Determination of Some Trace Elements in Breast Cancer Serum by Atomic Absorption Spectroscopy

In the breast cancer significant differences occurs in the normal distribution of the trace elements, playing an important role in carcinogenic process. The aim of this study was to investigate the serum levels of some trace elements (Chromium, Cadmium, Manganese, Cobalt, Nickel, Selenium, Zinc, Iron, Copper and Magnesium) by using flame atomic absorption spectroscopy (FAAS). The present study included 150 females, the participates were divided into two main groups, control group which consisted of 75 apparently healthy females, 75patients with breast cancer group .The serum levels of Chromium (tcal2.9631 ,ttab1.960), Cadmium (tcal2.0798 ,ttab1.960), Manganese (tcal18.5676, ttab1.960), Selenium (tcal2.2759,ttab1.960), Iron (tcal2.9296,ttab1.960), Copper (tcal4.0869, ttab1.960),Magnesium (tcal2.6648, ttab1.960), Cobalt (tcal3.8615 , ttab1.960) and Zinc (tcal7.0160, ttab1.960) were statistically significant higher in the breast cancer patients group than the control group. Similarly, Nickel (tcal1.5988,ttab1.960) levels showed significant lower level in breast cancer patients group. On the other hand, the higher levels of trace elements could lead to formation of free radicals or other reactive oxygen species. The evaluation of these trace elements in serum maybe used as helpful tool in diagnosis of the breast cancer.


Introduction
Almost 1.7 million new cases of breast cancer diagnosed in 2012, 12% new of cancer cases and 25% of all cancers in women, causing death to women worldwide (Ferlay et al., 2012).The etiology of breast cancer is multi-factors which including, hormone, genetic and environmental can be affect the pathogenesis of the disease, but the exact factor are unknown (McPherson, Steel & Dixon 2000) and (Russo, Fu, Yang & Russo, 2000).Oxidative stress in the cells will predominate when the formation of reactive oxygen species (ROS) becomes too much, overpowering the normal antioxidant defense system, reduce the antioxidant system in addition to formation more free radicals (McKeown, 1999) and (Halliwell & Cross, 1994).Many trace elements can activate or inhibit enzymes through rival with the other elements and metalloprotiens for binding sites or disturbing the cell membrane permeability, playing direct or indirect role on the carcinogenic routes (Huang, Sheu & Lin, 1999).The increasing formation of free radicals may harm the tissue through the reaction with lipid cell membrane, proteins and nucleic acids (Halliwell, 2007).The cellular defense mechanisms (antioxidant mechanism) can control the free radicals species levels through, enzymatic (glutathione peroxidase, superoxide dismutase) and non-enzymatic compounds (vitamin E, vitamin C, glutathione and uric acid) (Badid et al., 2010).The oxidative damage to the biological molecules will create lipid peroxydation, mutagenesis, carcinogenesis (Kasapovic, Pejic,Todorovic, Stojiljkovic & Pajovic, 2008).The air, drinking water and food are the natural resources of trace elements which the human can be exposure possibly modified (Bertini & Cavallaro, 2008).The normal levels of the trace elements are essential to the human internal stability balance.The trace elements determination is important for their main role in the biochemical and physiological processes (Navoro & Rohan, 2007).
The latest investigations discovered the relationship between the trace elements and cancer, some of them inducing the toxicity effect during the production of free radicals and acting as cofactors in oxidative destruction of the macromolecules and DNA (Leonard, Bower & Shi, 2004).
The exact role of the serum trace elements levels in carcinogenesis, breast cancer, oxidative stress and different tumor markers are still shortage and pellicular (Navoro et al., 2007).The exact mechanism of tumor creation, high oxidative DNA damage after metal exposure is unclear.
The variance of the trace elements levels in neoplastic breast tissues has been found through many investigations, on the other hand the metabolic and microenviromental factors which are responsible of the tumors creation are complex and not completely known (Silva & Poletti, 2009).The aim of the present study was to investigate the association of the serum levels of some trace elements ,Chromium (Cr),Cadmium (Cd), Manganese (Mn),Cobalt (Co),Nickel (Ni),Selenium (Se), Zinc (Zn),Iron (Fe),Copper (Cu) and Magnesium (Mg) with breast cancer in order to add more knowledge to their rule in the patients ( Silva & Poletti, 2009).

Apparatus
Aurora atomic absorption spectrophotometer model (AI-1200) was used with an air-acetylene burner (slot length 11 cm).Instrument setting were: lamp current, 5-10 mA, slit width, 0.2 nm, air flow, 1.8 L/min, ignition flow 2.4 L /min as shown in Table 1.These conditions were maintained constant throughout the measurements.The operating conditions adjusted according to the standard guideline of the manufacturer.A standard solution of each element was prepared immediately by serial dilution of 1000 mg/L stock solution (Scharlau Chemie) with deionized water prior to use.

Preparation of Samples
Seventy five female patients, with histopathological diagnosis of breast cancer, aged 26-62 years were selected from Al-sader public hospital in Misan-Iraq.Seventy five matched healthy women who attended routine health examination in the same hospital were selected from the same area with no history of any tumor or breast disease.10 ml of venous blood without using anticoagulant samples were collected from patients and healthy volunteers in plain screw cap specimen bottles and then left for 30 min for retraction after, which centrifugation was carried out at 3000 rpm for 5 min and the supernatant serum was aspirated by using pasteur pipette into another plain bottle.The collected supernatant was then pooled and stored at −20°C until the time of analysis.Serum samples was diluted with deionized water and homogenized before analysis.

Statistical Analysis
The data were subjected to statistical analysis using the student t-test for comparison of elements levels between patient and control groups.All the data were expressed as mean and standard deviation of the mean.The quantitative determination of the elements in samples was done by using calibration curves, linearity which obtained from diluted stock solution of each element 1000 mg/L as shown in Table 1.The concentration of the analytes was obtained directly from the FAAS, through the linear regression's equations after the correction of the absorbance signal from the blank.

Results and Discussion
Ten trace elements as follows; chromium(Cr), cadmium (Cd), manganese (Mn)), cobalt(Co), nickel(Ni), selenium(Se), zinc (Zn),iron(Fe),copper(Cu),and magnesium (Mg)were identified and their concentrations were determined for each serum sample.The obtained concentration data was averaged separately for the control and breast cancer groups.The compares average concentrations along with the corresponding standard deviations for each element between75 patients and 75 control groups are shown in Table 2.
The order of the mean trace elements concentrations in serum samples of breast cancer group is Mg Fe Cu  ZnSe Ni Co Mn Cd  Cr.Overall mean concentrations of Mg (20.51μg/mL),Fe (1671.4μg/L),Cu(1338.5μg/L),Zn (920.75μg/L),Se(89.4μg/L)Ni (24.5μg/L),Co (20.3μg/L),Mn (15.3μg/L),Cd (14.5 μg/L) and Cr (13.2 μg/L) in breast cancer group, in contrast the order of the mean trace elements concentrations in serum samples of control group is Mg

Figure 1 .
Figure 1.Mean concentrations of trace elements (Cr, Cd, Mn, Co, Ni, Se, Zn, Cu and Fe ) in Breast Cancer and control groups (n=75)

Table 1 .
Operating parameters of FAAS, wavelength, slit width, (HCL) current, range of calibration standardsand flame This study warrant further investigations on larger sample size, more other trace elements and multiple serum samples. http://ijc.ccsene