METHOD AND TEST STRIP OF DETECTING OXIDIZING ADULTERANT IN URINE

Abstract: A single reagent system and a method to detect and measure oxidizing adulterants in bodily fluid being screened for drugs of abuse are disclosed. The system comprising a strip containing 0.05 to 0.2 micromole/25 sq. mm. of a benizidine derivative and is used to detect sodium hypochlorite (bleach), chlorine, hydrogen peroxide, sodium bromide, sodium iodide, sodium nitrite, and pyridinium chlorochromate adulterants in urine, sweat, saliva, blood or other bodily fluids during screening for drugs of abuse.

Description: METHOD AND TEST STRIP OF DETECTING OXIDIZING ADULTERANT IN URINE Field Of The Invention The present invention relates to a single reagent system and method designed to detect and measure oxidizing adulterants in urine, sweat, saliva, blood or other bodily fluids being screened for drugs of abuse. Specifically, the present invention is direct to detect and measure sodium hypochlorite (bleach), chlorine, hydrogen peroxide, sodium bromide, sodium iodide, sodium nitrite and pyridinium chlorochromate adulterants in urine during screening for drugs of abuse. Background Of The Invention As the use of illicit drugs in public, workplace, sports and the like has grown, public concern for the health and safety of individuals and the negative impact of such drugs use on productivity of industry has grown as well. Such concern obligated the use of analysis of urine, sweat, saliva, blood, or other bodily fluids, as a way to detect and defer drug use. Such testing for drugs of abuse in industry 6r prospective and current employees, particularly government employees, military personnel, professional and amateur athletes, truck drivers, pilots, as well as people under supervision of the criminal justice system, has become a routine practice. Because of the significance of a positive result in such testing commonly performed by examining urine, sweat, saliva, blood, or other bodily fluids sample, the testing procedure must withstand vigorous scrutiny. A positive test result of screening for drugs of abuse can have serious impact on the life of a person being tested. The incentive for a user to alter the test specimen is high. The users of drugs of abuse have developed ways to adulterate the collected specimen in an attempt to produce a false negative result in the drug-screening test being conducted. A drug user may affect the test results by dilution- reduce the drug concentration in the urine sample; substitution- with liquids such as clean (drugfree) urine, apple juice, tea, soda for the drug containing specimen; or adulteration- addition to the urine specimen of foreign substances in an attempt to invalidate the test. Such adulterations of urine sample can be achieved by the addition of several readily available agents, including household products; i.e. bleach (sodium hypochlorite), chlorine, sodium bromide, sodium iodide, hydrogen peroxide, pyridinium chlorochromate, finger nail polish remover, Visine, soap, together with commercial available adulteration products, such as 'URINE-AID®" (gluteraldehyde), and 'KLEA®" (sodium nitrite). Additionally, drug users may eliminate some drugs rapidly from their bodies by altering their urinary pH. This includes using NH4CI to hasten the elimination of phencyclidine or amphetamines from the bodies. While the use of some in vitro adulterants can be eliminated by the direct observation of the test subject during the collection process, such direct observation is often deemed unacceptable. Such adulteration can affect the four commonly used methods for drugs of abuse, namely: enzyme immunoassay (EIA or EMIT), lateral flow immunoassay (LFIA), radioimmunoassay (RIA) and florescent polarization immunoassay (FPIA). Thus, clinical chemistry literature recommends that adulterants be tested prior to testing of drugs of abuse in urine samples. Accordingly, a need exists for providing an easy and convenient manner by which to make a determination of the presence of oxidizing adulterants in urine samples prior to the testing for drugs of abuse. Particularly, the need exists to provide an easy and convenient manner to detect bleach, chlorine, sodium bromide, sodium iodide, hydrogen peroxide, sodium nitrite, and pyridinium chlorochromate in urine samples prior to the testing for drugs of abuse. Description Of The Prior Art The patent to Ehrenkranz discloses a urine sample collection apparatus having ability for detecting the presence of adulterants, such as bleach, intentionally added into the urine sample to tamper or mask urine test results. The apparatus is provided with an enzyme/bleach poison detector or strip having an indicator formed from glucose oxidase, peroxidase and potassium iodide. The patent to Smith discloses a method of detecting adulterants, such as bleach and gluteraldehyde, in urine samples through the use of an indicator containing essentially phenylhydrazine. The patents to Ramana et al; Serrat; and Wu all disclose the specific use of tetramethylbenzidine as an indicator on a test strip (or in a kit) for detecting the presence of chlorine in water samples. The patents to Gretton et al; Magers et al; and Charlton et al all disclose the use of tetramethylbenzidine or benzidine per se for detecting glucose in body fluids, such as blood or urine. The patents to Pugia and Kardos et al both disclose the use of benzidine- containing compositions per se for detecting peroxidative substances such as blood in urine or other body fluids. All of the above patents had to use very complicated and/or expensive reagent system and are troublesome to get quick and easy test results. The use of an aqueous enzyme system makes it difficult to handle and has a limited shelf life. Summary Of The Invention Accordingly, it is an object of the present invention to provide for a test strip - a single reagent system and a method to tetect and measure oxidizing adulterants in urine or other bodily fluids, being screened for drugs of abuse. In a preferred embodiment, a 0.1 to 0.9 of a reagent-benzidine derivative (including o-tolidine, o-tolidine dihydrochloride, benzidine, benzidine dhydrochloride, 3,5,3',5' tetramethylbenzidine and 3,5,3', 5' tetramethylbenzidine dihydrochloride) is dissolved in 100-300 ml of a pH5 phosphate or acetate buffer and (050 ml) 2.5 percent Gantrez (ISP Technologies Inc. manufacturer of GAF Chemicals). A filter paper such as Whatman #3 type, or a paper of cellulose base, woven or nonwoven, or other material like fiberglass, is then soaked in the solution of 0.1 to 10 minutes. The excess solution is then squeezed out and dried in an oven at 5075 degree C for 10-30 minutes. The amount of the benzidine derivative on the test strip is calculated to be from about 0.05 to 0.2 micromole or 0.15 to 60 microgram per 25 Sq. ml (millimeter). In an alternative embodiment, the process can be accomplished by slipping the paper material in a roll configuration and then use warm air to blow dry the material instead of putting it into an oven to dry. The roll of paper is then cut into the strip for testing purposes. During an analysis, the paper containing the dried reagent is contacted with a liquid, such as urine, to be analyzed. After the liquid has been in contact with the dried reagent for a selected time up to 15 minutes, the paper containing the dried reagent is evaluated for detecting color change. For a solution containing 0.5 to 20% (by volume) CHLOREX® (sodium hypochlorite-bleach) and iodide, a blue color is detected. For a solution containing 0.05 to 10% (by weight) pyridinium chlorochromate, a reddish-brown color is detected. For a solution of containing 10 to 30% (by weight) hydrogen peroxide, a light blue color is detected. For a solution containing 0.01 to 10% (by weight) of sodium nitrite, a very dark blue color is detected. Other objects and advantage of the invention will become apparent from the following detailed disclosure. Brief Description Of The Drawings The novel features which are believed to be characteristic of the invention are set forth with particularity in the appended claims. The invention itself, however, both as to its organization and method of operation, may best be understood by reference to the following description, taken in connection with the accompanying drawing in which: Fig. 1 is a perspective view of a test strip showing the 3,5,3'5' tetramethylbenzidine reagent for detecting oxidizing adulterants in urine. Fig. 2 is a perspective view of a commercial INTECT™test strip for adulterants detection with tests for Creatinine, Nitrite, Glutoraldehyde, PH, Bleach and Specific Gravity. Detailed Description Of The Invention Referring now to FIG.1 , a test strip 1 used to detect oxidizing adulterants in urine is prepared by using about 0.5 g of the reagent-3,5,3',5' tetramethylbenzidine (TMB) dissolved in about 200 ml of a pH 5 phosphate or acetate buffer and about 2.5% Gantrez. A Whatman #3 type filter paper is then soaked in the solution for about 10 minutes. The excess solution is then squeezed out and dried in an oven at about 60 degree C for about 15 minutes. The urine for the testing is collected in a clean plastic container. No centrifuging or preservative is added to the urine sample. The paper containing the dried reagent TMB 2 is contacted with the urine to be analyzed. After the urine has been in contact with the dried reagent for about 1-5 minutes, the paper containing the dried reagent is evaluated for detecting color change. EXAMPLE 1 - Detection of Sodium Hypochlorite (bleach) A one-volume part of aqueous 5.25% by weight sodium hypochlorite solution is diluted with the indicated volume parts of normal urine. Test strip was white prior to immersion into the testing solution. Readings were taken about 1 minute after immersion. Dilution 1/10 1/20 1/50 1/100 1/200 1/500 Color Very dark Dark blue Blue Light blue Very light white blue blue EXAMPLE 2 - ■ Detection of Pyridinium Chlorochromate The following concentrations were percentage by weight (w/w) in urine. Readings were taken 1 minute after immersion. Cone. 10% 5% 1 % 0.5% 0.1% 0.05% Color Reddish Brown Very dark Dark blue Light blue white Brown blue EXAMPLE 3 - Detection of Iodine and Sodium lodate The starting solution contains 2% iodine and 2.4% sodium iodate in 47% of ethanol. Dilutions are made by diluting indicated volume with volume of normal urine. Readings were taken 1 minute after immersion. Dilution 100% 50% 25% 18% 12% Color dark blue dark blue deep blue purple white EXAMPLE 4 - Detection of Hydrogen Peroxide The following concentrations were obtained by diluting certain volume part of aqueous 30% by weight of hydrogen peroxide solution with indicated volume parts of normal urine. Test strip was white before immersion into the testing solution. Readings were taken 5 minutes after immersion. Dilution 2 parts/1 part 2 parts/1 part 2 parts/1 part H2O2/urine H202/urine H2O2/urine Color light blue slight blue white EXAMPLE 5 - Detection of Sodium Nitrite The following concentrations were percentage by weight (w/w) in urine. Readings were taken 1 minute after immersion. Cone. 10% 5% 1% 0.5% 0.1% 005% 0.01% Color very dark dark blue light light very light white blue blue blue blue blue Referring now to FIG. 2, a commercially available INTECT™ strip 10 is shown. The test strip can be used to test for adulteration in urine for Creatinine (3), Nitrite(4), Glutoraldehyde (5), PH (6), Specific Gravity (7), and Bleach (2).

Claims: What is claimed is: Having described our invention, what we claim and desire by letter patent is: 1. A method for the detection of oxidizing adulterants in a bodily fluid comprising: (a) obtaining an aliquot of urine from an unknown sample; (b) contacting said urine with a test strip comprising a benzidine derivative; (c) performing color change observation on said test strip. 2. The method according to claim 1 wherein the benzidine derivative is in a range of 0.05 to 0.2 micromole/25 sq. mm. 3. The method according to claim 1 wherein the benzidine derivative is selected from the group consisting of o-tolidine, otolidine dihydrochloride, benzidine, benzidine dihydrochloride, 3,5, 3', 5' tetramethylbenzidine and 3, 5,3', 5' tetramethylbenzidine dihydrochloride. 4. The method according to claim 1 wherein the oxidizing adulterant is selected from the group consisting of sodium hypochlorite, chlorine, hydrogen peroxide, sodium bromide, sodium iodide, pyridinium chlorochromate and sodium nitrite. 5. The method according to claim 1 , wherein the bodily fluid is selected from the group consisting of urine, sweat, saliva and blood. 6. The test strip for the detection of oxidizing adulterants ina bodily fluid comprising 0.05 to 9.2 micromole/25 sq. mm. Of 3,5,3', 5' tetramethylbenzidine. 7. A method for the detection of oxidizing adulterants in a bodily fluid comprising: (a) obtaining an aliquot of urine from an unknown sample. (b) contacting said urine with a test strip comprising 3, 5, 3', 5' tetramethylbenzidine; (c) performing color change observation on said test strip. 8. The method according to claim 6 wherein the 3,5,3', 5' tetramethylbenzidine is in a range of 0.05 to 0.2 micromole/25 sq. mm. 9. The method according to claim 6 wherein the oxidizing adulterant is selected from the group consisting of sodium hypochlorite, chlorine, hydrogen peroxide, sodium bromide, sodium iodide, pyridinium chlorochromate and sodium nitrite. 10. The method according to claim 6, wherein the bodily fluid is selected from the group consisting of urine, sweat, saliva and blood. 11. A test strip for the detection of oxidizing adulterants in a bodily fluid comprising 0.05 to 0.2 micromole/25 sq. mm. Of 3,5,3',5'tetramethylbenzidine.

Inventors: CHEN SHUENN TZONG, CHEN, SHUENN, TZONG
Applicants: BRANAN MEDICAL CORP, BRANAN MEDICAL CORP.
Application year: 2001
Main classification: G01N33
IPC codes: G01N33