The present work is a descriptive, cross-sectional study carried out with individuals selected from the control series of a hospital-based case-control study conducted in Petropolis to investigate the association between exposure to pesticides and the development of different types of cancer. This study excluded individuals with cognitive impairment that made it impossible to communicate or understand the research questionnaire questions, individuals hospitalized for treatment of any cancer or symptoms potentially associated with neoplasms, and individuals with a previous history of cancer. Thus, the 151 participants included in the present study are part of a sample of individuals over 18 years old admitted to the Alcides Carneiro Municipal Hospital, from December 2014 to March 2020, with no previous history of cancer. Trained personnel used a structured questionnaire to interview the participants at the health unit. At the end of the interview, a peripheral blood sample was extracted (4 ml in a tube with EDTA) and processed by ultracentrifugation at 8,000 rpm for 20 minutes to separate the plasma from the cell layer. Analysis of plasma samples executed through gas chromatography coupled to triple quadrupole mass spectrometry (GC-MS/MS). The persistent organic pollutants (POPs) of the organochlorine (OC) class analyzed were: alpha-HCH, beta-HCH, gamma-HCH, delta-HCH, 2.4'-DDD, 2.4'-DDE, 2.4 '-DDT, 4,4'-DDD, 4,4'-DDE, 4,4'-DDT, Aldrin, Dieldrin, Endrin, Alpha-Clordan, Gamma-Clordan, Alpha-Endosulfan, Beta-Endosulfan, Endosulfan Sulfate, Dicofol, Heptachlor, Heptachlor epoxide, Hexachlorobenzene, Methoxychlor, Mirex, Pentachloroanisole, trans-Nonachlor. The polychlorinated biphenyls analyzed were: PCB 101 (2,2',4,5,5'-Pentachlorobiphenyl); PCB 105 (2,3,3',4,4',-Pentachlorobiphenyl); PCB 118 (2,3',4,4',5-Pentachlorobiphenyl); PCB 126 (3,3',4,4',5-Pentachlorobiphenyl); PCB 128 (2,2',3,3',4,4'-Hexachlorobiphenyl); PCB 138 (2,2',3,4,4',5'-Hexachlorobiphenyl); PCB 153 (2,2',4,4',5,5'-Hexachlorobiphenyl); PCB 156 (2,3,3',4,4',5-Hexachlorobiphenyl); PCB 169 (3,3',4,4',5,5'-Hexachlorobiphenyl); PCB 170 (2,2',3,3',4,4',5-Heptachlorobiphenyl); PCB 180 (2,2',3,4,4',5,5'-Heptachlorobiphenyl); PCB 28 (2,4,4'-Trichlorobiphenyl); PCB 31 (2,4',5-Trichlorobiphenyl); PCB 52 (2,2',5,5'-Tetrachlorobiphenyl); PCB 77 (3,3',4,4'-Tetrachlorobiphenyl).
The analytical method was adapted from SARCINELLI et al. (2003). Briefly, the samples were thawed at room temperature and subsequently denatured, then diluted with equal parts of methanol and water, then mixed and extracted in previously conditioned 500mg/6mL C18 solid-phase extraction cartridges (JT Baker, USA), dried, and eluted with 7 mL of hexane afterward. A pre-conditioned 1g/6mL florisil cleanup cartridge received the eluate, which was eluted with hexane followed by dichloromethane/hexane (1:1 v/v), a solution of petroleum ether: hexane (85:15). The extracts evaporated to dryness under a nitrogen atmosphere. The volume was resuspended to 100 µL with hexane and analyzed by GC-MS/MS, using 5 µL of dibromophenol 1, 1'-biphenyl-4, 4'dibromine at 1 µg mL− 1 as the internal standard.
A GC-MS/MS analysis using Thermo Scientific equipment, model TSQ 8000 EVO, containing a Trace GC 1310 chromatograph with AS 1310 autosampler with programmable temperature vaporization (PTV) injectors, operating in splitless mode with ThermoFisher Xcalibur™ and TraceFinder™ software. An Agilent® DB-5MS column that contained phenylmethyl siloxane (30 m x 250 µm x 0.25 µm), with injection without flow splitting, received 2 µL of the sample. Compounds were identified by selected individual reaction monitoring (SRM) adjusted for retention time. The National Institute of Standards and Technology (NIST) libraries, included in the Tracefinder™ software, defined the spectrometric conditions, and Accu Standard's high purity standards confirmed those conditions. Therefore, transitions were defined according to their high specificity combined with high abundance.
Plasma levels of each POP were presented as mean, median, and standard deviation (SD) in ng mL− 1 and ng g− 1 of lipid to compare the results obtained in the literature. Since the lipid content was not measured individually in each sample, the average reference values for adults of fasting triglycerides (below 150 mg dL− 1) and fasting total cholesterol (below 190 mg dL− 1) served as the basis. The total lipid content was estimated using the formula established by Phillips et al. (1989), whose ratio is total lipids = 2.27 * total cholesterol + triglycerides + 0.623. As a result, the calculated total lipid content was 2.27 * 180 + 150 + 0.623 mg dL− 1, or 560 mg dL− 1.
The IBM-SPSS 20.0 program provided data analysis. Characterization of the study population was according to age group, gender, skin color, schooling, BMI, alcohol consumption, smoking, and variables related to exposure to pesticides. The use of the Mann-Whitney or Kruskal-Wallis U tests established the existence of a correlation between the plasma levels of POPs and these variables.
The procedures of this project followed the recommendations of Resolution No. 466/12 of the National Health Council. This project received the approval of the Research Ethics Committees of the Public Health National School (ENSP – Escola Nacional de Saúde Pública) (CAAE: 31126514.0.0000.5240) and the Alcides Carneiro Municipal Hospital.