The experiments of this proposal will test the hypothesis that accumulation of systemically administered drug molecules involves ionic and covalent biochemical assimilation of the drugs into the structural matrix of the hair, and that this type of accumulation results in much higher concentrations than random mechanical occlusion of drugs during the keratinization process. It is proposed that in many studies to date a significant portion of drugs deposited in hair, by virtue of covalent assimilation into matrix protein, has likely escaped conventional extraction and analytical detection methods.
The specific aims of this proposal will be accomplished using 23 day-old balb or C57 mice entering their second anagen of 24 days duration. This will insure controlled hair growth and allow a determination of the effect of pigmentation on drug deposition.
The aims consist of 1) quantitation of hair growth rate by microscopic measurements, and calculation of hair volume per unit length for expression of drug concentration levels in hair. Water accessible volume will be ascertained with tritiated water. These volumes may define differential compartmentation possible for deposited drugs. 2) As a comparative standard, the hair concentrations of ubiquitous, non-covalently interacting species will be determined following 1 or 5 days administration of Cl-, Ca++ or [14C]-urea. Temporal deposition of radioactivity in the hair will be determined by autoradiography for the anagen period. 3) The hair concentrations of heavily accumulated, covalently binding [35S]-cysteine will be examined. 4) With these guidelines established, the hair concentrations resulting from 1 or 5 day administration of several radiolabeled model drugs will be examined. These amines, quaternary ammonium compounds and amino esters, strategically labeled at various sites to detect hydrolysis of the drug, have the capability to react ionically or covalently with several functionalities of matrix proteins. 5) The efficacy of a number of wash, extraction and digestive procedures at removing the deposited radioactivity will be evaluated and compared. 6) The efficacy of these wash and extraction procedures will be examined after external application of the same array of radiolabels in control hair. 7) Non-removable radioactivity will support the covalent association of drug molecules with structural elements of the hair. This will be examined by 2-dimensional gel electrophoresis of S-carbamoylmethylated keratins. By accounting for the disposition of total radiolabeled drug, the differences in storage mechanisms for systemically accumulated versus externally applied drugs can readily be discerned.
|Dehn, D L; Claffey, D J; Duncan, M W et al. (2001) Nicotine and cotinine adducts of a melanin intermediate demonstrated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Chem Res Toxicol 14:275-9|
|Claffey, D J; Stout, P R; Ruth, J A (2001) 3H-nicotine, 3H-flunitrazepam, and 3H-cocaine incorporation into melanin: a model for the examination of drug-melanin interactions. J Anal Toxicol 25:607-11|
|Claffey, D J; Ruth, J A (2001) Amphetamine adducts of melanin intermediates demonstrated by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Chem Res Toxicol 14:1339-44|
|Stout, P R; Ruth, J A (2000) Histologic localization of serum constituents, (45)Ca(2+), (36)Cl(-), [(14)C]urea, and [(35)S]cysteine in forming hair after systemic administration. Drug Metab Dispos 28:113-7|
|Claffey, D J; Stout, P R; Ruth, J A (2000) A comparison of sodium hydroxide and sodium sulfide digestion of mouse hair in the recovery of radioactivity following systemic administration of [3H]-nicotine and [3H]-flunitrazepam. J Anal Toxicol 24:54-8|
|Stout, P R; Ruth, J A (1999) Deposition of [3H]cocaine, [3H]nicotine, and [3H]flunitrazepam in mouse hair melanosomes after systemic administration. Drug Metab Dispos 27:731-5|
|Stout, P R; Claffey, D J; Ruth, J A (1998) Fentanyl in hair. Chemical factors involved in accumulation and retention of fentanyl in hair after external exposure or in vivo deposition. Drug Metab Dispos 26:689-700|
|Stout, P R; Ruth, J A (1998) Comparison of in vivo and in vitro deposition of rhodamine and fluorescein in hair. Drug Metab Dispos 26:943-8|
|Stout, P R; Dehn, D; Ruth, J A (1998) Deposition and retention of radiolabeled serum constituents in hair after systemic administration. Drug Metab Dispos 26:900-6|