j. Cosmet. Sci., 52, 297-311 (September/October)2001 Evaluation of a new X-ray fluorescent analysis technique for the creation of a Nordic hair database: Elemental distributions within the root and the virDin seDment of hair fibers B. STOCKLASSA, M. ARANSAY-VITORES, G. NILSSON, C. KARLSSON, D. WIEGLEB, and B. FORSLIND, Cox Analytical Systems AB, Gothenburg (B.S.), National Laboratory of Forensic Science, Linkb}Oing (M.A.-V., G.N, C.K.), and EDRG, Department of Medical Biophysics, Karolinska Institute, Stockholm (D. W., B.F.), Sweden. Accepted for p•blication May 31, 2001. Synopsis A new, non-destructive X-ray fluorescence technique for quantitative estimation of elemental content in biological tissues has been developed. Technical and instrumental characteristics of the ITRAX X-ray spectrometer have been evaluated in relation to the properties of biological samples, i.e., human hair fibers. Thus, attenuation variations of the fluorescent X-rays in the hair bulk mass were demonstrated by analysis of sulfur, calcium, and zinc in a virgin part near the root of one hair fiber with elliptical cross section. By rotation of the hair fiber and successive analyses made of the same part of the hair fiber, the results showed that concentrations of elements varied as functions of the diameter of the analyzed hair volume. Other sources of errors are also discussed. The ITRAX instrument allows for precise, fast, non-destructive, simultaneous, quantitative recording of the detected elements and trace elements down to levels of 1 ppm (l•g/g). It was used for assessment of normal values of physiologically important elements present in hair in a cohort of normal, healthy Swedish, Caucasian individuals. The database constructed from data retrieved from a conceivably homogenous ethnic set of individuals represents, to our knowledge, the first of its kind. INTRODUCTION In recent years an increasing interest in hair research has evolved, partly as a result of the introduction of pharmaceutical agents (minoxidil, finasteride) promoting hair growth in certain individuals with inherited androgenic alopecia (AnA). In addition, the "green wave" of dietary consciousness has resulted in a layman's interest in trace elements and their proposed beneficial properties to the status of health. Assessing the content of trace elements in hairs allegedly reflecting the health of an individual has become a particular mercantile niche. A great number of commercial laboratories have appeared that provide 297

298 JOURNAL OF COSMETIC SCIENCE customers with analyses of hair fibers using sophisticated techniques that are accurate methods per se. The basis for an analysis of such data remains an uncertain one as yet (cf. refs. 9-11). From the biologist's/physician's point of view there are two main objections to such analyses. The first concerns the manner in which hair is collected for analysis. Generally this has not taken the state of the hair cycle or the scalp area of origin into consideration i.e., the analyses are made on bulk specimens rather than on selected hair fibers in the anagen (growing) phase of the hair cycle. If the analysis data are to be related to the physiologic history of the proband, such as is the case in lawsuits and forensic applica- tions, the timing of ingestion of (a) specific element(s) can be deduced from the distance to the hair root, since hair growth in normal individuals is relatively constant (approxi- mately 0.4 ram/day). Using bulk specimens would mean that we lose this kind of information. The increased mass of the specimen will not speed up the analysis process, which is a matter of a few minutes with single fibers, and adds the problem of correction for self absorption, which is virtually nil with single-fiber analysis. The second is that there is presently no reference material publicly available for comparison of individual data. In addition, ideally such quantitative elemental data from hair fibers should be correlated to quantitative data from blood analyses of the same individuals. Furthermore, there is a need for the establishment of precision and accuracy in single-fiber measure- ments. These topics have been scrutinized in the past (5,14), but the effect of such analyses and discussions does not seem to have influenced the commercially interested parties. In forensic applications, hair fibers collected during crime investigations are often ex- amined for morphological characteristics, and sometimes also for cosmetic treatments, chemical analysis of shampoo residues (1), and DNA or enzyme analysis. In some instances such analyses allow a specific hair fiber to be assigned to a certain individual. However, only in a minor number of cases are such characteristics distinctive enough to be used as conclusive evidence. With the development of new DNA techniques, hair fibers can sometimes be identified as to their origin by comparative analysis, being a reason for replacement of enzymatic tests. Elemental analysis has not yet proven to be useful for identification of single hair fibers, but as a method of analysis complementary to that of DNA techniques, it may provide information on subjects related to physi- ological status, residential area (rural/urban), certain occupations (e.g., uptake of ele- ments at a particular time), etc., of specific individuals. A basis for such evaluations must rely on good baseline data of the normal elemental content in hair fibers from a normal population. The present study makes it clear that, when the goal is to assess an individual's physi- ological status, elemental analysis of hair should be made on a virgin part of a single anagen hair fiber, i.e., within the segment 2-5 mm from the root of a plucked hair fiber in order to minimize effects of contamination or leakage out of the hair shaft. However, the question of how representative such a measurement is, as related to a database of normal values that is thought to represent a large population, remains a crucial one. A previous investigation suggested that even the hair fibers from one and the same location in an individual might show conspicuous variations in the concentration of elements such as sulfur (S) and zinc (Zn) (9). Many analytical techniques are essentially bulk techniques requiring at least milligram