**3. Results**

#### **3.1 Validation of analytical methods**

The analytical methods used were validated previous to the execution of this study. The parameters determined were linearity, repeatability, recovery from the rainwater matrix and method detection limits (MDL). For the determination of the organochlorine pesticides, polychlorinated biphenyls, brominated flame retardants, phthalates, artificial musks and organotin compounds internal standards were added to each sub-sample prior to analysis. For the perfluorinated compounds two relevant samples are spiked with the compounds of interest and analysed. The recovery of the internal standard and spikes were in the range of 70 to 140% with the exception of artificial musks where recoveries in the range of 56 to 87% were found, and the organotin compounds where recoveries were in the range of 67 to 99%. With the exception of PCBs and organotin compounds, the results are not corrected for the recovery of the internal standards or spikes since the spikes used were not compound specific and their recovery is only used to evaluate the performance of the method. With each series of ten samples a blank sample was included. For the blank analysis the complete analytical procedure was followed, including all chemicals and solvents, but no sample was added. Blank results were only found for the phthalates DEHP, DIBP and DBP, The results were corrected for these blank values and the detection limits were raised to 10 ng/g for DIBP and DBP, and to 20 ng/g for DEHP.

#### **3.2 Organochlorine pesticides**

72 Pesticides in the Modern World - Risks and Benefits

partitioning the hexane extract is extracted with dimethylformamide (DMF) to isolate the phthalates and artificial musks. After removal of the hexane layer, water is added to DMF fraction and the analytes are re-extracted into fresh hexane. This extract was concentrated and purified over a glass chromatographic column packed with florisil and capped with anhydrous sodium sulphate to isolate the fraction containing the phthalates and artificial musks. The eluent was concentrated to a small volume, the syringe standard was added and this final extract was analysed with the identical GC/MS system as described above. The mass spectrometer was operated in the selected ion monitoring mode and typically two or three characteristic ion masses were monitored for each analyte. The samples were analyzed for the following phthalates; di-methyl- (DMP), di-ethyl- (DEP), di-isobutyl- (DIBP), dibutyl- (DBP), benzylbutyl- (BBP), di-(2-ethylhexyl)- (DEHP), di-isononyl- (DINP) and diisodecyl phthalate (DIDP): The following artificial musks; musk ketone (MK), musk xylene

AP's and the brominated flame retardant tetrabromobisphenol-A (TBBPA) were isolated using a steam distillation procedure described by Guenther et al., 2002. In the round bottom flask of the steam distillation apparatus typically 10 g of the laboratory sample was spiked with an internal standard and mixed with 250 ml Milli-Q water to which was added 1 ml of concentrated hydrochloric acid and 20 g of sodium chloride. During the overnight distillation process the organic phenols are isolated in the organic solvent, in this case hexane, in the backflow cooling system of the apparatus. The hexane extract is isolated, dried with anhydrous sodium sulphate and concentrated. Following drying the extract was reduced in volume nearly to dryness under a stream of nitrogen and re-dissolved in 1 ml of methanol. During concentration of the hexane extract care has to be taken to avoid losses of the AP's and rinsing of the glass surfaces with methanol is necessary. The methanol extract is analysed with liquid chromatography coupled with mass spectrometry (LC/MS) in the

For the determination of perfluorinated compounds, specifically perfluoro-octanoate (PFOA), perfluoro-octane sulphonate (PFOS) and perfluoro-octane sulphonamide (PFOSA) sub-samples of 5 gram were collected in 50 ml poly-propylene tubes and extracted using acetonitrile. The samples were centrifuged and the clear liquid is decanted and purified over a glass chromatographic column packed with florisil, silica, LC-NH2 and activated carbon. The residue in the poly-propylene tube is extracted two times more and each extract is decanted over the same chromatographic column, thus combining the purified extracts. 0.5

The analytical methods used were validated previous to the execution of this study. The parameters determined were linearity, repeatability, recovery from the rainwater matrix and method detection limits (MDL). For the determination of the organochlorine pesticides, polychlorinated biphenyls, brominated flame retardants, phthalates, artificial musks and organotin compounds internal standards were added to each sub-sample prior to analysis. For the perfluorinated compounds two relevant samples are spiked with the compounds of interest and analysed. The recovery of the internal standard and spikes were in the range of 70 to 140% with the exception of artificial musks where recoveries in the range of 56 to 87% were found, and the organotin compounds where recoveries were in the range of 67 to 99%.

ml octanol is added as a keeper and the extracts are concentrated to a small volume.

(MX), tonalide (AHTN) and galaxolide (HHCB).

selected ion monitoring mode.

**3.1 Validation of analytical methods** 

**3. Results** 

Organochlorine pesticides (OCPs) include compounds like DDT, lindane, hexachlorobenzene and chlordane, among others. DDT is a well-known agricultural insecticide that has been used extensively on a global basis for over 40 years. Although their manufacture and application are now largely prohibited or restricted in industrialized western countries because of their toxicity and persistence, they can still be found in environmental and biological matrices due to their persistence. Pesticide exposure has been associated with arthritis, diabetes, neurobehavioral changes and DNA damage Cox et al., 2007; Lee et al. 2007; Jurewicz et al., 2008; Rusiecki et al., 2008). The structures of p,p'-DDT and it's breakdown product p,p'-DDE, are shown below.

 As a part of its monitoring program, the Food and Drug Administration (FDA) determines the levels of pesticide residues in a wide variety of foods typically consumed by Americans. Over the past ten years, these surveys have detected DDE and other OCPs in a variety of foods including meat, fish and shell fish products, eggs, root vegetables, legumes (beans, peas, and peanuts), some fruits, and leafy greens. In 1999 DDE or DDT were detected in 22% of the 1,040 food items analysed in the FDA Total Diet Study (FDA, 1999). The results for the 2003 Total Diet Study indicate DDT, but mainly DDE in 18% of the various food items in concentrations ranging from 0.1 to 11 ng/g (FDA, 2003). Those for chlordane and lindane range from 0.1 to 3.8 and from 0.1 to 8.4 ng/g product. In general, the concentrations as well as the frequency of detection of OCPs were lower in the 2003 study. The results of the OCPs in this study are presented in table 2 at the end of this section, and graphically in figure 1. OCPs are found in 17 of the 25 samples. The predominant OCPs that are detected are p,p'-DDE and HCB both found in 15 of the 25 samples. In addition o,p'-DDE and cis-chlordane were found in one and two samples respectively. The maximum concentration found for p,p'-DDE was 5.6 ng/g in a sample of pickled herring. The median concentrations for p,p'-DDE and HCB were 0.43 and 0.14 ng/g. Compared to the FDA's Total Diet Study, p,p'-DDE and HCB are found more frequently but in lower concentrations. This compares with a recent study of Schecter et al. who reported p,p'-DDE in 23 out of 31 samples in concentrations ranging from 0.06 to 9.0 ng/g with a median value of 0.51 ng/g (Schecter et al., 2010).

Industrial Contaminants and Pesticides in Food Products 75

section. A graphic summary is given in figure 2. PCBs are found in every sample with PCB-18, -28 and -52, with the highest frequency. The highest PCB concentrations were found in the pickled herring and the salmon with concentrations for the sum of i-PCBs of 13 and 3.3 ng/g. The highest concentration of an individual PCB was for PCB-153, 5.5 ng/g in the salmon. The results found are comparable with those reported by EFSA for different food

32

types sampled across Europe.

total-PCBs (ng/g)

Butter

Cheese

Bacon

**3.4 Brominated flame retardants** 

Sausages

Eggs

Milk

Olive oil

predominantly found in fish products, especially in eel.

Chicken

Fish fingers

Fig. 2. Graphic presentation of the total-PCB concentration in 24 food items

Salmon

Honey

Tuna

Brominated flame retardants (BFRs) are widely used in electronic household equipment, plastics, textile and polyurethane foam in furniture and cars for safety reasons. Of the brominated products, about one-third contain tetrabromobisphenol-A (TBBPA) and derivatives, another third contains various bromines, including hexabromocyclododecane (HBCD) and the last third contains polybrominated diphenylethers (PBDEs). All three types of BFRs are determined in this study. The PBDEs are commercial mixtures with different degrees of bromination and used as additives to fireproof polymers. HBCD is a cycloaliphatic brominated chemical introduced as a replacement for the PBDEs and with the same applications. TBBPA is mainly used as a reactive (chemically bound) flame retardant in epoxy polymers such as printed circuit boards in electronic equipment. The structure of BDE-209 (better known as deca-BDE), HBCD and TBBPA is shown below. In a study by the Dutch National Institute for Public Health and the Environment the levels of various PBDEs, HBCD and TBBPA in 84 food products were determined (de Winter-Sorkina et al., 2003, 2006). With the exception of fish products PBDEs were absent or present in low concentrations (<0.1 ng/g) in food products. For the fish products the concentrations of the congeners BDE-28, -47, -99, -100, -153 and -154 ranged from 0.1 to 14 ng/g. BDE-209 was found in none of the 84 products while TBBPA was found in 7 products in concentrations ranging from 0.1 to 3.4 ng/g. Surprisingly, HBCD was found in 28 of the 84 samples in concentrations ranging from 0.1 to 8.9 ng/g product. TBBPA and HBCD were

Brown bread

Frankfurter sausages

Reindeer meat

Pickled herring

Minced beef

Pork chops

Cottage cheese

Salami

Cheese

Ham

Cheese

Scottish cheese

Fig. 1. Graphic presentation of the total-OCP concentration in 25 food items

#### **3.3 Polychlorinated biphenyls**

Polychlorinated biphenyls (PCBs) are marketed as cooling or insulating fluids for transformers, as softeners in the varnish and adhesive industries, and as hydraulic fluids. These compounds are not combustible, are heat resistant and make good solvents. On the other hand there is a severe toxic effect of PCBs, which damage the organs responsible for metabolism and also the nervous system (Guo et al., 1995; Ribas-Fito et al., 2001). Because of their persistence, PCBs are widely spread in the environment and, due to their good liposolubility, are easily deposited and concentrated in human, animal and plant tissue. The structures of two of the seven indicator PCBs, PCB-28 and PCB-180 are shown below.

 In the Total Diet Study of 2003 the levels of the sum of i-PCBs (e.g. PCB-28, PCB-52, PCB-101, PCB-118, PCB-138, PCB-153 and PCB-180) ranged from 6 to 70 ng/g product. The Estonian Environment Research Centre determined levels of the sum of i-PCBs in Estonian butter and found a relative narrow concentration range of 5.2 to 8.8 ng/g lipid, translating to a range of 4.2 to 7.0 ng/g on a wet weight basis (EU, 2004). A study of the European Food Safety Association (EFSA) of different food types sampled across Europe between 1997 and 2003 reports median concentrations for the sum of i-PCBs ranging from 1.4 to 12 ng/g. Only fish oil showed a higher median concentration of 52 ng/g for the sum of i-PCBs (Gallani et al., 2004). The results for the PCBs in this study are presented in table 2 at the end of this

total-OCBs (ng/g)

Butter

Cheese

Bacon

Sausages

**3.3 Polychlorinated biphenyls** 

Eggs

Milk

Olive oil

Chicken

Fish fingers

Salmon

Fig. 1. Graphic presentation of the total-OCP concentration in 25 food items

Honey

Orange juice

Polychlorinated biphenyls (PCBs) are marketed as cooling or insulating fluids for transformers, as softeners in the varnish and adhesive industries, and as hydraulic fluids. These compounds are not combustible, are heat resistant and make good solvents. On the other hand there is a severe toxic effect of PCBs, which damage the organs responsible for metabolism and also the nervous system (Guo et al., 1995; Ribas-Fito et al., 2001). Because of their persistence, PCBs are widely spread in the environment and, due to their good liposolubility, are easily deposited and concentrated in human, animal and plant tissue. The structures of two of the seven indicator PCBs, PCB-28 and PCB-180 are shown below.

 In the Total Diet Study of 2003 the levels of the sum of i-PCBs (e.g. PCB-28, PCB-52, PCB-101, PCB-118, PCB-138, PCB-153 and PCB-180) ranged from 6 to 70 ng/g product. The Estonian Environment Research Centre determined levels of the sum of i-PCBs in Estonian butter and found a relative narrow concentration range of 5.2 to 8.8 ng/g lipid, translating to a range of 4.2 to 7.0 ng/g on a wet weight basis (EU, 2004). A study of the European Food Safety Association (EFSA) of different food types sampled across Europe between 1997 and 2003 reports median concentrations for the sum of i-PCBs ranging from 1.4 to 12 ng/g. Only fish oil showed a higher median concentration of 52 ng/g for the sum of i-PCBs (Gallani et al., 2004). The results for the PCBs in this study are presented in table 2 at the end of this

Tuna

Brown bread

Frankfurter sausages

Reindeer meat

Pickled herring

Minced beef

Pork chops

Cottage cheese

Salami

Cheese

Ham

Cheese

Scottish cheese

section. A graphic summary is given in figure 2. PCBs are found in every sample with PCB-18, -28 and -52, with the highest frequency. The highest PCB concentrations were found in the pickled herring and the salmon with concentrations for the sum of i-PCBs of 13 and 3.3 ng/g. The highest concentration of an individual PCB was for PCB-153, 5.5 ng/g in the salmon. The results found are comparable with those reported by EFSA for different food types sampled across Europe.

Fig. 2. Graphic presentation of the total-PCB concentration in 24 food items
