Roland K. Pomeroy,' Nikolajs Drikitis, ond Yoshikata Koga
University o f British Columbia Vancouver, B. c.,Canada V6T 1W5
The Determination of Zinc in Hair Using Atomic Absorption S p e ~ t r 0 ~ ~ 0 p y
The determination of heavy metals in trace amounts has become increasingly important in such fields a s medicine, forensic science, space and pollution research. For this purpose such techniques as atomic absorption spectroscopy, X-ray fluorescent spectroscopy, and neutron activation analysis are commonly used. There is the general trend that while students are learnine the basics of analvtical chemistrv. thev like to see thise techniques applied to a "relevait;' pr&lem. Therefore. we have introduced the aoolication of atomic absorotion spectroscopy to the laborat'ory program of the course, "analytical physical inorganic chemistry" for second year non-chemistry majors. Students were shown a n audio-visual program (1) describing the principles of atomic absorption spectroscopy and its application prior to the operation. Such analyses a s the iron content in commercial cereals and the trace metal content in local sea water have been carried out with some success. Below we describe the use of atomic absorption spectroscopy in the determination of the zinc content in human hair. Since about 400 students take this course each year, a large number of samples can be collected and analyzed. The compilation of the zinc content together with other characterization of samoles (ace. sex. color of hair, etc.), and a statistical analysis mayualso cbntrihute some meaningful results to the actual research field. Introduction to the Experiment
A number of enzymes contain zinc (2), e.g., carbonic anhydrase, alcohol and lactic dehydrogenases, and various peptidases. Studies have shown that zinc depletion increases lethargy and slows learning rates in animals (3). Zinc deficiency in the rat is manifested by retarded growth, loss of hair, and lesions of the skin (4a, 5). Since zinc is widely distributed in foods i t is unlikely that zinc deficiency will occur in human beings eating a n otherwise adequate diet. However, there is a report where a team of university wrestlers were found to have low zinc levels not only in their hair, but also in their red blood cells and urine (6). A group of Iranian dwarfs have been shown to he zinc deficient 146, 5). The analysis of zinc in hair has also been used for the assessment of zinc nutriture (13). Hair consists mainly of the protein keratin which contains a s much as 14% sulfur. However, a number of elements can be detected in trace amounts in hair: Mg, Al, CI, Ca, Cr, Mn, Fe, Co, Cu, Zn, As, Se, Cd, I, Hg, Au (7). The amount of the trace element present in hair can often be a measure of the amount of the element ingested for the period during which growth of the hair took place. This fact has been used to prove arsenic poisoning even many years after the actual time of death. Contamination of hair from external sources is known to take place for some elements (8, 9). This however has not been found to be the case for zinc, even for those people using commercial shampoos containing zinc pyrithione (8, 91. Zinc is present in hair in such concentrations a s to he suitable for determination by atomic absorption spectroscopy. This concentration for a particular individual has also been found to be fairly constant along the length of 544 / Journal of Chemical Education
hair and also from the position on the head from which the sample is taken (7). Although there is some discrepancy in the earlier literature (10) concerning the normal range of the zinc concentration in human hair i t is now well established a s being between 150-200 pglg (7-9, 111. Values greater than 250 hg/g are, however, not uncommon. Studies are underway to correlate the factors affecting the trace metal content in human hair. These factors include such things a s variation with geographical location, diet, age, sex, race, etc. (8, 12,14). Experimental
A sample of clean hair (about 0.3 g) is accurately weighed in a 100-ml Erlenmeyer flask. The hair is dissolved by gently hoiling it with 10 ml of concentrated nitric acid (carriid out in the fume hood). When the volume is reduced to about a half, the solution is cooled and 2 ml of perchloric acid is added. The further digestion is carried out on a hat plate until the volume has decreased to 1-2 ml. After cooling, the solution is made up to 100 ml in a volumetric flask. Among solutions even from the same hair sample, some are of pinkish color while others are colorless. Upon standing, colorless solutions turn to pinkish. What causes this phenomena is not known, hut the values of the zinc content are identical within the experimental error of about *I0 pg/g, independent of the color of solutions. The solution thus prepared normally contains 0.5-1 ppm of zinc. A series of solutions of known zinc concentration covering this range are prepared. This is best done by diluting a more concentrated standard solution. The absorhanees of these solutions are measured using the atomic absorption spectrometer set up for the determination of zinc. A graph of absorbance versus concentration is drawn. The calibration curve is usually straight up to 1.5 ppm. The absorbance of the solution of dissolved hair is measured immediately after the measurement of the standard solutions. The concentration of zinc this corresponds to is determined from the calibration curve. From this value the amount of zinc in the 100 ml of the solution and hence in the original hair sample can be calculated. The above procedure should be carried out at least in duplicate for each hair sample. A blank correction may be applied to the absorbance of the hair solution. However in practice this correction was negligible. Similarly, a more thorough washing of the hair before weighing has been found both in the present experiment and by other workers (7, 9, l l J to have only a very small effect on the results. Thus in a typical experiment by using Unicam Sp 90 spectrometer, 0.29M) g of the sample hair (from male, age 32, brown) when treated as described above had an absorbance 0.223. This corresponded to 0.540 ppm Zn in the solution and hence 183 pg/g of zinc in hair. Similarly 0.3166 g of the same hair gave an absorbance 0.245, which indicated 0.595 ppm of Zn in the solution and hence 188 pglg. The other hair sample (from female, age 12, black) was repeatedly analyzed. The values obtained were 309, 317, 297, 310, 301, 318, and 310 &gig, the average being 309 pg/g and the standard deviation 8pg/g. Acknowledgment
The authors wish to thank Dr. J. B. Farmer for valuable discussion.
' I'resrnt nddrrrs. Departmenr 01' Chmmicrry. Llniversitv of Alberta. Edmontun. Alhrrrn, Cnnado.
Literature Cited (11 Robinson. J. W.. "Principieo of Atomic Absorption." Program 3Wl CornmunieatianSkillCorp.. 1912. (21 White. A.. Handler. P.. and Smith, E. L.. "Principles of Biochemistry." McGrawHill BmkCa.. NewYork. 1968.p. 10L6. 131 Caldwell. D. F., Obnleas. D.. Clancy. J. J., and Pvasad. A. S., k c . Sor. Ezp. BiolMed.. 133, 1717 119701. 141 (4 Prasad. A. S.. "Zinc Metabolism," CC Thomsa, Springfield. Ill. 1966. p. 142: (bl P. 2,W. (51 Hurley, R.S., Ph.D.Theds. UniversityofUtah. 1972. 161 Chem Eng. News, May 1st. p. 17 (19721.
(71 Obrurnik. I.. Gilasan. J . , Maes. D., McMillan. D. K., D'Auria. I., and Pate. B. D.J. Rodimnnl. Chem.. IS. 1151L973);J. ForrnrirSciancea. 17.426119721. 181 Gordus. A , J Rodioonol. Chem., li.229I19731. 191 Verghwe, G. C.. Kisho*. R.. and Gunn. V. P., J . Rodiwnol. Chem., 15. 329
,.".",.
,la??,
110) Porkom. A. K..andJervis. R. E . J . Forensir Sciences. 11.6911966l. (111 HarRaun, W. W. Yuraeck. 1. P., and Benson. C. A,. Clin. Chim. Acto. 23. 83 (19691. 53.71 119691. (12) Schroeder.H.A.. endNason, A.P.. J. Inuerti~aiiueDermnfolopy, (131 Klsvsy, L. M..Arner. J . Clin. Nutrition, 23. 284 119101. (141 Petering. H. G., Yeaper. D. W., and Witherup, S. 0 ,Arch. Enuimn. H d f h . 23, 202 119711.
Volume 52, Number 8, August 1975 / 545
University o f British Columbia Vancouver, B. c.,Canada V6T 1W5
The Determination of Zinc in Hair Using Atomic Absorption S p e ~ t r 0 ~ ~ 0 p y
The determination of heavy metals in trace amounts has become increasingly important in such fields a s medicine, forensic science, space and pollution research. For this purpose such techniques as atomic absorption spectroscopy, X-ray fluorescent spectroscopy, and neutron activation analysis are commonly used. There is the general trend that while students are learnine the basics of analvtical chemistrv. thev like to see thise techniques applied to a "relevait;' pr&lem. Therefore. we have introduced the aoolication of atomic absorotion spectroscopy to the laborat'ory program of the course, "analytical physical inorganic chemistry" for second year non-chemistry majors. Students were shown a n audio-visual program (1) describing the principles of atomic absorption spectroscopy and its application prior to the operation. Such analyses a s the iron content in commercial cereals and the trace metal content in local sea water have been carried out with some success. Below we describe the use of atomic absorption spectroscopy in the determination of the zinc content in human hair. Since about 400 students take this course each year, a large number of samples can be collected and analyzed. The compilation of the zinc content together with other characterization of samoles (ace. sex. color of hair, etc.), and a statistical analysis mayualso cbntrihute some meaningful results to the actual research field. Introduction to the Experiment
A number of enzymes contain zinc (2), e.g., carbonic anhydrase, alcohol and lactic dehydrogenases, and various peptidases. Studies have shown that zinc depletion increases lethargy and slows learning rates in animals (3). Zinc deficiency in the rat is manifested by retarded growth, loss of hair, and lesions of the skin (4a, 5). Since zinc is widely distributed in foods i t is unlikely that zinc deficiency will occur in human beings eating a n otherwise adequate diet. However, there is a report where a team of university wrestlers were found to have low zinc levels not only in their hair, but also in their red blood cells and urine (6). A group of Iranian dwarfs have been shown to he zinc deficient 146, 5). The analysis of zinc in hair has also been used for the assessment of zinc nutriture (13). Hair consists mainly of the protein keratin which contains a s much as 14% sulfur. However, a number of elements can be detected in trace amounts in hair: Mg, Al, CI, Ca, Cr, Mn, Fe, Co, Cu, Zn, As, Se, Cd, I, Hg, Au (7). The amount of the trace element present in hair can often be a measure of the amount of the element ingested for the period during which growth of the hair took place. This fact has been used to prove arsenic poisoning even many years after the actual time of death. Contamination of hair from external sources is known to take place for some elements (8, 9). This however has not been found to be the case for zinc, even for those people using commercial shampoos containing zinc pyrithione (8, 91. Zinc is present in hair in such concentrations a s to he suitable for determination by atomic absorption spectroscopy. This concentration for a particular individual has also been found to be fairly constant along the length of 544 / Journal of Chemical Education
hair and also from the position on the head from which the sample is taken (7). Although there is some discrepancy in the earlier literature (10) concerning the normal range of the zinc concentration in human hair i t is now well established a s being between 150-200 pglg (7-9, 111. Values greater than 250 hg/g are, however, not uncommon. Studies are underway to correlate the factors affecting the trace metal content in human hair. These factors include such things a s variation with geographical location, diet, age, sex, race, etc. (8, 12,14). Experimental
A sample of clean hair (about 0.3 g) is accurately weighed in a 100-ml Erlenmeyer flask. The hair is dissolved by gently hoiling it with 10 ml of concentrated nitric acid (carriid out in the fume hood). When the volume is reduced to about a half, the solution is cooled and 2 ml of perchloric acid is added. The further digestion is carried out on a hat plate until the volume has decreased to 1-2 ml. After cooling, the solution is made up to 100 ml in a volumetric flask. Among solutions even from the same hair sample, some are of pinkish color while others are colorless. Upon standing, colorless solutions turn to pinkish. What causes this phenomena is not known, hut the values of the zinc content are identical within the experimental error of about *I0 pg/g, independent of the color of solutions. The solution thus prepared normally contains 0.5-1 ppm of zinc. A series of solutions of known zinc concentration covering this range are prepared. This is best done by diluting a more concentrated standard solution. The absorhanees of these solutions are measured using the atomic absorption spectrometer set up for the determination of zinc. A graph of absorbance versus concentration is drawn. The calibration curve is usually straight up to 1.5 ppm. The absorbance of the solution of dissolved hair is measured immediately after the measurement of the standard solutions. The concentration of zinc this corresponds to is determined from the calibration curve. From this value the amount of zinc in the 100 ml of the solution and hence in the original hair sample can be calculated. The above procedure should be carried out at least in duplicate for each hair sample. A blank correction may be applied to the absorbance of the hair solution. However in practice this correction was negligible. Similarly, a more thorough washing of the hair before weighing has been found both in the present experiment and by other workers (7, 9, l l J to have only a very small effect on the results. Thus in a typical experiment by using Unicam Sp 90 spectrometer, 0.29M) g of the sample hair (from male, age 32, brown) when treated as described above had an absorbance 0.223. This corresponded to 0.540 ppm Zn in the solution and hence 183 pg/g of zinc in hair. Similarly 0.3166 g of the same hair gave an absorbance 0.245, which indicated 0.595 ppm of Zn in the solution and hence 188 pglg. The other hair sample (from female, age 12, black) was repeatedly analyzed. The values obtained were 309, 317, 297, 310, 301, 318, and 310 &gig, the average being 309 pg/g and the standard deviation 8pg/g. Acknowledgment
The authors wish to thank Dr. J. B. Farmer for valuable discussion.
' I'resrnt nddrrrs. Departmenr 01' Chmmicrry. Llniversitv of Alberta. Edmontun. Alhrrrn, Cnnado.
Literature Cited (11 Robinson. J. W.. "Principieo of Atomic Absorption." Program 3Wl CornmunieatianSkillCorp.. 1912. (21 White. A.. Handler. P.. and Smith, E. L.. "Principles of Biochemistry." McGrawHill BmkCa.. NewYork. 1968.p. 10L6. 131 Caldwell. D. F., Obnleas. D.. Clancy. J. J., and Pvasad. A. S., k c . Sor. Ezp. BiolMed.. 133, 1717 119701. 141 (4 Prasad. A. S.. "Zinc Metabolism," CC Thomsa, Springfield. Ill. 1966. p. 142: (bl P. 2,W. (51 Hurley, R.S., Ph.D.Theds. UniversityofUtah. 1972. 161 Chem Eng. News, May 1st. p. 17 (19721.
(71 Obrurnik. I.. Gilasan. J . , Maes. D., McMillan. D. K., D'Auria. I., and Pate. B. D.J. Rodimnnl. Chem.. IS. 1151L973);J. ForrnrirSciancea. 17.426119721. 181 Gordus. A , J Rodioonol. Chem., li.229I19731. 191 Verghwe, G. C.. Kisho*. R.. and Gunn. V. P., J . Rodiwnol. Chem., 15. 329
,.".",.
,la??,
110) Porkom. A. K..andJervis. R. E . J . Forensir Sciences. 11.6911966l. (111 HarRaun, W. W. Yuraeck. 1. P., and Benson. C. A,. Clin. Chim. Acto. 23. 83 (19691. 53.71 119691. (12) Schroeder.H.A.. endNason, A.P.. J. Inuerti~aiiueDermnfolopy, (131 Klsvsy, L. M..Arner. J . Clin. Nutrition, 23. 284 119101. (141 Petering. H. G., Yeaper. D. W., and Witherup, S. 0 ,Arch. Enuimn. H d f h . 23, 202 119711.
Volume 52, Number 8, August 1975 / 545
