Acetaminophen is 0.25ug/ml. Acetaminophen Concentration was considered as <0.25ug/ml internal standard - mepivacaine Phenyltoloxamine IRM. Heptabarbital IRM Chlorpheniramine seen not reported. IS - Mepivacaine ( 21 )

22 TABLE 1F Item 1 Webcode XVNN2X ZEB9V8 Item 1 - Comments Promazine used as Internal Standard for Screening/Confirmatory testing No Basic Drugs Detected ( 22 )

23 Screening Results - Item 2 TABLE 2A Item 2 Item Scenario: A 17 year old male was pulled over by police for an expired registration. The officer noticed that the individual exhibited slow movements and loss of memory. The result of a breath alcohol test was 0.00%. A Drug Recognition Expert arrived and noted that the individual had flaccid muscles and constricted pupils with no reaction to light. A blood sample was collected 90 minutes after the arrest. Item Contents and Preparation Concentration: (260 ng/ml) EDDP (70 ng/ml) Alprazolam (100 ng/ml) Webcode 2ABGZF 2ATE97 2B8WAQ 2BQRRT 2RALHH 2RCBME 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4L4FBH 4LXETP 4VKDKU 69BD8G 6GZC6F 6NK9FG 6VFHDT 6VH8HQ 78TYLQ 7GE4VB Screening Results Benzodiazepines, methadone alprazolam Alprazolam Benzodiazepines Benzodiazepines Possible metabolite and Alprazolam Benzodiazepines Benzodiazepines and methadone. Benzodiazepines Benzodiazepines benzodiazepines alprazolam methadone EDDP Benzodiazepines Benzodiazepines Benzodiazepines Alprazolam, methadone. Benzodiazepines Alprazolam, EDDP, Benzodiazepines Alprazolam Benzodiazepines and Alprazolam, methadone., EDDP, Alprazolam ( 23 )

24 TABLE 2A Item 2 Webcode 82LJZA 84NQUL 84TRBE 87VYAX 89ZEUH 9CYZ87 9LJJFJ 9RPCAE 9RYXEB A2YURD ADNQTB AHZW7W B3JDYN B9MCAD DAEXB9 DAUDAH DERKZ8 DGY6N8 DLPFY6 DRD8Z7 E2JFA6 EAPBY4 EP9RC2 ERRTBU ERUNPX EYBZ3Y FK2ZBE FVM4KY FZ76WX G8BQ3Y Screening Results Alprazolam and Benzodiazepines Benzodiazepines Benzodiazepines Benzodiazepines Benzodiazepines, methadone Alprazolam : Benzodiazepines class indicative. alprazolam, methadone, and eddp found in LCMSMS screen. Benzodiazepines Benzodiazepines Benzodiazepines Benzodiazepines; methadone Benzodiazepines, Alprazolam, caffeine, EDDP, Benzodiazepines Benzodiazepines and Alprazolam Benzodiazepines Alprazolam EDDP, methadone, and alprazolam Benzodiazepines Benzodiazepines methadone alprazolam Acetaminophen, Alprazolam, Chlorpheniramine, Gabapentin,., EDDP, Alprazolam Benzodiazepine/Alprazolam,Benzodiacepines Alprazolam,, Atenolol, Ibuprofen, Salicylic Acid, Naproxen, Paracetamol METHADONE ( 24 )

25 TABLE 2A Item 2 Webcode GGLV93 GUUGV2 H3EERY HA4KU2 HGNKCZ HGZDWF HL9YKD HZRJ48 J3T68X JCQLDU JM2ZA2 JPTCUT K9GZUM KGBA3W KKRJ63 KWKWQT LMXB4V LQWNPZ LQWTXG LRBZLJ LVBCFX M2NQCZ MRPWBN MWG4TW NC99QT NTCXQW Screening Results Benzodiazepines (alprazolam) EDDP (methadone metabolite) (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine) benzodiazepines (alprazolam); methadone; EDDP (methadone metabolite) alprazolam, EDDP and methadone alprazolam, methadone, EDDP benzodiazepines: alprazolam methadone 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) [No screening results reported.] EMIT Screening: Benzodiazepines Screening: possible methadone possible alprazolam possible gabapentin possible promethazine possible lamotrigine possible cannabinol possible THC possible dehydroisoandrosterone possible chlorpheniramine Alprazolam, EDDP, Benzodiazepines and ELISA Benzodiazepines positive,, Alprazolam, Omeprazole, Paracetamol, benzodiazepines Benzodiazepines, Benzodiazepines Benzodiazepines Benzodiazepines, Benzodiazepines and methadone Benzodiazepines and Benzodiazepines Acetyl Fentanyl, Alprazolam, EDDP, Benzodiazepines Benzodiazepines [No screening results reported.] Alprazolam, caffeine, EDDP, Benzodiazepines Benzodiazepines and indicated positive by ELISA Screening. Benzodiazepines ( 25 )

26 TABLE 2A Item 2 Webcode NXQKGU PJKNXT PUJKWN PYA4QQ QP9NRE QTMLWF RB4UVV RD2H36 RK7GEF TVG9HF TVLCMR U26DNN U6RN9N U8CMFQ UHBEMW UPQ9NK UVYG6N UX3CQD VE3L63 VH6KFV VNU3YD VUY26W VVHQNH W82PYC WF4M2M WFYJXA WL9JLM WXWVPJ Screening Results Benzodiazepines Benzodiazepines, benzodiazepines methadone [No screening results reported.] Specimen screened positive for Benzodiazapines drug class. Benzodiazepine Positive ELISA Benzodiazepines [No screening results reported.] methadone alprazolam METHADONE : OPIODS ALPRAZOLAM: BENZODIAZEPINES Benzodiazepines Benzodiazepines Possible Opiates EDDP,, Alprazolam Alprazolam Alprazolam Benzodiazepines and, Benzodiazepines Benzodiazepines /Metabolite Alprazolam alprazolam methadone and metabolites and alprazolam Benzodiazepines, methadone methadone, benzodiazepines methadone alprazolam Benzodiazepines - indicative Alprazolam benzodiazepines, alprazolam, methadone, EDDP (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine) Benzodiazepines, ( 26 )

27 TABLE 2A Item 2 Webcode XANPDY XLZXGQ XVM6YF XVNN2X XY9RWA YFPCRK YLZJDH YN3D3M YUQZXR YWWCZM ZEB9V8 ZFMUK9 ZGHQT3 Screening Results Benzodiazepines, Benzodiazepines, Benzodiazepines, Benzodiazepines, Possible Opiates Benzodiazepine (drug class) Benzodiazepines benzodiazepines, alprazolam, methadone, and 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) We did not screen sample by immunoassay METHADONE BENZODIACEPINES Alprazolam Benzodiazepines methadone alprazolam Acetyl Fentanyl, Alprazolam, EDDP, Item 2 - Response Summary Participants Reporting Screening Results: 115 / Opiates: 87 EDDP: 19 Alprazolam/ Benzodiazepines: 112 : 1 *Other: 4 Participants can report multiple drugs/metabolites therefore the sum of the values here may be greater than the total number of participants responding for this item. *This category represents the total number of participants that reported a response other than that which is listed above. ( 27 )

28 Confirmatory Results - Item 2 What drugs/metabolites were detected in Item 2? Item Scenario: TABLE 2B Item 2 A 17 year old male was pulled over by police for an expired registration. The officer noticed that the individual exhibited slow movements and loss of memory. The result of a breath alcohol test was 0.00%. A Drug Recognition Expert arrived and noted that the individual had flaccid muscles and constricted pupils with no reaction to light. A blood sample was collected 90 minutes after the arrest. Item Contents and Preparation Concentration: (260 ng/ml) EDDP (70 ng/ml) Alprazolam (100 ng/ml) Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units 2ABGZF Alprazolam 2ATE97 2BQRRT mg/l Alprazolam mg/l Alprazolam ng/ml 2RALHH 2RCBME Alprazolam Alprazolam 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4LXETP 220 ng/ml Alprazolam 98 ng/ml 0.21 ±0.04 μg/ml Alprazolam 0.11 ±0.03 μg/ml ug/ml Alprazolam ng/ml / ug/ml Alprazolam 106 +/- 17 ng/ml mg/l EDDP lower than the lowest N/A μg/l calibrator of 50 Alprazolam mg/l / ug/ml Alprazolam 82 +/ ng/ml Alprazolam ng/ml ( 28 )

29 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units 4VKDKU Alprazolam 69BD8G 6GZC6F 6NK9FG 6VFHDT Alprazolam 97 +/- 14 ng/ml ng/ml EDDP ng/ml Alprazolam ng/ml /-13.20% ug/ml Alprazolam 119 =/-15.89% ng/ml ng/ml Alprazolam ng/ ml 6VH8HQ Alprazolam 78TYLQ ,2 ng /ml Alprazolam 67 6,4 ng/ml 7GE4VB 82LJZA EDDP Alprazolam Alprazolam 84NQUL 87VYAX Alprazolam ng/ml Alprazolam ng/ml 89ZEUH EDDP (methadone Metabolite) Alprazolam 9CYZ ng/ml Alprazolam ng/ml 9LJJFJ Alprazolam 9RPCAE 9RYXEB ng/ml Alprazolam ng/ml mg/l EDDP Alprazolam mg/l ( 29 )

30 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units A2YURD ADNQTB AHZW7W ug/ml ug/ml Alprazolam 77 ng/ml ng/ml / ug/ml Alprazolam 98 +/- 16 ng/ml ug/ml Alprazolam ng/ml B3JDYN Alprazolam B9MCAD DAEXB % ug/ml ng/ml EDDP ng/ml Alprazolam ng/ml DAUDAH Alprazolam DERKZ8 DGY6N8 DLPFY6 DRD8Z7 E2JFA ug/ml % Alprazolam 83 ng/ml % % ug/ml Alprazolam % ng/ml / ug/ml Alprazolam 96 +/- 15 ng/ml ng/ml Alprazolam ng/ml ng/ml EDDP (2-ethylidene-1, 5-dimethyl-3, <50 ng/ml 3-diphenylpyrrolidine) Alprazolam ng/ml EAPBY4 EP9RC2 EDDP Alprazolam Alprazolam ERRTBU ERUNPX mg/l Alprazolam mg/l mg/l Alprazolam 97 6 ng/ml ( 30 )

31 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units EYBZ3Y 230 EDDP 9 Alprazolam 120 FK2ZBE FVM4KY Alprazolam EDDP Alprazolam FZ76WX ng/ml Alprazolam 90 6 ng/ml Atenolol <50 ng/ml Ibuprofen <5000 ng/ml Naproxen <5000 ng/ml Paracetamol <5000 ng/ml Salicylic Acid <5000 ng/ml G8BQ3Y GGLV93 GUUGV2 H3EERY HA4KU2 HGNKCZ ng/ml EDDP (2-ethylidene-1, 5-dimethyl-3, <50 N/A ng/ml 3-diphenylpyrrolidine) Alprazolam ng/ml ng/ml EDDP (methadone metabolite) <50 ng/ml Alprazolam ng/ml EDDP Alprazolam ng/ml mg/l EDDP (2-ethylidene-1, 5-dimethyl-3, 3-diphenylpyrrolidine) Alprazolam mg/l mg/l 2-ethylidene-1, 5-dimethyl-3, 3-diphenylpyrrolidine (EDDP) lower than the lowest calibrator of 0.05 mg/l Alprazolam mcg/l HGZDWF HL9YKD Alprazolam ( 31 )

32 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units HZRJ48 EDDP Alprazolam J3T68X Alprazolam JCQLDU JM2ZA2 K9GZUM % mg/l Alprazolam Approx % mg/l Omeprazole Paracetamol % ug/ml Alprazolam % ng/ml 0.20 ± 0.03 ug/ml Alprazolam 84 ± 13.3 ng/ml KGBA3W KKRJ63 Alprazolam KWKWQT LMXB4V LQWNPZ LQWTXG LRBZLJ LVBCFX 0.21 ±0.04 μg/ml Alprazolam 0.11 ±0.03 μg/ml 223 ng/ml (ng/ml) Alprazolam 89 ng/ml (ng/ml) Alprazolam 96 +/-14 ng/ml ng/ml EDDP ng/ml Alprazolam ng/ml Alprazolam ng/ml / ug/ml Alprazolam 125 +/- 20 ng/ml M2NQCZ EDDP Alprazolam MRPWBN ng/ml EDDP ng/ml Alprazolam ng/ml NC99QT Alprazolam ( 32 )

33 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units NXQKGU /-0.02 ug/ml Alprazolam 81 +/-13 ng/ml PJKNXT Alprazolam PUJKWN ug/ml Alprazolam ug/ml QP9NRE Alprazolam QTMLWF RB4UVV /-0.02 ug/ml Alprazolam 91 +/-15 ng/ml % ug/ml Alprazolam % ng/ml RD2H36 Alprazolam RK7GEF Alprazolam mg/l TVG9HF TVLCMR U26DNN Alprazolam Alprazolam Alprazolam U6RN9N U8CMFQ UHBEMW UPQ9NK EDDP Alprazolam / ng/ml ng/ml Alprazolam ng/ml ng/ml Alprazolam ng/ml ng/ml Alprazolam ng/ml UVYG6N VE3L63 Alprazolam Alprazolam ( 33 )

34 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units VH6KFV 269 ng/ml Alprazolam 87 ng/ml VNU3YD EDDP Alprazolam EMDP VUY26W VVHQNH W82PYC WF4M2M WFYJXA WL9JLM % Alprazolam % 220 ng/ml Alprazolam 90 ng/ml mg/l Alprazolam mg/l mg/l 2-ethylidene-1, 5-dimethyl-3, <0.05 mg/l 3-diphenylpyrrolidine (EDDP) Alprazolam mcg/l % ng/ml Alprazolam 91.04l 13.9% ng/ml ng/ml EDDP (2-ethylidene-1, 5-dimethyl-3, 3-diphenylpyrrolidine) see note below [Table 2F] ng/ml Alprazolam ng/ml WXWVPJ XANPDY XLZXGQ Alprazolam Alprazolam Alprazolam XVM6YF mg/l EDDP Alprazolam mg/l XVNN2X Alprazolam XY9RWA YFPCRK / μg/ml Alprazolam 71 +/- 11 ng/ml /-0.01 ug/ml Alprazolam 100 +/-16 ng/ml ( 34 )

35 TABLE 2B Item 2 Webcode Analyte Reported Qualitative Only Reported Concentration Uncertainty Units YLZJDH mg/l 2-ethylidene-1, 5-dimethyl-3, 3-diphenylpyrrolidine (EDDP) lower than the lowest calibrator of 0.05 mg/l Alprazolam mcg/l YN3D3M Alprazolam YUQZXR YWWCZM ZEB9V8 ZFMUK9 ZGHQT3 0.2 mg/l Alprazolam 0.09 mg/l /-0.03 ug/ml Alprazolam 82 +/-13 ng/ml 0.21 ±0.03 ug/ml Alprazolam 80 ±12.7 ng/ml mg/l Alprazolam mg/l ng/ml EDDP ng/ml Alprazolam ng/ml Item 2 - Response Summary Participants Reporting Confirmatory Results: 111 : EDDP: Alprazolam: 107 *Other: 3 Participants can report multiple drugs/metabolites therefore the sum of the values here may be greater than the total number of participants responding for this item. *This category represents the total number of participants that reported a response other than that which is listed above. ( 35 )

36 Raw Data - Item 2 List of raw data determinations in ng/ml. TABLE 2C Item 2 Item 2 Raw Data - Preparation concentration: (260 ng/ml) Webcode Raw Data (ng/ml) Participant Mean 2ATE YBZ4J L6ZEK LJJYH QG3ZG DDRFF DNZXE NK9FG VFHDT TYLQ CYZ RPCAE RYXEB A2YURD ADNQTB AHZW7W B9MCAD DAEXB DERKZ DGY6N DLPFY DRD8Z E2JFA ERRTBU ERUNPX EYBZ3Y FZ76WX GGLV ( 36 )

37 TABLE 2C Item 2 Item 2 Raw Data - Preparation concentration: (260 ng/ml) Webcode Raw Data (ng/ml) Participant Mean GUUGV HA4KU HGNKCZ JCQLDU JM2ZA K9GZUM KWKWQT LMXB4V LQWTXG LVBCFX MRPWBN NXQKGU PUJKWN QTMLWF RB4UVV U8CMFQ UHBEMW UPQ9NK VH6KFV VUY26W VVHQNH W82PYC WF4M2M WFYJXA WL9JLM XVM6YF XY9RWA YFPCRK YLZJDH YUQZXR ( 37 )

38 TABLE 2C Item 2 Item 2 Raw Data - Preparation concentration: (260 ng/ml) Webcode Raw Data (ng/ml) Participant Mean YWWCZM ZEB9V ZFMUK ZGHQT Statistical Analysis for Item 2 - Grand Mean Standard Deviation Number of Participants Included Number of Participants Excluded 62 0 Number of Participants without Raw Data or Data that was not reported in ng/ml 0 ( 38 )

39 TABLE 2C Item 2 Item 2 Raw Data - EDDP Preparation concentration: (70 ng/ml) Webcode Raw Data (ng/ml) Participant Mean 4DDRFF DAEXB E2JFA EYBZ3Y GGLV GUUGV HA4KU HGNKCZ LQWTXG MRPWBN WF4M2M WL9JLM YLZJDH ZGHQT Statistical Analysis for Item 2 - EDDP Grand Mean Standard Deviation Number of Participants Included Number of Participants Excluded 14 0 Number of Participants without Raw Data or Data that was not reported in ng/ml 0 ( 39 )

40 TABLE 2C Item 2 Item 2 Raw Data - Alprazolam Preparation concentration: (100 ng/ml) Webcode Raw Data (ng/ml) Participant Mean 2ATE BQRRT YBZ4J L6ZEK LJJYH QG3ZG DDRFF DNZXE LXETP BD8G NK9FG VFHDT TYLQ NQUL VYAX CYZ RPCAE RYXEB A2YURD ADNQTB AHZW7W DAEXB DERKZ DGY6N DLPFY DRD8Z E2JFA ERRTBU ERUNPX EYBZ3Y ( 40 )

41 TABLE 2C Item 2 Item 2 Raw Data - Alprazolam Preparation concentration: (100 ng/ml) Webcode Raw Data (ng/ml) Participant Mean FZ76WX GGLV GUUGV H3EERY HA4KU HGNKCZ JCQLDU JM2ZA K9GZUM KWKWQT LMXB4V LQWNPZ LQWTXG LRBZLJ LVBCFX MRPWBN NXQKGU PUJKWN QTMLWF RB4UVV RK7GEF U6RN9N U8CMFQ UHBEMW UPQ9NK VH6KFV VUY26W VVHQNH W82PYC WF4M2M ( 41 )

42 TABLE 2C Item 2 Item 2 Raw Data - Alprazolam Preparation concentration: (100 ng/ml) Webcode Raw Data (ng/ml) Participant Mean WFYJXA WL9JLM XVM6YF XY9RWA YFPCRK YLZJDH YUQZXR YWWCZM ZEB9V ZFMUK ZGHQT Statistical Analysis for Item 2 - Alprazolam Grand Mean Standard Deviation Number of Participants Included Number of Participants Excluded 71 0 Number of Participants without Raw Data or Data that was not reported in ng/ml 0 ( 42 )

43 TABLE 2C Item 2 Item 2 Raw Data - Other Webcode Analyte Raw Data (ng/ml) FZ76WX Atenolol Ibuprofen 1, Naproxen Paracetamol Salicylic Acid Statistical Analysis for Item 2 - Other Please note statistical analysis has not been provided due to the low number of raw data responses. ( 43 )

44 Reporting Procedures - Item 2 If quantitative analysis was performed, the reported concentrations are: TABLE 2D Item 2 Webcode 2ATE97 2BQRRT 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4LXETP 69BD8G 6GZC6F 6NK9FG 6VFHDT 78TYLQ 84NQUL 87VYAX 9CYZ87 9RPCAE 9RYXEB A2YURD ADNQTB AHZW7W B9MCAD DAEXB9 DERKZ8 DGY6N8 DLPFY6 DRD8Z7 E2JFA6 Quantitative Reporting Procedures The mean of duplicate/several determinations. The mean of duplicate/several determinations. The mean of duplicate/several determinations. The mean of duplicate/several determinations. The mean of duplicate/several determinations. both The mean of duplicate/several determinations. The mean of duplicate/several determinations. ( 44 )

45 TABLE 2D Item 2 Webcode ERRTBU ERUNPX EYBZ3Y FZ76WX GGLV93 GUUGV2 H3EERY HA4KU2 HGNKCZ J3T68X JCQLDU JM2ZA2 K9GZUM KWKWQT LMXB4V LQWNPZ LQWTXG LVBCFX M2NQCZ MRPWBN NXQKGU PUJKWN QTMLWF RB4UVV RD2H36 RK7GEF U6RN9N U8CMFQ UHBEMW UPQ9NK VH6KFV VUY26W Quantitative Reporting Procedures The mean of duplicate/several determinations. alprazolam- single determination; methadone - mean of duplicate/several determinations The mean of duplicate/several determinations. The mean of duplicate/several determinations. ( 45 )

46 TABLE 2D Item 2 Webcode VVHQNH W82PYC WF4M2M WFYJXA WL9JLM XVM6YF XY9RWA YFPCRK YLZJDH YUQZXR YWWCZM ZEB9V8 ZFMUK9 ZGHQT3 Quantitative Reporting Procedures The mean of duplicate/several determinations. The mean of duplicate/several determinations. The mean of duplicate/several determinations. Response Summary for Item 2 Participants: 75 A single determination: The mean of duplicate/several determinations: Other: 60 (80.0%) 13 (17.3%) 2(2.7%) ( 46 )

47 Methods of Analysis - Item 2 TABLE 2E Item 2 Webcode Method Screening Confirmatory 2ABGZF 2ATE97 2B8WAQ 2BQRRT 2RALHH 2RCBME 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4L4FBH 4LXETP 4VKDKU 69BD8G 6GZC6F Quantitation LC/MS GC/FID GC/FID GC/FID ( 47 )

48 TABLE 2E Item 2 Webcode Method Screening Confirmatory 6NK9FG 6VFHDT 6VH8HQ 78TYLQ 7GE4VB 82LJZA 84NQUL 84TRBE 87VYAX 89ZEUH 9CYZ87 9LJJFJ 9RPCAE 9RYXEB A2YURD ADNQTB AHZW7W B3JDYN B9MCAD Quantitation LC/MS GC/FID GC/FID LC/MS GC/FID GC/NPD &/MS ( 48 )

49 TABLE 2E Item 2 Webcode Method Screening Confirmatory DAEXB9 DAUDAH DERKZ8 DGY6N8 DLPFY6 DRD8Z7 E2JFA6 EAPBY4 EP9RC2 ERRTBU ERUNPX EYBZ3Y FK2ZBE FVM4KY FZ76WX G8BQ3Y GGLV93 Quantitation LC-QTOF GC/FID GC/FID LC/MS LC-QTOF MS UPLC-QTOF-MS ( 49 )

50 TABLE 2E Item 2 Webcode Method Screening Confirmatory GUUGV2 H3EERY HA4KU2 HGNKCZ HGZDWF HL9YKD HZRJ48 J3T68X JCQLDU JM2ZA2 JPTCUT K9GZUM KGBA3W KKRJ63 KWKWQT LMXB4V LQWNPZ LQWTXG LRBZLJ LVBCFX M2NQCZ Quantitation LC/MS LC-QTOF-MS GC/FID GC/FID ( 50 )

51 TABLE 2E Item 2 Webcode Method Screening Confirmatory MRPWBN MWG4TW NC99QT NTCXQW NXQKGU PJKNXT PUJKWN QP9NRE QTMLWF RB4UVV RD2H36 RK7GEF TVG9HF TVLCMR U26DNN U6RN9N U8CMFQ UHBEMW UPQ9NK UVYG6N UX3CQD Quantitation LC-QTOF GC FID GC/FID GC/FID LC/MS HPLC/UV-DAD ( 51 )

52 TABLE 2E Item 2 Webcode Method Screening Confirmatory VE3L63 VH6KFV VNU3YD VUY26W VVHQNH W82PYC WF4M2M WFYJXA WL9JLM WXWVPJ XANPDY XLZXGQ XVM6YF XVNN2X XY9RWA YFPCRK YLZJDH YN3D3M Quantitation LC/MS GC/NPD LC/MS LCMSQTOF LC/QTOF GC/FID GC/FID ( 52 )

53 TABLE 2E Item 2 Webcode Method Screening Confirmatory YUQZXR YWWCZM ZEB9V8 ZFMUK9 ZGHQT3 Response Summary for Item 2 Quantitation GC/FID GC/FID LC/MS Participants: 118 Screening Confirmatory Quantitation : : LC/MS: : Other: ( 53 )

54 Additional Comments for Item 2 TABLE 2F Item 2 Webcode 2RALHH 3L6ZEK 4DDRFF 4L4FBH 69BD8G 6NK9FG 6VFHDT 6VH8HQ 78TYLQ 82LJZA 84TRBE 89ZEUH 9RYXEB B9MCAD DAUDAH E2JFA6 EAPBY4 EP9RC2 ERUNPX FK2ZBE FVM4KY Item 2 - Comments Butyl Acetate- Promazine ISTD, Benzo Confirmation- Prazepam ISTD (20ug/mL), Opiate Confirmation- Nalorphine ISTD ELISA screening panel includes: amphetamine, benzodiazepines, cannabinoids, carisoprodol, cocaine and metabolites, methadone, opiates, oxycodone/oxymorphone, phencyclidine and zolpidem. Benzodiazepine confirmation panel includes alprazolam, diazepam, 7-aminodiazepam, clonazepam, lorazepam, nordiazepam, oxazepam and temazepam. Alprazolam-D5 and methadone-d3 used as internal standards. LOQ/LOD for alprazolam is 5ng/ml. LOD/LOQ for methadone is 10ng/ml. EDDP's chemical name is 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine, and is a metabolite of methadone. I am currently not competent in confirming Benzodiazepines. Alprazolam-D5 was used as an internal standard with a limit of detection of 10 ng/ml. Quantitation by GC/FID Internal Standard: flurazepam / nalorphine Internal Standards used: phenyltoloxamine and heptabarbital Gabapentin was detected in all items, which is considered a component of the biological matrix and does not correlate with the context of the case. EDDP indications not reported. The sample was screened for the following type / class of drugs: Amphetamines, Benzodiazepines, Cannabinoids, Cocaine, Opiates, PCP. No confirmatory method was used on this sample since that is beyond my current qualifications. Diazepam-D5 was used as internal standard Alprazolam quantification was a single determination. was a mean of two values. EDDP concentration is lower than a lowest calibrator of mg/l. (eddp = 2-ethylidene-1,5-dimethyl- 3,3-diphenylpyrrolidine) Benzodiazepines are only screened in-house. Confirmations are currently outsourced to [laboratory]. Therefore, no benzodiazepine confirmations were able to be performed. Note: was screened using GC/NPD and and confirmed using GC/NPD. Hexobarbital and Phenyltoloxamine were the internal standards used. Internal standards used: mepivacaine, diazepam-d5; clonazepam-d4; gabapentin-d4. Quantitation for EDDP was less than the lowest calibrator of 50 ng/ml Phenyltoloxamine - Internal Standard Gabapentin related compound indicated. Chlorpheniramine indicated. EDDP indicated Confirmation of acetaminophen done by UPLC. Butyl acetate screen used promazine as the internal standard. Opiate confirmation used nalorphine as the internal standard. Benzodiazepine confirmation used prazepam as the internal standard. Alprazolam: we used clonazepam -D4 as internal standard. LOD: 10 ng/ml;, EDDP : we used trimipramine D3 as internal standard LOD : 50 ng/ml ( 54 )

55 TABLE 2F Item 2 Webcode FZ76WX G8BQ3Y GGLV93 GUUGV2 HA4KU2 HGNKCZ HGZDWF HL9YKD J3T68X JCQLDU KGBA3W KKRJ63 KWKWQT LMXB4V LQWNPZ LQWTXG MWG4TW Item 2 - Comments, Atenolol, Ibuprofen, Salicylic Acid, Naproxen, Paracetamol. Screening & Quantitative analysis. Instrument: UPLC-QTOF MS (Waters) (which is ). Salting-out assisted extraction. Internal Standards: Cyclobarbitone, Prazepam & D3-. Limits of detection: 5 ng/ml. Alprazolam Quantitative Analysis. Instrument: UPLC-TQD (Waters). Internal Standard: D5-Diazepam. LOD: 1 ng/ml Methaqualone was used as internal standard (ISTD). mepivacaine and/or diazepam-d5 was used as internal standard. Lowest calibrator for EDDP is 50 ng/ml Mepivacaine and/or diazepam-d5 used as internal standard; lowest calibrator for EDDP (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine)= 50 ng/ml I.S. mepivacaine, diazepam d5 EDDP (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine) quantitative result of 18 μg/l is lower than the lowest calibrator of 50 μg/l and above the limit of report of 3.0 μg/l internal standards used: mepivacaine, nalorphine, diazepam-d5, clonazepam-d4; amphetamine-d11, methamphetamine-d11. Limit of Report (LoR) for 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP): 3 mcg/l Screening was not performed. STI used was tetracosane, limit of detection is 300 ng/ ml, positive methadone detected. Other possible drugs in sample include: possible gabapentin, possible promethazine, possible lamotrigine, possible cannabinol, possible THC, possible dehydroisoandrosterone, possible chlorpheniramine. Internal Reference Materials Used : Phenyltoloxamine, Hexobarbital, and 11-Hydroxy-Delta 9-THC Gabapentin related compound and EDDP seen. Trace peaks of Xylazine, Chlorpheniramine, Promethazine, and Lamotrigine seen. D3 methadone ISTD used to quantitate methadone, prazepam ISTD used to quantitate alprazolam. This sample would be forwarded for further benzodiazepine testing. IS - Mepivacaine ELISA screening panel includes: amphetamine, benzodiazepines, cannabinoids, carisoprodol, cocaine and metabolites, methadone, opiates, oxycodone/oxymorphone, phencyclidine, and zolpidem. Benzodiazepine confirmation/quantitation panel includes 7-aminoclonazepam, alprazolam, diazepam, clonazepam, lorazepam, nordiazepam, oxazepam, and temazepam using 7-aminoclonazepam-D4, alprazolam-d5, diazepam-d5, oxazepam-d5, and temazepam-d5 as internal standards. LOQ/LOD for all aforementioned benzodiazepines is 5 ng/ml. Perform confirmation/quantitation of methadone using methadone-d3 as internal standard following positive methadone screen. LOQ/LOD for methadone is 10 ng/ml. limit of detection for both drugs is 10 ng/ml. IS-Alprazolam-D5 and -D9 1) This specimen screened negative for the following assays: cannabinoids, cocaine/metabolites, amphetamines, opiates, and phencyclidine. 2) The specimen was 'none detected' for the following compounds: oxazepam, alpha-hydroxyalprazolam, nordiazepam, lorazepam, clonazepam, and diazepam. 3) The cutoff for alprazolam is 10ng/mL. Alprazolam: Internal standard - Alprazolam-D5; LOQ - 5ng/mL. EDDP: Internal standard - EDDP-D3; LOQ - 5ng/mL. : Internal standard - -D9; LOQ - 5ng/mL Screening Cut-Off's: Amphetamine: 20 ng/ml. Benzoylecgonine: 50 ng/ml. Carboxy-THC: 20 ng/ml. Methamphetamine: 20 ng/ml. Morphine: 20 ng/ml. Oxazepam: 50 ng/ml ( 55 )

56 TABLE 2F Item 2 Webcode NC99QT NTCXQW PUJKWN RD2H36 TVG9HF U26DNN UPQ9NK UVYG6N UX3CQD VE3L63 WF4M2M WL9JLM XANPDY XLZXGQ XVNN2X XY9RWA YLZJDH ZGHQT3 Item 2 - Comments Internal Standard: Mepivacaine. Indicated but not reported: Chlorpheniramine and EDDP. Amphetamine/Methamphetamine/Morphine/C-THC: 20ng/mL, BE/Oxazepam: 50ng/mL ELISA screening panel includes: amphetamine, benzodiazepines, cannabinoids, carisoprodol, cocaine and metabolites, methadone, opiates, oxycodone/oxymorphone, phencyclidine, and zolpidem. alprazolam-d5 used as internal standard. LOD/LOQ for alprazolam is 5 ng/ml. methadone-d3 used as internal standard. LOD/LOQ for methadone is 10 ng/ml. Flurazepam as Internal Standard INTERNAL STANDARD USED FOR ANALYSIS: PHENOBARBITAL D5. INTERNAL SATDARD USED FOR HPLC/UV-DAD ANALYSIS : DEXTROPROPOXYPHENE. INETRNAL SATANDARD USED FOR ANALYSIS : CODEINE D3 Butyl Acetate - Promazine (ISTD); extra methadone positive control run with second butyl acetate test to verify retention time. Benzodiazepine - Prazepam (ISTD), Opiate - Nalorphine (ISTD) Alprazolam: Internal Standard: Alprazolam-d5, LOD/LLOQ: 5 ng/ml. : Internal Standard: -d9, LOD/LLOQ: 10 ng/ml Prazepam Internal Standard, Major ions of EDDP detected Screening test only performed. Internal standard: estazolam. LCMSMS Internal Standard - mepivacaine. GCMS Internal Standard - mepivacaine. LCMSMS Quantitative internal standard (alprazolam) - diazepam-d5. LCMSMS Quantitative internal standard (methadone & 2-ethylidene-1,5-dimethyl-3,3- diphenylpyrrolidine (EDDP)) - mepivacaine. 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) lowest calibrator is 0.05 mg/l. EDDP reported lower than the lowest calibrator of 50 ng/ml. Internal standards - mepivacaine and diazepam-d5. Limit of report: Alprazolam = ng/ml. methadone = 25 ng/ml. EDDP (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine) = 3 ng/ml Phenyltoloxamine IRM Heptabarbital IRM EDDP seen not reported. Promazine Internal Standard used for screening. Prazepam Internal Standard used for Benzo extraction. Nalorphine Internal Standard used for Opiate extraction Internal standard used for the GC was Mepivicaine. Internal standards used for the LC were Alpha-Hydroxyalprazolam-D5 and Diazepam-D5. lowest calibrator for 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP) quantitation is 0.05 mg/l (limit of report is mg/l) Internal Standards Alprazolam-D5, EDDP-D3, -D9 LOQ 5ng/ml Alprazolam, EDDP, ( 56 )

57 Screening Results - Item 3 TABLE 3A Item 3 Item Scenario: A 32 year old female was arrested for assaulting a bystander and the arresting officer. The officer reported that she exhibited rigid muscles, slurred speech, and had a blank stare. A blood sample was collected 50 minutes after the arrest. Item Contents and Preparation Concentration: (65 ng/ml) Webcode 2ABGZF 2ATE97 2B8WAQ 2BQRRT 2RALHH 2RCBME 2YBZ4J 3L6ZEK 3LJJYH 3QG3ZG 4DDRFF 4DNZXE 4L4FBH 4LXETP 4VKDKU 69BD8G 6GZC6F 6NK9FG 6VFHDT 6VH8HQ 78TYLQ 7GE4VB 82LJZA 84NQUL 84TRBE 87VYAX 89ZEUH 9CYZ87 9LJJFJ 9RPCAE 9RYXEB A2YURD Screening Results phencyclidine phencyclidine (PCP) [Participant reported that drugs were detected, but did not report the drug class or name] (PCP) and benzodiazepines PCP ( phencyclidine) PCP [No screening results reported.] (PCP) phencyclidine was found in an LCMSMS screen. ( 57 )