Cotinine Research Articles

Changes in Cotinine Levels During Pregnancy

Changes in Cotinine Levels During Pregnancy

A New Gas Chromatography–Mass Spectrometry Method for Simultaneous Determination of Total and Free trans-3′-Hydroxycotinine and Cotinine in the Urine of Subjects Receiving Transdermal Nicotine

trans-3'-Hydroxycotinine (THOC) has been recognized as the most abundant metabolite of nicotine. In an attempt to assess THOC and cotinine (COT) concentrations during nicotine transdermal therapy, we developed a new quantitative gas chromatography-mass spectrometry (GC-MS) method for simultaneous determination of total and free THOC and COT in human urine. The method utilizes the following: (a) hydrolysis of conjugated THOC and COT by beta-glucuronidase; (b) basic extraction of THOC and COT with mixed dichloromethane and n-butyl acetate; (c) derivatization of THOC with bis(trimethylflurosilyl)acetamide; and (d) separation and identification by GC-MS with selective ion monitoring. Lower limits of quantification for the assay were 50 and 20 microg/L for THOC and COT, respectively. The intra- and interassay CVs were 4.4% and 11% for THOC, and 3.9% and 10% for COT at 1000 microg/L. The results from six consecutive 24-h urine collections in 71 subjects administered daily transdermal nicotine doses of 11, 22, and 44 mg showed that, on average, free THOC was 76% of total THOC and free COT was 48% of total COT in all subjects. THOC is the major metabolite of nicotine and constitutes 20% of total nicotine intake at steady state, whereas urinary nicotine and COT excretion were 8% and 17%, respectively. The method is useful for simultaneous determination of free and total THOCand COT and can be used to assess the urinary excretion of these metabolites during transdermal nicotine therapy.

Development of a simple and rapid elisa of urinary cotinine for epidemiological application.

Highly stereospecific polyclonal antibodies (anti-CN) to cotinine (CN), a major metabolite of nicotine, were prepared from rabbit antisera to CN-linked keyhole limpet hemocyanin (KLH) by removing the antibodies to KLH and to its binding regions of CN. This was achieved by using immunoadsorbents consisting of insolubilized KLH onto CNBr-activated-Sepharose 4B. A new simple and rapid enzyme-linked immunosorbent assay (ELISA) of urinary CN was developed using the anti-CN. A brief outline of the method is as follows: CN-bovine thyroglobulin complex is coated onto wells of microtiter plates (1 ng/well), and then aliquots of urine samples or standard CN solutions were added followed by appropriate dilution of the anti-CN. The bound anti-CN antibodies are quantified spectrophotometrically with horseradish peroxidase-labelled anti-rabbit IgG and 2,2'-azino-di (3-ethylbenz- thiazoline)-6-sulfonic acid. Measurement of CN concentration in urine samples can be read off on a calibration curve drawn by using standard CN solutions. The standard curve ranged from 1 ng to 4 μ g /ml with an estimated lower limit of sensitivity of 7-8 ng/ml, resulting in within/between-assay CV (coefficient of variation) of lower than 10%. The method allowed one to assay more than 40 samples in duplicate by using just one plate, and is thus easily applicable to epidemiological investigations into exposure status to tobacco smoke.

The in-vitro effects of nicotine and cotinine on sperm motility

Swim-up spermatozoa from the seminal samples of non-smokers, usually not exposed to passive smoking, were treated in vitro with nicotine (NIC) and cotinine (COT) at the average levels found in smokers' seminal plasma and at levels 500 times higher than this average. This was done to evaluate the action of these drugs on sperm motility. Each sample was allowed to swim up in Tyrode's solution with or without the drug; the study was carried out at time 0 and +1, +2, +4, +8 and +24 h of incubation, using a light microscope and a CASA system (experiment 1). In addition, the direct action of smoke on spermatozoa was studied using aspirated cigarette smoke (experiment 2). Kinetic parameters were then measured at 30 min, 45 min and 60 min starting from the last smoke injection. The first experiment showed that NIC and COT at average levels did not produce statistically significant variations of the kinetic parameters studied up to 24 h. However, the much higher concentration significantly altered all the kinetic variables in relation to the time of incubation. The second experiment with smoke in toto demonstrated a sharp reduction in all the sperm kinetic parameters. This reduction was seen after 30 min exposure to smoke and increased progressively until almost complete immotility at 1 h of exposure. These results suggest that NIC and COT are not responsible for the harmful effects of cigarette smoke on sperm kinetic parameters reported in the literature.

Interference of Nicotine Metabolites in Cotinine Determination by RIA

Cotinine (COT), a major metabolite of nicotine (NIC), has been used as a biomarker in many studies of active and passive smoking (1)(2)(3)(4). Methods of analysis for COT in biological fluids include gas chromatography, gas chromatography–mass spectrometry, HPLC, HPLC–mass spectrometry, and immunoassays (5)(6)(7)(8)(9)(10). In particular, the RIA developed for COT and NIC by Langone and Van Vunakis (11) was the method reported by the International Agency for Research on Cancer (IARC) for determination of these analytes in several biological matrices (12)(13). In the description of the assay (11), no mention was made about cross-reactivity of anti-NIC and anti-COT antibodies with trans -3′-hydroxycotinine (THOC) and cotinine glucuronide (COT-G). Indeed, these two compounds were found to be the most abundant metabolites of NIC in urine of smokers (14). Other studies, however, have investigated the cross-reactivity of the anti-COT antiserum with THOC and other metabolites in an ELISA (15). The objective of this study was to investigate the cross-reactivity of the anti-COT and anti-NIC antisera with THOC and COT-G in the RIA (11)(12)(13). We also used urine samples from active and passive smokers to compare data from the RIA with data obtained by HPLC. The rabbit antiserum, (−)[3H]COT, and (−)[3H]NIC were …

In-vitro Effect of Some Common Xenobiotics on the Activity of Human Erythrocyte Pi Class Glutathione S-Transferase

It has been suggested that human erythrocyte anionic glutathione S-transferase rho (GST-p) may be used as a marker of chemical exposure. In the present study, the activity of GST-p in human erythrocyte was measured after incubation of the erythrocytes with some common xenobiotics, and since it has also been suggested that human erythrocyte GST-p and human placental GST-π are identical, the effect of nicotine on purified human placental GST was also determined. Nicotine and cotinine caused an inhibition of the erythrocyte enzyme. There were no effects of malathion, monocrotophos or endosulfan on the erythrocyte enzyme activity. Acrolein, N-ethylmaleimide and styrene oxide, were confirmed as inhibitors of the enzyme.

女子短大生の家庭における父の喫煙習慣と母及び娘の尿中ニコチン排せつ量との関連について

In order to assess the effect of passive smoking at home, 39 female junior college students and their mothers were surveyed concerning paternal smoking habits and asked to provide early morning urine samples in which the concentrations of cotinine (COT) and hydroxyproline (HOP) were measured. The concentration of urinary COT per gm creatinine (COT/Cr) of smoker was 1, 000 μg/g or more. There was no correlation between the number of cigarettes smoked per day by the father and the concentration of COT/Cr of either the mother or daughter. Their levels of COT/Cr remained 0-30μg/g irrespective of the father's smoking. The cases in which no COT was detected in either daughters or mothers were actually more frequent with heavier paternal smoking. There was also no relationship between the father's smoking and the concentration of HOP/Cr of the daughters and their mothers. One possible explanation of these findings may be that fathers who smoke at home do so outside of the room. Smoking by mothers was also found to not increase the concentration of COT/Cr of their daughters or rather low level of 5 μg/g. Thus, passive smoke at home did not show any effects on nicotine metabolite of daughters and mothers. The COT/Cr levels of daughters did increase significantly by 61 percent from 11.0 μg/g, as the result of a one hour exposure to heavy smoke in a meeting room.

Nicotine, Cotinine, and trans-3-Hydroxycotine Levels in Seminal Plasma of Smokers

Sperm samples from 44 cigarette smokers and 50 nonsmokers attending an infertility clinic were examined by high-performance liquid chromatography (HPLC) assay and HPLC-mass spectrometry for the presence of nicotine (NIC), cotinine (COT), and trans-3'-hydroxycotinine (THOC) in seminal plasma. Smokers were found to have levels of COT and THOC in seminal plasma that were similar to those found in serum. The level of NIC was significantly increased in seminal plasma compared to serum. Total motility of spermatozoa was significantly and negatively correlated to COT and THOC levels in seminal plasma. Forward motility of spermatozoa was correlated only with cotinine semen levels. On the basis of these results, we suggest that the presence of tobacco smoke constituents in seminal plasma could provide a warning of the adverse effects of cigarette smoke on the physiology of reproduction.

Limited Value of Urinary Cotinine to Creatinine Ratio as an Indicator for Tobacco Smoke Exposure

Usefulness of urinary cotinine (COT)/creatinine (Cr) ratio, a widely accept practical indicator of tobacco smoke uptake, was evaluated in 4 groups of healthy male volunteers (21 in total) with different cigarette doses of 0, 10, 20 and 30 cigarettes per day. 24-hour urine samples were collected on the 7th day of successive controlled smoking to determine urinary COT, thiocyanate (SCN) and Cr concentrations. Although urinary COT concentration exhibited close correlation (r=0.91, p<0.01) with cigarette dose, this correlation was weakened to r=0.82 (p<0.01) when it was standardized by urinary Cr concentration. Urinary SCN concentration was, in contrast, unrelated to cigarette dose (r=0.09), which however become correlated (r=0.71, p<0.05) after Cr standardization, in part due to the variation of urinary Cr concentration which was found inversely linked to cigarette dose (r= -0.47, p<0.05). Multiple regression analyses revealed that extent of urine volume-dependent decrease in urinary COT concentrationwas of several orders smaller in magnitude than that of urinary SCN or Cr concentrations as estimated on the basis of standardized partial regression coefficients. These results suggested that urinary COT/Cr ratio, compared to urinary COT concentration, tends to exaggerate the actual smoke uptake in the individuals with increased urine flow as well as in the smokers with smoking-associated decrease in urinary Cr concentration. J Epidemiol, 1992; 2 : 91-95.

Thiocyanate vs cotinine as a marker to identify smokers.

Thiocyanate vs cotinine as a marker to identify smokers.

Effects of cotinine on the prostacyclin and thromboxane byosynthesis

Effects of cotinine on the prostacyclin and thromboxane byosynthesis

Demethylation of Cotinine in Vivo1

Demethylation of Cotinine in Vivo¹