Determination of the concentration of Fe, Se, and Zn elements in nails of Vietnamese women with breast cancer using k0-INAA method

Science & Technology Development Journal, 22(4):370-377 Open Access Full Text Article Research Article 1Department of Nuclear Physics, University of Science, VNU-HCM, 227 Nguyen Van Cu, Distr. 5, Ho Chi Minh City, Vietnam 2Department of Oncology, Dong Nai General Hospital, 2 Dong Khoi, Bien Hoa City, Vietnam 3Nuclear Technique Labolatory, Building B23, Linh Trung Campus, University of Science, VNU-HCM 4Institute of Nuclear Research, 1 Nguyen Tu Luc, Dalat City, Vietnam 5Department of Analytical Chemistry, University of Science, VNU-HCM, 227 Nguyen Van Cu, Distr. 5, Ho Chi Minh City, Vietnam Correspondence Huynh Truc Phuong, Department of Nuclear Physics, University of Science, VNU-HCM, 227 Nguyen Van Cu, Distr. 5, Ho Chi Minh City, Vietnam Email: htphuong@hcmus.edu.vn History  Received: 2019-07-17  Accepted: 2019-12-24  Published: 2019-12-31 DOI : 10.32508/stdj.v22i4.1698 Determination of the concentration of Fe, Se, and Zn elements in nails of Vietnamese womenwith breast cancer using k0-INAA method Huynh Truc Phuong1,*, Tran PhamNgoc Trinh2, Dinh Thanh Binh2, Nguyen Thi Truc Linh3, Truong Thi Hong Loan1,3, Tran Tuan Anh4, HoManh Dung4, Nguyen Van Dong5 Use your smartphone to scan this QR code and download this article ABSTRACT Introduction: Breast cancer is themost common type of cancer resulting in death. Trace elements of Fe, Se, and Zn can play a key role in the onset and prevention of breast cancer. Trace elements in the fingernails may be used as bioindicators for breast cancer diagnosis. The purpose of this work is to determine the concentrations of Fe, Se, and Zn in the fingernails of women with breast cancer and healthy women, which used to find the difference and correlation of these elements in the fingernail. Methods: This research was approved by the Ethics Committee of Dong Nai General Hospital. The fingernail collected from 29 women with breast cancer and 30 healthy women, who are the same age and living in Dong Nai province, Vietnam. The concentrations of Fe, Se, and Zn in the fingernails were determined using the k0-INAA method. The analytical data were evaluated using some statistical analysis for the correlation of trace elements in the fingernails of both groups. Results: As a result, the mean concentrations of Fe, Se, and Zn in fingernails of womenwith breast cancer were 102.87 mg/g, 0.75 mg/g and 65.49 mg/g, respectively, while those of healthy women were 69.74 mg/g, 0.78 mg/g and 107.75 mg/g. The assessment of these elements in fingernails for both two sample groups, including t-test and correlation coefficients, was also carried out in this study. As a result, the significant difference (P<0.05) was found for the Fe and Zn in fingernails of women with breast cancer, while those of Se was not found. The correlation between Se and Zn was found in both groups. The correlation between Fe and Zn was found in fingernails of healthy women, but it disappears in the women with breast cancer. Conclusions: From our findings, it can be concluded that Fe and Zn significantly associated with the risk of breast cancer, while Se is not associated. Key words: Essential elements in fingernails, breast cancer, the k0-INAA method INTRODUCTION Vietnam is one of the countries with the cancer rate at high levels. Prof. Nguyen Chan Hung suggested that, up to 2020, cancer is a reasonable cause of death for 115,000 people per year in Vietnam, approximately 315 people per day. In those, breast cancer is one of the common cancer types cause leading death for Vietnamese women. The reason for cancer-causing maybe start from the intake of the toxic elements into the human body, and/or it also can be due to reducing the concentration of essential elements in tissues. Trace elements entered the body by eating, activities, and living environment. The trace elements are ac- cumulated in human body tissues like hair and nails that have distinct advantages of the application as biomarkers. Therefore, they are recognized as biolog- ical tools for disease diagnosis. A large amount of evi- dence suggested that overload or deficiency of certain heavy metals linked with the risk of chronic diseases, including cancer and other ones1,2. Many studies claimed that there is a relationship be- tween trace elements and cancer risk3–5. Breast can- cer is the third most common cancer worldwide, and the most common cancer among women6. Its in- cidence increases with age, with greater frequency at menopause. It was reported that the lower level of Zn may be associated with an increased risk of breast cancer3, while the lower levels of Zn and Se, and the high level of Fe may be associated with an increased risk of prostate cancer7. The most accu- rate way to determine the level of trace elements in the body is the analysis of hair and nails8. Differ- ent analysis methods can be used to identify and measure trace elements, such as Total reflection X- Ray Fluorescence spectroscopy (TXRF)9, Atomic Ab- sorption Spectroscopy (AAS)10–12, Inductively Cou- pled Plasma Mass Spectroscopy (ICP-MS)13–15, In- ductively Coupled Plasma Atomic Emission Spec- troscopy (ICP-AES)16,17, and Instrumental Neutron Cite this article : Truc PhuongH, PhamNgoc Trinh T, Thanh Binh D, Thi Truc Linh N, Thi Hong Loan T, Tuan Anh T, Manh Dung H, Van Dong N. Determination of the concentration of Fe, Se, and Zn elements in nailsofVietnamesewomenwithbreast cancerusingk0-INAAmethod. Sci. Tech. Dev. J.; 22(4):370-377. 370 Copyright © VNU-HCM Press. This is an open- access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Science & Technology Development Journal, 22(4):370-377 ActivationAnalysis (INAA)18–20. Mostly thesemeth- ods are required for standard reference materials in comparison with analyzing samples. Meanwhile, the k0-INAA method is one of the methods which give high sensitivity and accuracy and not required for standard reference materials21,22. In this study, the k0-INAA standardization method was chosen for the determination of the concentra- tion of Fe, Se, and Zn in the nails of Vietnamese women with breast cancer and healthy. Besides, the statistical analysis was also considered in this study, such as the significant differences and correlation co- efficients between the elements in the fingernails of both sample groups. MATERIALS - METHODS This research was approved by the Ethics Committee of Dong Nai General Hospital. Sample collection The present study was conducted in Dong Nai province in the southeast of Vietnam, where locates many large industrial zones of the country. The study had involved a group of breast cancer females and a group of healthy persons as reference. The former group included twenty-nine female patients whose ages between 45 and 60, treated breast cancer at the Oncology Department of the General Hospital Dong Nai. The latter group was healthy women whose the same ages as the first group and living also in Dong Nai province. All the patients in the study were in first stage breast cancer and had not treated by chem- ical or radiation. The mean age was 47.28 for both studied groups. The fingernail samples were collected from August 2017 to February 2018 on the persons who had been aware of the investigation. The nail donors were asked to clean their hands with fresh water, then with distilled water. The fingernail from 10 fingers was taken by a stainless-steel snail clipper, kept in a pre-cleaned plastic bag, and stored in ambient laboratory conditions. The information on the nail donors, including name, ages, place of living, health conditions, etc. was recorded. Sample preparation For neutron activation analysis, the fingernail samples were treated as described elsewhere6,19,23,24. Briefly, the treatment procedure for fingernail based on 5 steps as follows: i) The fingernail samples were first kept soaked in distilled water for 10 minutes followed by another 5 minutes in rubbing alcohol with light shaking. This step was to reduce the risks of micro- biological activities from fungi and bacteria. ii) The fingernail samples were triplicated soaked in acetone with ultrasonic agitation (Model B2510-DTH, Bran- son, USA) for oneminute. For each replicate, the ace- tone was discarded, and new acetone was added to the nail sample. iii)The fingernail samples were treated in the same manner with step 2, using 2% Triton X100 (CAS 9002-93-1, Merck KGaA, Germany) instead of acetone. iv) The fingernail samples were triplicated cleaned by soaking in distilled water and ultrasonic agitation for one minute. v) The cleaned fingernail samples pre-dried by placing on the filter paper for 12 hours at ambient temperature. For neutron activation analysis, approximately 30-70 mg of each sample was placed in a cleaned polyethy- lene bag and sealed before irradiation. The certified reference materials, namely, NIST 1566b (Oyster Tis- sue) and NIST 1577a (Bovine Liver) were also used as quality control samples. For dry based calculation, the humidity of fingernail samples and certified ref- erence materials - NIST 1566b, and NIST 1577a were measured (Model MB45, Ohaus, USA). For this de- termination, approximately 80mg of NIST 1566b and 130 mg of NIST 1577a were dried in an oven (Model UFB 500, Memmert, Australia) at 80oC for 12 hours. The humidity was 4.1% and 11% for NIST 1566b and NIST 1577a, respectively. For the fingernail samples, the humidity was between 8.2% and 10.7%. Irradiation, measurements, and calculation The samples were divided into two groups. The first group was included with 29 fingernail samples from patients, NIST 1566b, cleaning blank (~ 123 mg) and Al-0.1%Au (wire form, ~ 3.6 mg) as a neutron flux monitor. The second group was included with 30 fin- gernail samples of healthy women and NIST 1577a. Samples from each sample group were placed to- gether, wrapped with aluminum foil and placed in an aluminum irradiation device called ”rabbit”. The neutron irradiation was performed in the Dalat Nu- clear Reactor, Vietnam for 10 hours under a thermal neutron flux of 3.2 x 1012 n.cm2.s1. The deviation of the epithermal neutron spectrum and the ratio of thermal/epithermal neutron fluxes were a = 0.071  0.001, and f = 39.5 0.4, respectively. The irradiated, the sample was measured using a gamma-ray spectrometer with HPGe detector (Can- berra, USA) which its resolution (FWHM) of 1.9 keV at 1332.5 keV peak of 60Co. Each sample was counted in a time of 10 hours, after 12 days of decay. At the full energy peak of radioisotopes, the net area was ob- tained using software GENIE 2000. For the fingernail samples, the full energy peaks of 59Fe (192.3 keV and 371 Science & Technology Development Journal, 22(4):370-377 1099.3 keV), 75Se (279.5 keV) and 65Zn (1115.5 keV) were measured. The element as Fe, Se, and Zn were also detected in the blank, but them at low levels (level of mg.kg1) and ignorable. The k0-INAA standard- ization method was used to calculate the concentra- tion of elements21,22. Data and statistical analysis The element concentrations were expressed as the arithmetic mean, standard deviation, standard error of the mean, minimum and maximum values, and median which calculated using Microsoft Office Ex- cel. The statistical significance of mean values be- tween cancer and the healthy group was determined by applying Student’s t-test. When a probability value (P-value) is smaller than 0.05, the difference was con- sidered to be significant. Besides, the compositional relationships among elements in fingernails are eval- uated by using the correlation matrix. RESULTS Analytical quality control After the correction of the blank, the concentration of elements in Certified Reference Materials was cal- culated. Here, the uncertainty of concentrations ob- tained was calculated using the propagation of error. Table 1 and Table 2 shows the results obtained in the analysis of NIST 1566b Oyster Tissue and NIST 1577a Bovine Liver, respectively. The relative devia- tion between measured and certified values for NIST 1566b Oyster Tissue and NIST 1577a Bovine Liver were lower than 7% and 9%, respectively. To evalu- ate the accuracy of the results obtained in the analy- sis of certified reference materials, the Z-score index was used in this work25,26. A result is considered ac- ceptable when the value of Z-score is between -3 and 3. The Z-score values obtained were below 2 indicate that the results obtained are within the range of cer- tified values at a level of significance of 5% 26. In this work, the Z-score values obtained are also shown in Table 1 andTable 1 forNIST 1566bOyster Tissue and NIST 1577a Bovine Liver, respectively. As a result, all of the elements obtained in the analysis of the cer- tified reference materials were satisfactory. It means that the analysismethod in this workwas trustworthy. Elemental concentrations in fingernails Table 3 and Table 4 show the results obtained in the analysis of fingernail samples of women with breast cancer and healthy women, respectively. As a re- sult, in the fingernails of both womenwith cancer and healthy, elements including Fe, Se, and Zn were rec- ognized in this work. The concentration of Se ele- ment was obtained at levels of the order of mg.g1, while those of elements, including Fe and Zn in order of much higher of mg.g1. The results showed that the concentration of Fe in the fingernails of women with breast cancer was higher than of those healthy women, while Zn level was at the lower level. The con- centration of Se was the same for both groups. The schemes for comparison on concentrations between the two groups shows in Figure 1. Statistical analysis Table 5 showed a significant difference in element concentrations in fingernails between breast cancer and healthy women. In this statistical analysis, the t- test, T critical at two tails and P values were carried out with a significant level of 0.05. As a result, there is a significant difference in the concentration of Fe and Zn between two groups (P < 0.05), while Se element was not different. Correlation analysis In order to examine the interrelationships between elements, the correlation coefficients between deter- mined elements in the fingernails of women with breast cancer and healthy women are represented in Table 6 and Table 7, respectively. In the fingernails of women with breast cancer, strong positive correla- tions are observed between Zn and Se (Table 6). For healthy women, strong positive correlations are found to the elements Zn and Fe, Zn and Se (Table 7). The significant correlation between Fe and Se was not found in both groups. In addition, Zn and Fe have correlated for healthy women, but they were disap- peared for women with breast cancer. DISCUSSION Themetals have diversified biological functions from essential elements to toxic elements, and it is the rea- son possibly causes cancer or other diseases. The es- sential metal elements such as Fe, Se, Zn, etc., are essential metal elements at common levels. The es- sential metal elements are very important in the pro- cess of metabolism, respiration, and in the process of growing up and death of the cells27–29. The change in the concentration of trace elements can be lead to illness or toxicity27,30,31. Table 6 and Table 7 shown that these results are com- pletely suitable for the correlation between Se and Zn. Because, the element zinc (Zn) is exciting for gene 372 Science & Technology Development Journal, 22(4):370-377 Table 1: Concentrations of elements in NIST 1566b Oyster Tissue Element Measured value (mg.g1) Relative deviation (%) jZ scorej Certified value (mg.g1) Ag 0.712 0.021 +6.5 2.01 0.666 0.009 As 7.29 0.47 -4.9 0.45 7.65 0.65 Co 0.347 0.040 -6.9 0.59 0.371 0.009 Fe 219.5 17.8 +6.2 0.72 205.8 6.8 K 6229 498 -4.7 0.58 6520 90 Na 3237 162 -1.9 0.35 3297 53 Rb 3.23 1.06 -0.9 0.03 3.26 0.14 Se 1.99 0.21 -3.5 0.27 2.06 0.15 Zn 1376 59 -3.5 0.64 1424 46 Table 2: Concentrations of elements in NIST 1577a Bovine Liver Element Measured value (mg.g1) Relative deviation (%) jZ scorej Certified value (mg.g1) Co 0.23 0.02 +8.7 0.37 0.21 0.05 Fe 204 22 +4.9 0.34 194 20 Na 2442 140 +0.5 0.06 2430 130 Rb 13.1 0.8 +4.6 0.74 12.5 0.1 Se 0.69 0.12 -2.9 0.14 0.71 0.07 Zn 121 4 -1.7 0.22 123 8 Table 3: Concentration of elements (mg.g1) in the fingernail samples of womenwith breast cancer Element No. sample Arithmetic mean Standard deviation Median Min. Max. Fe 20 102.87 37.53 91.220 49.447 188.746 Se 21 0.75 0.30 0.672 0.328 1.550 Zn 26 65.49 23.39 67.452 26.623 103.812 Table 4: Concentration of elements (mg.g1) in the fingernail samples of healthy females Element No. sample Arithmetic mean Standard deviation Median Min. Max. Fe 25 69.74 34.21 62.261 26.125 195.477 Se 29 0.78 0.42 0.742 0.161 1.947 Zn 28 107.75 38.95 103.561 50.406 204.000 373 Science & Technology Development Journal, 22(4):370-377 Figure 1: Scheme for comparison on the element concentrations between cancer and healthy women. Table 5: The statistical parameters of element concentrations in fingernails Element t-test T critical P-Value Sign. different Fe 3.060 2.021 0.007 Yes Se 0.295 2.012 0.841 No Zn 4.872 2.014 1.4x10-5 Yes Table 6: Correlation coefficients between elements in fingernails of women with breast cancer Fe Se Zn Fe 1 Se 0.267 1 Zn 0.067 0.613 1 Table 7: Correlation coefficients between elements in fingernails of healthy women Fe Se Zn Fe 1 Se 0.078 1 Zn 0.458 0.716 1 374 Science & Technology Development Journal, 22(4):370-377 transcription and cell proliferation, and increasing of Zn concentration in cells contributed to themultiple - cell processes, even if cells of tumors32. While the se- lenium (Se) has an effect to prevent for development of cancer cells according to a certain mechanism. Se- lenium (Se) helps for protection and against chromo- some injury which may be caused to cancer33. For this reason, Zn and Se are always to have a correlation. This result is also found in noncancerous and cancer- ous breast tissues34. In the fingernails of women with breast cancer, the Zn concentration was lower than that of healthywomen, while the Se concentrationwas still not different. Hence, the correlation between Zn and Se was slightly decreased for women with breast cancer. However, these changes were not broken for correlation with them. In this study, the result of the correlation between Zn and Fe was fairly interesting. In researching the correlation between Zn and Fe in the breast tis- sues of healthy and cancer women, Ammar Mubarak Ebrahim35 shown that, no correlation between Zn and Fe in the breast tissues of cancer women, but they are correlated in breast tissues of healthy women. This result was agreeing with our study for fingernails. Thus, the evaluation of the correlation between Zn and Fe in fingernails was not different from that in breast tissues. However, there are significant differ- ences in the average concentration of Zn and Fe be- tween fingernails and breast tissues. For the breast tissue of women with breast cancer, the concentra- tion of the element Zn was higher than that of healthy women. It is explained that the development of the tu- morwas brought about the increasing quantity of cells so that they are needed for the element Zn of trans- formation and metabolism. In the cancerous cells, increasing element Zn concentration was also repre- sented at other cancers, such as prostate cancer36, gas- tric cancer32,37. In addition, cancer cells need more blood than that compared with healthy cells, so that the concentration of element Fe has slightly increased, but there is not enough for significant statistical. In breast tissue, the concentration of element Zn was highly increased, while the concentration element Fe was not changed. This is shown that the correlation between Zn and Fe in breast tissues was reversed in comparison with fingernail tissues. In this study, the correlation between Zn and Fe in fingernails was dis- appeared for women with breast cancer (Table 6). In comparison, with healthy women, it was shown that the concentration of element Zn decreased, while the concentration of element Fe increased (Table 4). One can understand these cases as follows, because there no blood vessels in fingernail tissues so that the con- centration of element Fe in fingernails of women with breast cancer can not similar to cancerous breast tu- mors. In researching metal exposure in the nails of the population at Punjab, India, Blaurock has shown that the concentration of element Fe in nails of breast cancer patients was much higher than to healthy peo- ple and intake Fe into the body due to environmental exposure38. Recently, it was not found any research for the eval- uation of the concentration of element Zn in nails of women with breast cancer and healthy women. How- ever, a few of research has indicated that, for the pa- tients who contract a chronic As exposure, the con- centration of elemental Zn in hair was strongly de- creased in comparison with healthy humans39. Fur- ther, as the above discussions, the concentration of element Zn has increased in tissues of stomach can- cer patients32,37. However, in the research of Cam- pos et al.40,41 shown that there is a reverse correlation between the concentration of Zn in nails and stom- ach cancer. In our study, the concentration of Zn in the fingernails of womenwith breast cancer was lower than that in comparison with healthy women. It was proved that increasing the concentration Zn in can- cerous cells leads to deficient at the other organs in the body. Selenium is considered an essential trace element be- cause it is the primary component of selenoproteins, which have roles in counteracting oxidative stress and regulating the redox status of other molecules 42. In case–control studies such as those of Van’t Veer et al.43 and Ghadirian et al.44, both of which examined the level of Se in the toenail, which is consideredmore representative of long term Se exposure, found no as- sociation with breast cancer risk. This is a good agree- ment with the obtained results of Se in our study. CONCLUSION The present study k0-INAA method was used to de- termine the concentrations of Fe, Se, and Zn in finger- nail samples of women with breast cancer and healthy women. In the elements determined, the concentra- tion of Fe was found a much higher level in women with breast cancer than those of healthywomen, while the concentration of the element Zn was found at a lower level. The element Se was the same in concen- tration for both sample groups. The significant difference (P<0.05) between cancer and healthy women was found for the elements as Fe and Zn. The results of correlation analysis show that a strong positive correlation is found between Zn and Se in fingernails for both cancer and healthy women, 375 Science & Technology Development Journal, 22(4):370-377 while the correlation between Zn and Fe is found in fingernails for healthy women, but this correlation is disappeared for cancer women. These resultsmay be concluded that Fe andZn are sig- nificantly associated with breast cancer of the women, while Se is not associated with breast cancer risk. However, because of the small sample size, the results in this paper are insufficient to indicate that the con- centration of Fe, Se, and Zn in fingernails can be used as an indicator of breast cancer. Therefore, more ev- idence is needed to confirm that the elements in this study are associated with breast cancer. LIST OF ABBREVIATIONS INAA: Instrumental Neutron Activation Analysis NIST: National Institute of Standards andTechnology HPGe: High Pure Germanium FWHM: Full Width at Half Maximun AUTHOR CONTRIBUTIONS All authors contributed to the study conception and design. Material preparation, data collection, and analysis were performed by Huynh Truc Phuong, Tran Tuan Anh, Tran Pham Ngoc Trinh, and Nguyen Thi Truc Linh. Huynh Truc Phuong wrote the first draft of themanuscript and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. CONFLICT OF INTEREST The authors declare that they have no competing in- terests. ACKNOWLEDGMENTS This research is funded by the Vietnam National Foundation for Science andTechnologyDevelopment (NAFOSTED) under grant number 103.04-2017.311. Thank you to the ethical committee of Dong Nai General Hospital for approving of doing this study. The authors thank the staff of INAA Lab, Dalat Nu- clear Reactor, supported during neutron irradiation. Thank to doctor - nurses at the Department of Oncol- ogy, Dong Nai General Hospital, supported for sam- ple collection. REFERENCES 1. Gerhardsson L, Englyst V, Lundström NG, Sandberg S, Nord- bergG. Cadmium, copper and zinc in tissues of deceased cop- per smelter workers. J Trace Elem Med Biol. 2002;16(4):261– 6. PMID: 12530590. Available from: 10.1016/S0946-672X(02) 80055-4. 2. He K. Trace elements in nails as biomarkers in clinical research. Eur J Clin Invest. 2011;41(1):98–102. PMID: 20813017. Avail- able from: 10.1111/j.1365-2362.2010.02373.x. 3. Garland M, Morris JS, Colditz GA, Stampfer MJ, Spate VL, Bas- kett CK, et al. Toenail trace element levels and breast can- cer: a prospective study. Am J Epidemiol. 1996;144(7):653– 60. PMID: 8823061. Available from: 10.1093/oxfordjournals. aje.a008977. 4. Lappano R, Malaguarnera R, Belfiore A, Maggiolini M. Recent advances on the stimulatory effects of metals in breast can- cer. Mol Cell Endocrinol. 2017;457:49–56. PMID: 27765682. Available from: 10.1016/j.mce.2016.10.017. 5. Silva MP, Soave DF, Ribeiro-Silva A, Poletti ME. Trace elements as tumor biomarkers and prognostic factors in breast cancer: a study through energy dispersive x-ray fluorescence. BMC Res Notes. 2012;5(1):194. PMID: 22534013. Available from: 10.1186/1756-0500-5-194. 6. Bu-Olayan AH, Al-Yakoob SN, Alhazeem S. Lead in drinking water from water coolers and in fingernails from subjects in Kuwait City, Kuwait. Sci Total Environ. 1996;181(3):209–14. PMID: 8820436. Available from: 10.1016/0048-9697(95)05011- 6. 7. Karimi G, Shahar S, Homayouni N, Rajikan R, Bakar NFA, Oth- manMS. Association between trace element and heavymetal levels in hair and nail with prostate cancer. Asian Pac J Cancer Prev. 2012;13(9):4249–53. PMID: 23167323. Available from: 10.7314/APJCP.2012.13.9.4249. 8. Hubert A. Scoble and Robert Litman. Preparation of hair and nail samples for trace element analy- sis. Anal Lett. 1978;11(2):183–9. Available from: 10.1080/00032717808067866. 9. Magalhaes T, Becker M, Carvalho ML, von Bohlen A. Study of Br, Zn, Cu and Fe concentrations in healthy and cancer breast tissues by TXRF. Spectrochim Acta B At Spectrosc. 2008;63(12):1473–9. Available from: 10.1016/j.sab.2008.10. 014. 10. Sukumar A, Subramanian R. Relative element levels in the paired samples of scalp hair and fingernails of patients from New Delhi. Sci Total Environ. 2007;372(2-3):474–9. PMID: 17140638. Available from: 10.1016/j.scitotenv.2006.10.020. 11. Nowak B, Chmielnicka J. Relationship of lead and cadmium to essential elements in hair, teeth, and nails of environmentally exposed people. Ecotoxicol Environ Saf. 2000;46(3):265–74. PMID: 10903823. Available from: 10.1006/eesa.2000.1921. 12. Nowak B. Occurrence of heavy metals, sodium, calcium, and potassium in human hair, teeth, and nails. Biol Trace ElemRes. 1996;52(1):11–22. PMID: 8860662. Available from: 10.1007/ BF02784086. 13. Krachler M, Irgolic KJ. The potential of inductively coupled plasma mass spectrometry (ICP-MS) for the simultaneous de- termination of trace elements in whole blood, plasma and serum. J Trace Elem Med Biol. 1999;13(3):157–69. PMID: 10612079. Available from: 10.1016/S0946-672X(99)80006-6. 14. Rodushkin I, Axelsson MD. Application of double focusing sector field ICP-MS for multielemental characterization of hu- man hair and nails. Part I. Analytical methodology. Sci Total Environ. 2000;250(1-3):83–100. PMID: 10811254. Available from: 10.1016/S0048-9697(00)00369-7. 15. Yoshinaga J, Shibata Y, Morita M. Trace elements determined along single strands of hair by inductively coupled plasma mass spectrometry. Clin Chem. 1993;39(8):1650–5. PMID: 8353951. 16. Carl A. Burtis, Edward R. Ashwood, and David E. Bruns. Tietz textbook of clinical chemistry and molecular diagnostics. St. Louis, MO, USA: Elsevier Health Sciences; 2012. 17. Statistical analysis of selected heavy metals by ICP-OES in hair and nails of cancer and diabetic patients of Pakistan. EJEAFChe. 2012;11(3):163–71. 18. Arriola H, Longoria L, Quintero A, Guzman D. INAA of trace elements in colorectal cancer patients. Biol Trace Elem Res. 1999;71-72(1):563–8. PMID: 10676532. Available from: 10. 1007/BF02784244. 19. Aguiar AR, Saiki M. Determination of trace elements in hu- man nail clippings by neutron activation analysis. J Radioanal Nucl Chem. 2001;249(2):413–6. Available from: 10.1023/A: 1013235123875. 376 Science & Technology Development Journal, 22(4):370-377 20. Xiao L, Zhang YH, Li QG, Zhang QX, Wang K. INAA of ele- mental contents in fingernails of esophageal cancer patients. J Radioanal Nucl Chem. 1995;195(1):43–9. Available from: 10.1007/BF02036471. 21. F. De Corte. The k0-Standardization method: A move to the optimization of neutron activation analysis. PhD Thesis, GENT, 1987. 22. Corte FD, Simonits A. Recommended nuclear data for use in the k0 standardization of neutron activation analysis. At Data Nucl Data Tables. 2003;85(1):47–67. Available from: 10.1016/ S0092-640X(03)00036-6. 23. Gault AG, Rowland HA, Charnock JM, Wogelius RA, Gomez- Morilla I, Vong S, et al. Arsenic in hair and nails of individu- als exposed to arsenic-rich groundwaters in Kandal province, Cambodia. Sci Total Environ. 2008;393(1):168–76. PMID: 18234288. Available from: 10.1016/j.scitotenv.2007.12.028. 24. Brockman JD, Guthrie JM,Morris JS, Davis J,Madsen R, Robert- son JD. Analysis of the toenail as a biomonitor of supranu- tritional intake of Zn, Cu, and Mg. J Radioanal Nucl Chem. 2009;279(2):405–10. Available from: 10.1007/s10967-007- 7279-3. 25. Thomson M, et al. The international Harmonized Protocol for the Proficiency Testing of Analytical Laboratories. Pure Appl Chem. 2006;78(1):145–96. Available from: 10.1351/ pac200678010145. 26. . Bode. Instrumental andorganization aspects of a neutron ac- tivation analysis labolatory. PhDThesis - University of Technol- ogy, Delft, 1996. 27. Chan S, Gerson B, Subramaniam S. The role of copper, molyb- denum, selenium, and zinc in nutrition and health. Clin Lab Med. 1998;18(4):673–85. PMID: 9891606. Available from: 10.1016/S0272-2712(18)30143-4. 28. Christianson DW, Cox JD. Catalysis by metal-activated hy- droxide in zinc and manganese metalloenzymes. Annu Rev Biochem. 1999;68:33–57. Available from: 10.1146/annurev. biochem.68.1.33. 29. Kist AA, Zhuk LI, Danilova EA. On question of biological role of scandium. Abstracts of international conference on nuclear science and its application. Section. Environ Sci. 2012;44(50). 30. Cantley LC, Aisen P. The fate of cytoplasmic vanadium. Implications on (NA,K)-ATPase inhibition. J Biol Chem. 1979;254(6):1781–4. PMID: 217870. 31. Chan PC, Peller OG, Kesner L. Copper(II)-catalyzed lipid per- oxidation in liposomes and erythrocyte membranes. Lipids. 1982;17(5):331–7. PMID: 7098774. Available from: 10.1007/ BF02535190. 32. Magálová T, Bella V, Brtková A, Beno I, Kudlácková M, Volkovová K. Copper, zinc and superoxide dismutase in pre- cancerous, benign diseases and gastric, colorectal and breast cancer. Neoplasma. 1999;46(2):100–4. PMID: 10466433. 33. Zaichick V, Zaichick S. Fifty trace element contents in nor- mal and cancerous thyroid. Acta Scientific Cancer Biology. 2018;2(8):21–38. 34. Garg AN, Weginwar RG, Sagdeo V. Minor and trace elemental contents of cancerous breast tissue measured by instrumen- tal and radiochemical neutron activation analysis. Biol Trace Elem Res. 1990;26-27(1):485–96. PMID: 1704754. Available from: 10.1007/BF02992704. 35. Ebrahim AM, EltayebMA, Shaat MK, Mohmed NM, Eltayeb EA, Ahmed AY. Study of selected trace elements in cancerous and non-cancerous human breast tissues from Sudanese subjects using instrumental neutron activation analysis. Sci Total En- viron. 2007;383(1-3):52–8. PMID: 17570463. Available from: 10.1016/j.scitotenv.2007.04.047. 36. Kwiatek WM, Banas A, Banas K, Gajda M, Galka M, Falkenberg G, et al. Iron and other elements studies in cancerous and non cancerous prostate tissues. J Alloys Compd. 2005;401(1):178– 83. Available from: 10.1016/j.jallcom.2005.03.090. 37. Magálová T, Beno I, Brtková A, Mekinová D, Volkovová K, Staruchová M, et al. [Levels of Cu, Zn, Se and their relation to levels of ceruloplasmin and the activity of antioxidative en- zymes]. Bratisl Lek Listy. 1997;98(1):8–11. PMID: 9264806. 38. Blaurock-Busch E. Yvette M. Busch, Albrecht Friedle, Hol- ger Buerner, Chander Parkash and Anudeep Kaur. Compar- ing the metal concentration in the nails of health and can- cer patients living in the Malwa region of Punjab, India with a random Euopean group - A follow up study. Br J Med Med Res. 2015;5(4):480–98. Available from: 10.9734/BJMMR/2015/ 13124. 39. Trace elements in the hair of normal and chronic arsenism people. Global Advanced Research Journal of Environmental Science and Ecotoxicol. 2013;2(7):163–73. 40. Campos F, Carrasquilla G, Koriyama C, Serra M, Carrascal E, Itoh T, et al. Risk factors of gastric cancer specific for tumor location and histology in Cali, Colombia. World J Gastroen- terol. 2006;12(36):5772–9. PMID: 17007041. Available from: 10.3748/wjg.v12.i36.5772. 41. Campos FI, Koriyama C, Akiba S, Carrasquilla G, Serra M, Car- rascal E, et al. Toenail zinc level and gastric cancer risk in Cali, Colombia. J Cancer Res ClinOncol. 2008;134(2):169–78. PMID: 17619905. Available from: 10.1007/s00432-007-0266-1. 42. Goldhaber SB. Trace element risk assessment: essentiality vs. toxicity. Regul Toxicol Pharmacol. 2003;38(2):232–42. PMID: 14550763. Available from: 10.1016/S0273-2300(02)00020-X. 43. van ’t Veer P, van derWielen RP, Kok FJ, Hermus RJ, Sturmans F. Selenium in diet, blood, and toenails in relation to breast can- cer: a case-control study. Am J Epidemiol. 1990;131(6):987– 94. PMID: 2343870. Available from: 10.1093/oxfordjournals. aje.a115619. 44. Ghadirian P, Maisonneuve P, Perret C, Kennedy G, Boyle P, Krewski D, et al. A case-control study of toenail selenium and cancer of the breast, colon, and prostate. Cancer Detect Prev. 2000;24(4):305–13. PMID: 11059562. 377