Cotinine

Manufactured by Cerilliant

Sourced in United States

Cotinine is a biomarker used for the analysis and detection of nicotine exposure. It is a metabolite of nicotine and can be measured in various biological samples such as blood, urine, or saliva to assess an individual's tobacco use or exposure.

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Cotinine-d3 (6 citations)

6 protocols using cotinine

Nicotine and Cotinine Quantification

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Ethyl acetate (Chromasolv HPLC Grade, Sigma Aldrich), methanol (HPLC Super Gradient Reagent, VWR Chemicals), carbon dioxide (food grade, AGA), ammonium hydroxide (ACS Reagent, Sigma Aldrich), AOAC Method 2007.01 Extraction salts (DisQuE, Waters Corp, Milford USA), AOAC Method 2007.01 clean-up tubes (DisQuE, Waters Corp, Milford USA), nicotine, nicotine-D4, cotinine and cotinine-D3 (all from Cerilliant, Round Rock Texas, USA), and 3’-hydroxycotinine (Toronto Research Chemicals, Ontario Canada).

Wright S.L., Rowe D., Reid M.J., Thomas K.V, & Galloway T.S. (2015). Bioaccumulation and biological effects of cigarette litter in marine worms. Scientific Reports, 5, 14119.

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TSNA Analysis via SPE and HPLC

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SupelMIP™ TSNA (50 mg/3 ml) SPE cartridges were purchased from Sigma Aldrich. Oasis hydrophilic–lipophilic balance (HLB, 30 mg/1 ml) and mixed‐mode cation exchange (MCX, 30 mg/1 ml) SPE cartridges were purchased from Waters (Milford, MA, USA). The NIP material was bought from Biotage (Uppsala, Sweden).
HPLC grade acetonitrile, dichloromethane (DCM), heptane, methanol (MeOH), tetrahydrofuran (THF) and water (H₂O) were purchased from VWR International (Radnor, PA). Ammonium formate (97%) and orthophosphoric acid (85% wt.) were purchased from Sigma Aldrich (St Louis, MO, USA) and ammonium acetate and triethyl amine (TEA) were purchased from Fisher Scientific (Waltham, MA, USA). Methyl tert‐butyrl ether (MtBE) was purchased from Honeywell (Charlotte, NC, USA). Cotinine (99.7%), Cotinine‐d3 (99.8%), morphine (99.7%), nicotine (99.6%) and NNN (99,8%) were purchased from Cerilliant (Round Rock, TX, USA).

Mulder H.A., Pearcy A.C, & Halquist M.S. (2022). Characterization of molecularly imprinted polymers for the extraction of tobacco alkaloids and their metabolites in human urine. Biomedical Chromatography, 36(6), e5361.

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Certified Reference Standards for Drugs

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Certified reference standards of 6-monoacetylmorphine, alprazolam, amphetamine, buprenorphine, benzoylecgonine, carisoprodol, chlordiazepoxide, clonazepam, cocaine, codeine, cotinine, dihydrocodeine, diazepam, EDDP, fentanyl, flurazepam, hydrocodone, hydromorphone, lorazepam, MDA, MDEA, MDMA, methamphetamine, meperidine, morphine, methylphenidate, methadone, naloxone, norbuprenorphine, nicotine, nordiazepam, norfentanyl, noroxycodone, alpha-hydroxyalprazolam, oxazepam, oxycodone, oxymorphone, tapentadol, temazepam, ll-nor-9-Carboxy-Δ9-THC, and cis-tramadol were purchased, each at a concentration of 1 mg/mL, from Cerilliant Corporation (Round Rock, TX). Isotopically labeled internal standards (IS) morphine-D3, codeine- D6, hydrocodone-D3, 6-monoacetylmorphine-D6, hydromorphone-D6, oxymorphone-D3, noroxycodone-D3, and naloxone-D5 were purchased, each at a concentration of 100 μg/mL, from Cerilliant Corporation (Round Rock, TX). Drug-free urine was obtained from Bio-Rad Laboratories (Irvine, CA). High-performance liquid chromatography (HPLC) grade water, methanol, and formic acid were purchased from Fisher Scientific (Pittsburgh, PA) and ammonium formate was purchased from Sigma Aldrich (St. Louis, MO).

Pablo A., Breaud A.R, & Clarke W. (2019). Automated Analysis of Dried Urine Spot (DUS) Samples for Toxicology Screening. Clinical biochemistry, 75, 70-77.

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Quantitative Analysis of Nicotine and Cotinine in Biological Samples

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Nicotine was purchased from MilliporeSigma (St. Louis, MO), and nicotine-d₄, cotinine, and cotinine-d₃ were purchased from Cerilliant (Round Rock, TX). Water (Optima grade), acetonitrile (Optima grade), and acetic acid (Optima grade) were purchased from Fisher Scientific (Hampton, NH). PBS and ALI media were prepared (ALI media protocol is given in Supplementary Information) and used in pre-exposure HTBEC smoke exposure experiments as an apical lavage and basolateral cell media, respectively, to provide true matrix blanks for method development. Calibrators for nicotine, cotinine, and isotopically labeled standards were prepared in acetonitrile. To 60 μL of pre-exposure PBS or ALI sample, 10 μL of each standard was added and vortexed (30 s) for final nicotine and cotinine concentrations ranging from 10 to 1500 ng/mL and isotopically labeled standard concentrations of 1000 ng/mL. Nicotine and cotinine in pre-exposure PBS and ALI were used directly in PS-MS/MS. Nicotine calibrators for GC-MS/MS were prepared by adjusting the pH to 13 by addition of 2.5M NaOH followed by liquid-liquid extraction of PBS and ALI samples with 1 mL of 30:30:40 methyl tert-butyl ether:dichloromethane:ethyl acetate, sample drying, and reconstitution in 80 μL of acetonitrile.²⁰

Keating J.E., Minges J.T., Randell S.H, & Glish G.L. (2017). Paper Spray Mass Spectrometry for High-Throughput Quantification of Nicotine and Cotinine. Analytical methods : advancing methods and applications, 10(1), 46-50.

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Quantitative Analysis of Cotinine and Trace Elements

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High-performance liquid chromatography (HPLC)-grade acetonitrile (ACN) and methanol (MeOH) were obtained from TEDIA (Ohio, USA). Nitric acid (GR, approximately 68%) and perchloric acid (GR, approximately 72%) were purchased from Nanjing Chemical Reagent Co., Ltd. (Nanjing, China). Ultrapure water was produced using a water purification machine from a Millipore purification system (Millipore, MA, USA). Analytical standard cotinine (purity > 98%) and isotopic internal standard cotinine-d3 (purity > 98%) were purchased from Cerilliant (Texas, USA). Sodium standard (Na, 1 mg/mL), potassium standard (K, 0.1 mg/mL), calcium standard (Ca, 1 mg/mL), magnesium standard (Mg, 0.1 mg), iron standard (Fe, 1 mg/mL), and zinc standard (Zn, 1 mg/mL) were purchased from the National Research Center for Standard Materials (Beijing, China). Lanthanum (III) oxide (purity ≥ 99.99%) was supplied by Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). Cesium chloride (purity ≥ 99.99%) was obtained from Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). Standard compounds were stored following to the supplier’s recommendations. Stock solutions of individual compounds were prepared in water or MeOH, and the calibration solutions were diluted from individual stock solutions.

Xu L., Lu Y.T., Wu D.F., Li X., Song M., Hang T.J, & Su M.X. (2023). Application of the metal ions as potential population biomarkers for wastewater-based epidemiology: estimating tobacco consumption in Southern China. Environmental Geochemistry and Health.

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Optimized LC-MS Workflow for DOA

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Optima LC/MS grade methanol (MeOH), LC/MS grade 0.1% formic acid in water, LC/MS grade formic acid ampules, optima grade dichloromethane (DCM), optima LC/MS grade isopropanol (IPA), hydrochloric acid HCl, tert-butyl methyl ether, (MTBE), and hexane were purchased from Fisher Scientific. Ammonium hydroxide (NH4OH was purchased from Sigma. DOA analytical standards: (+/-) amphetamine, (+/-) methamphetamine, cocaine, benzoylecgonine, morphine, phencyclidine, (-) delta9-THC, (+/-) amphetamine -D5, (+/-) methamphetamine -D5, cocaine -D3, benzoylecgonine -D3, morphine -D3, phencyclidine -D5, and (-) delta9-THC -D3 were obtained as 1mg/mL solutions from Cerilliant (Round Rock, TX, USA). Interfering substances: Diphenhydramine HCl, Alprazolam, Dihydrocodeine, S-Nicotine, Caffeine, Cortisol, -Cotinine, Dextromethorphan, Ibuprofen, Naproxen, Prednisone, R-Pseudoephedrine, Amobarbital, Propranolol, Nicotinamide, Carbamazepine, Clobazam, Clonazepam, Valproic Acid, Verapamil, and Sertraline were purchased from Cerilliant (Round Rock, TX, USA). Stabilization buffer was purchased from Immunalysis (Pomona, CA). The Raptor Bipheyl LC column; 2.7um, 100mm x 2.1mm was purchased from Restek (Bellefonte, PA). Isolute SLE+ columns were purchased from Biotage (Charlotte, NC). The PRESSURE+ 48 positive pressure manifold was obtained from Biotage (Charlotte, NC).

Howard M.K., Russo P., Haymond A., Ortiz V.C., Andes S.R., Espina V., Luchini A., Liotta L.A., & Guirguis R. (2020). LC-MS/MS Validation for Drug of Abuse Testing Utilizing a Split Sample Oral Fluid Collection System.

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