Survey of Argentinean human plasma for ochratoxin A

Survey of Argentinean human plasma for ochratoxin A Authors: A. M. Pacin ab; E. V. Ciancio Bovier a; E. Motta c; S. L. Resnik bd; D. Villa e; M. Olsen f Affiliations: a Fundación de Investigaciones Científicas Teresa Benedicta de la Cruz, Centro de Investigación en Micotoxinas, Universidad Nacional de Luján, Buenos Aires, Argentina b Comisión de Investigaciones Científicas de la Provincia de Buenos Aires (CIC), Argentina c Universidad Nacional de Mar del Plata, Buenos Aires, Argentina d Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Argentina e Dirección Sistemas, Universidad Nacional de Luján, Buenos Aires, Argentina f National Food Administration, Microbiology Division, Uppsala, Sweden DOI: 10.1080/02652030701613709 Publication Frequency: 12 issues per year Food Additives & Contaminants Published in: First Published on: 21 December 2007 Abstract The presence of ochratoxin A (OTA) in human blood has been reported for many countries, especially in Europe. However, so far no report exists concerning such a presence in Argentina. The aim of this study was to assess OTA concentration in human plasma in two different areas of Buenos Aires province. OTA was determined by high-performance liquid chromatography (HPLC) in 199 plasma samples from blood donors in Mar del Plata and 236 from General Rodríguez. Solid-phase extraction with Bakerbond® C-18 cartridge and a final purification with Ochraprep® immunoaffinity columns was employed. The limit of quantification of ochratoxin A was 0.019ngml-1 and the confirmation of OTA was by formation of ochratoxin A methyl ester. The results showed that 63.8% of human plasma samples from Mar del Plata and 62.3% from General Rodríguez were positive for OTA, with Winsorized means of 0.15 and 0.43ngml-1, respectively. It is important to continue the research to detect the foods responsible of the presence of OTA in plasma. Keywords: Ochratoxin; mycotoxin; human plasma; exposure Introduction Ochratoxin A (OTA) is a mycotoxin produced by Aspergillus and Penicillium species. It can contaminate a variety of food items such as cereals, coffee, wine, beer, etc., resulting in chronic human exposure. OTA is very toxic to several animal species, the kidney being the main target organ. Two distinct pathological conditions have been associated with exposure to OTA: Balkan endemic nephropathy (BEN), described as progressive karyomegalic interstitial nephritis resulting ultimately in complete renal failure, and urinary tract tumours (UTT) that have been reported to occur with a higher incidence in endemic areas of BEN (Fink- Gremmels 2005). Although an association between intake of OTA and BEN in humans has been postulated, causality has not yet been established (Walker and Larsen 2005). Based on food consumption in Europe, the Joint Expert Committee on Food Additives (JECFA) (2001) estimated the mean total OTA intake to be 45ngkg-1 body weight week-1, assuming a body weight of 60kg (Walker and Larsen 2005). The wide range of food types, the sporadic occurrence, and the low levels at which OTA is found make assessment of exposure through analysis of foods particularly problematic (Gilbert et al. 2001; Thuvander et al. 2001). The occurrence of OTA has not been studied in depth in Argentina and only a small number of food samples have been found to be positive at low levels in the country (Soleas et al. 2001; Rosa et al. 2004; Pacin et al. 2005). However, monitoring of blood or urine samples can give an indication of the overall situation on the occurrence of OTA in food in the local market. OTA levels in plasma and urine have to be considered as typical biomarkers of exposure (Gilbert et al. 2001) and no other appropriate effect-related biomarkers have as yet been identified (Fink-Gremmels 2005). The presence of OTA in human blood has been reported in many studies (Scott 2005), but none has been performed in Argentina. There is a strong need for a risk assessment of human exposure to mycotoxins, including OTA, in the South Cone countries. The aim of this preliminary study was to monitor OTA in human blood samples in two areas of the Buenos Aires province in Argentina, with a standardized methodology for OTA determination implemented in Latin America South Cone laboratories. Materials and methods Mar del Plata is an Argentine city located on the coast of the Atlantic Ocean in the Buenos Aires Province, 400km south of Buenos Aires. Mar del Plata is one of the major fishing ports whose industry concentrates on fish processing. The area is also host to other light industry, such as textile and food manufacturing, and the biggest seaside beach resort in Argentina. With a population of 541733 (2001 census, INDEC) it is the seventh largest urban area in Argentina. Due to the fact that this is a tourist city, the population increases during the summer season, occasionally quadrupling the number of residents. General Rodriguez is located 55km to the west of Buenos Aires, its population being 67931 (2001 census, INDEC). The main industry is a milk factory; secondarily the population carries out rural work. Taking into account the last census (2001, INDEC) and keeping in mind the sociodemographic indicator, it can be inferred that the population attending Vicente López y Planes Hospital at General Rodriguez has less resources than the population attending Zonal Hospital at Mar del Plata. Collection of blood samples Blood samples from donors were collected in two different localities of Buenos Aires province, Argentina, in February 2004 in the Regional Hemotherapy Center of Mar del Plata and between April and July 2005 in Vicente Lopez y Planes Hospital of General Rodríguez. The first step of the blood donation procedure in the hospitals was to ask donors to fill in a questionnaire which allows a physician to set apart some of them (e.g. those with low weight, the elderly, those suffering from hepatitis). Moreover, it is compulsory in Argentina to make human immunodeficiency virus (HIV) and Chagas virus analysis on the blood before including it in the blood bank. The numbers of samples were 205 for Mar del Plata and 275 for General Rodríguez. However, after excluding samples from donors with HIV and Chagas virus, the numbers of samples were 199 plasma samples from Mar del Plata and 236 from General Rodríguez. In Argentina it is more common to receive men's blood than women's donations. Table I shows information about the blood donors in both localities. Table I. Data of blood donors in Mar del Plata and General Rodríguez. Age Weight (years) (kg) Sample Female Male Female Male amount City Female Male Total Mean SD* Mean SD Mean SD Mean SD *Standard deviation. Mar del Plata 57 142 199 40 13 37 12 70 13 80 15 General 43 193 236 36 9 35 11 68 11 80 12 Rodríguez Ochratoxin A extraction and clean-up The blood samples were conditioned with the anticoagulant K3EDTA (Vacutainer) immediately after collection. The samples were kept frozen at -18°C until analyses. The plasma was analysed for the presence of ochratoxin A according to the method of Scott et al. (1998) with some modifications, as reported below. A total of 1ml of plasma was mixed with 0.25ml saturated sodium chloride solution and 5ml methanol with a vortex mixer, for 15s in a centrifuge tube and centrifuged at an average relative centrifugal force of 500g for 15min. The supernatant was transferred to another tube and mixed with 5ml 0.015M o-phosphoric before adding to a Bakerbond® C-18 cartridge (art. 7020), preconditioned with 10ml MeOH followed by 6ml methanol-(0.015M) o-phosphoric acid (1:1, v/v). Solvents passed through the column by gravity at a flow rate of one to two drops s-1. Following the addition of supernatant, the column was washed with 5ml 0.015M o-H3PO4 followed by 5ml methanol:(0.015M) o-phosphoric acid (1:1, v/v). MeOH (2ml) was added to the cartridge and allowed to stand for 3min before elution. The evaporated extract was dissolved in 3×2.5ml phosphate-buffered saline solution (PBS; a mixture of 0.26g monoacid sodium phosphate, 1.14g diacid sodium phosphate was dissolved separately and then added to 7.02g sodium chloride, 0.201g potassium chloride and 0.5g sodium azide adjusted to pH 7.4 and diluted to 1 litre with bi-distilled water)-MeOH (85:15) and added to an Ochraprep® column. All solvents were passed through the column by gravity flow. The column was washed with 5ml PBS solution-MeOH (85:15) followed by 10ml distilled water. OTA was eluted in two back flushing steps. First, with 3ml MeOH and, second, with 1.5ml MeOH into a silanized vial. The eluate were pooled and evaporated to dryness under vacuum, at 30°C. The evaporated extract was dissolved in 200µl mobile phase. An injection of 100µl of sample extract was analysed by HPLC as described below. The normal process to evaluate the methodology of OTA analysis includes three spiked concentrations in the linearity range (low, mid and high). As we needed a large sample size to prepare triplicate samples at each OTA level, a diagnosis laboratory was asked to collect blood remaining in samples to perform these recovery studies. The pools obtained were always contaminated with OTA, so it was impossible to work at a low level because of errors deriving from the initial contamination. Recoveries of spiked samples with OTA standard (Sigma Chemical Co., St Louis, MO, USA) were 85% for a contamination level of 2.4ngml-1; 95% for 1.5ngml-1; and 96% for 0.8ngml-1. The mean recovery rate was 89.8%. They were all in the range between 50 and 120% (European Commission 1998, 2002). Since there is no appropriate certified reference material available, only the recovery was determined, but not the trueness. The stability of test results was tested following a spiked control sample over the period of the study. Chart control is shown in Figure 1. [Enlarge Image] Figure 1. Control chart for ochratoxin A (OTA) (a) lower action limit (mean-3 sigma), and (b) upper action limit (mean+3 sigma). The range of linearity was between 0.012 and 7.12ngml-1 (r2=0.99978, n=7). When samples presented higher contamination, dilution of the samples was made with mobile phase and a new injection was made to quantify the OTA. Determination of OTA by HPLC The HPLC system used was an Agilent®1100 series which included a degasser (G1322A), an autosampler (G1313A), a fluorescence detector (G1321A), a quaternary pump (G1311A) and a thermostatted column compartment (G1316A). The used column was a C18 reverse-phase (4mm i.d.×125mm containing 5µm particle size, Hypersil BDS, Hypersil® with a guard column of the same phase (Hypersil BDS C18 4mm i.d.×4mm, 5µm). The mobile phase was acetonitrile-water-acetic acid (49.5:67:1, v/v/v). The flow rate was 1mlmin-1. Fluorescence excitation and emission wavelengths were set at 330 and 470nm, respectively. Retention times of OTA were in the range of 2-3min. Detection and quantification limits (LOD and LOQ) for OTA were 0.012 and 0.019ngml-1, respectively (signal-to-noise ratios of 3:1 and 5:1, respectively). Confirmation of OTA by methyl ester The confirmation of the presence of OTA was performed through the formation of ochratoxin A methyl ester. Slight modifications of the procedure of Grosso et al. (2003) were made. Briefly, a quantitative portion of the methanolic elution phase from the immunological column was evaporated to dryness and resuspended into 200µl of a 12% methanolic solution of boron trifluoride (Baker C701-07). After heating for 15min at 60°C, the derivative was analysed by HPLC with the same chromatographic conditions as for OTA. Confirmation of OTA as a methyl ester was performed on all contaminated samples. The retention time of the OTA methyl ester was approximately 16.3min. Detection and quantification limits expressed as OTA were calculated with signal-to-noise ratios of approximately 3:1 and 5:1, respectively (0.017 and 0.028ngml-1). Estimation of ochratoxin A intake based on plasma levels The mean of ochratoxin level in blood was used to estimate the continuous dietary intake according to Klaassen equations (Breitholtz et al. 1991; Thuvander et al. 2004): where k0 is the continuous dietary intake (ngkg body weight day ); and Cp is the plasma concentration of OTA (ngml-1). -1 -1 Statistical analysis The alpha Winsorized mean with a percentage of substitution (or replacement) equal to 20 was used. This measure substitutes a percentage of extreme values for the last not replaced (Huber 1964; Hampel 1968; García Perez 2002). The Winsorized mean is a robust estimator of the population mean that is relatively insensitive to outlying values. Therefore, Winsorization is a method for reducing the effects of extreme values in the sample. The ktimes Winsorized mean is calculated as: Denoting X(1), ,X(n), the ordered sample, the Winsorized mean ( ), is computed replacing the 20\\\% smallest observations and the 20\\\% largest observations by X(k+1) and X(n-k), respectively, where k+1 is the index of the order statistic that leaves 20\\\% of the sample to its left. For a symmetrical distribution, the Winsorized mean is an unbiased estimate of the population mean. However, the Winsorized mean does not have a normal distribution even if the data are from a normal population. The sample standard deviation (SD) is a commonly used estimator of the population scale. However, it is sensitive to outliers. With robust scale estimators the estimates remain bounded even when a portion of the data points is replaced by arbitrary numbers. The Winsorized sum-of-squared deviations is defined as: The square root of swk² is noted as the Winsorized SD. To test the hypothesis that the means of the two groups from which the samples were drawn are equal, a Winsorized t-test was used (SAS Institute 1999). The statistic of this test is given by: where is the standard error of : S-Plus 7.0 software was used for the statistical analysis. To calculate the Winsorized mean, values for non-detected samples were assigned as LOD/2 and samples with OTA contamination levels between LOD and LOQ were reported with the obtained values. Results and discussion Table II shows the OTA distribution of the plasma samples in both areas of the Buenos Aires province. The results reveal that 63.8% (64.1% male, 63.2% female) from Mar del Plata and 62.3% (63.2% male, 58.1% female) from General Rodríguez of human plasmas sampled were OTA-positive. The highest concentration in Mar del Plata was 47.6ngml-1; and 74.8ngml-1 in General Rodríguez. Table II. Frequency and level of ochratoxin A contamination in blood plasma samples from two areas of Buenos Aires province. General Rodriguez Mar del Plata -1 Ochratoxin A (ngml ) Male Female Male Female -1 n.d., not detected. Limit of detection (LOD)=0.012ngml ; limit of quantitation (LOQ)=0.019ngml-1. n.d. 71 18 51 21 LOD to <LOQ 0 0 1 0 LOQ to 0.2 16 9 39 17 >0.2 to 0.4 25 3 19 7 >0.4 to 0.6 17 5 11 3 >0.6 to 0.8 14 2 5 3 >0.8 to 1.0 5 0 4 0 >1.0 to 1.2 2 0 2 1 >1.2 to 1.4 8 2 2 2 >1.4 to 1.6 5 1 0 0 >1.6 to 1.8 1 0 0 0 >1.8 to 2.0 1 0 1 0 >2.0 to 2.2 2 0 0 0 >2.2 to 2.4 0 0 1 0 >2.4 to 2.6 0 0 1 0 >2.6 to 2.8 3 1 1 0 >2.8 to 3.0 0 0 1 1 >3.0 to 3.4 3 0 2 0 >3.4 to 3.8 3 0 0 0 >3.8 to 4.2 0 0 0 0 >4.2 to 4.6 3 1 0 0 >4.6 to 5.0 0 0 0 0 >5.0 to 6.0 3 0 0 0 >6.0 to 7.0 2 0 0 0 >7.0 to 8.0 0 0 0 1 >8.0 to 9.0 2 1 1 0 >9.0 to 10.0 3 0 0 0 >10.0 to 20.0 1 0 0 0 >20.0 to 30.0 1 0 0 0 >30.0 to 40.0 1 0 0 0 >40.0 to 50.0 0 0 0 1 >50.0 to 60.0 0 0 0 0 >60.0 to 70.0 0 0 0 0 >70.0 to 80.0 1 0 0 0 Robust methods were used due to the fact that variables did not present a normal distribution (see Table II). These methods try to reduce the possible influence of the extreme data of a distribution. The median (measured of position not affected by extreme values) is one of the more used robust estimators; the OTA medians were 0.11ngml-1 in Mar del Plata and 0.24ngml-1 in General Rodriguez for the entire population. Other robust estimators such as the mean alpha Winsorized, the average alpha trimmed, or the estimator of location of Huber are also used. In the literature a criterion to select one of these estimators is proposed based on choosing the one that presents minor variance. The mean alpha Winsorized is recommended (García Perez 2002). Because of the presence of several samples with high OTA concentration, the Winsorized measures and the t-test based on them were used to compare the populations. Significant differences were found between both populations (Table III). Table III. Ochratoxin A (OTA) plasma concentrations (ngml-1) of the analysed population for the different age groups and comparison between the two areas of Buenos Aires province. Mar del General Plata Rodriguez Mean Population Mean* SD** n Mean SD n p differences *Winsorized mean. **Winsorized standard deviation. All 0.1537 0.1700 199 0.4319 0.4919 236 -0.2783 <0.01 Age 210.0940 0.1130 47 0.4640 0.5251 70 -0.3701 <0.01 group 30 310.2506 0.2735 53 0.4231 0.4846 71 -0.1725 <0.05 40 410.1627 0.1700 58 0.2675 0.3054 56 -0.1048 <0.05 50 510.1201 0.1311 24 1.7420 2.6191 19 -1.6219 <0.05 60 To study in detail the differences, four age groups were compared. Table III includes the results corresponding to these comparisons. Significant differences between both cities were found in all age groups. Considering the comparison of OTA levels between women and men in these areas of Buenos Aires province, it can be observed in Table IV that only in General Rodriguez was the difference significant. Table IV. Ochratoxin A (OTA) plasma Winsorized mean concentrations (ng ml-1): Comparison between males and females in two areas of the Buenos Aires province. Females Males Mean Population Mean* SD** n Mean SD n p differences *Winsorized mean. **Winsorized standard deviation. General 0.2059 0.2260 43 0.4752 0.5217 193 0.2693 <0.001 Rodriguez Mar del Plata 0.1524 0.1595 57 0.1644 0.1707 142 -0.1198 0.64 Preliminary estimated dietary intakes of OTA were made by using the Klassen formula (Breitholtz et al. 1991; Thuvander et al. 2004) using the factor 1.97 (or 1.34) and concentration values in both areas. With the alpha Winsorized mean (Table III), the dietary intake of OTA was calculated 0.30 (0.21) and 0.84 (0.58)ngkg-1 body weight day-1 in Mar del Plata and General Rodriguez, respectively. To compare the dietary intake estimation in these areas of Argentina with others it was necessary to recalculate considering the OTA median values. The results were 0.21 (0.15) and 0.47 (0.32)ngkg-1 body weight day-1 in Mar del Plata and General Rodriguez, respectively; and Mar del Plata had values smaller than all the countries present in SCOOP studies, where the General Rodriguez estimation is similar to the intake of Germany (Miraglia and Brera, 2002). The dietary intake estimation in these areas per week, calculated with the 95th percentile of OTA in plasma using the Klassen equations, was 32.0 (22.0) and 84.2 (57.3)ngkg-1 body weight week-1 in Mar del Plata and General Rodriguez, respectively; that is, below the provisional tolerable weekly intake (PTWI) of 100ngkg-1 body weight week-1 established in 2001 by JECFA. In any case, it is necessary to take into consideration that some high levels of OTA in plasma could exceed the suggested PTWI. Conclusions Argentinean people, represented in this study as the two areas of the Buenos Aires region, are exposed to OTA with values smaller or similar to those found in various European countries. Considering that the weight, age and race of both populations is very similar, the differences found in the presence of OTA in plasma could be attributed to various factors; the fact that the samples were obtained in different periods: February 2004 from Mar del Plata and April-July 2005 from General Rodriguez. It is known that fluctuations exist, not only seasonal fluctuations, but also regional ones as well as those due to individuals. Another factor could be that the population, and therefore the donors, at the General Rodriguez Hospital presents a lower socio-economical level than those in Mar del Plata Hospital. This difference could imply the intake of lower quality foods (Goodwin et al. 2006) and consequently containing a higher level of OTA contamination. Acknowledgements The authors acknowledge the technical assistance provided by Ms Gabriela Cano and Daniela Taglieri, as well as the financial support from the Comisión de Investigaciones Científicas of the Province of Buenos Aires, Universidad Nacional de Luján, Universidad de Buenos Aires, CONICET, BID 1201/OC-AR PICTOR 2002-00012, Argentina, and the European Union (Contract No. ICA4-CT-2002-10043). Special thanks are also directed to Mr Simon Bevis (RBiopharm Rhone Ltd) for the support including the Ochraprep® columns. References 1. Breitholtz, A., Olsen, M., Dahlbäck, Å and Hult, K. (1991) Plasma ochratoxin A levels in three Swedish populations surveyed using an ion-pair HPLC technique. Food Additives and Contaminants 8 , pp. 183-192. 2. European Commission (1998) Directive 98/53/EC of 16 July 1998 laying down the sampling methods and the methods of analysis for the official control of the levels for certain contaminants in foodstuffs. Official Journal of the European Commission L201 , pp. 93-101. 3. European Commission (2002) Decision 2002/657/EC of 12 August 2002 implementing Council Directive 96/23/EC concerning the performance of analytical methods and the interpretation of results. Official Journal of the European Commission L221 , pp. 8-36. 4. Fink-Gremmels, J. (2005) Conclusions from the workshops on Ochratoxin A in food: Recent developments and significance, organized by ILSI Europe in Baden (Austria), 29 June-1 July 2005. Food Additives and Contaminants 22:Suppl. 1 , pp. 15. 5. Perez, A García (2002) Métodos avanzados de estadística avanzada. Madrid: UNED pp. 32-36. Universidad Nacional de Educación a Distancia - 68-70 6. Gilbert, J., Brereton, P. and MacDonald, S. (2001) Assessment of dietary exposure to Ochratoxin A in the UK using a duplicate diet approach and analysis of urine and plasma simples. Food Additives and Contaminants 18 , pp. 1088-1093. 7. Goodwin, DK, Knol, LK, Eddy, JM, Fitzhugh, EC, Kendrick, O. and Donohue, RE (2006) Sociodemographic correlates of overall quality of dietary intake of US adolescents. Nutrition Research 26 , pp. 105-110. 8. Grosso, F., Saïd, S., Mabrouk, I., Fremy, JM, Castegnaro, M., Jemmali, M. and Dragacci, S. (2003) New data on the occurrence of Ochratoxin A in human sera from patients affected or not by renal disease in Tunisia. Food and Chemical Toxicology 41 , pp. 1133-1140. 9. Hampel, FR (1968) Contribution to the theory of robust estimation. Dissertation Department of Statistics, University of California - No. 119 10. Huber, PJ (1964) Robust estimation of a location parameter. Anales de Estadística Matemática 35 , pp. 73-101. 11. Joint Expert Committee on Food Additives (JECFA) (2001) Ochratoxin A. Safety evaluation of certain mycotoxins in food, prepared by the 56th Meeting of the Joint FAO/WHO Expert Committee on Food Additives World Health Organization pp. 281-387. Geneva - WHO Food Additives Series No. 47 12. Miraglia, M. and Brera, C. (2002) Assessment of dietary intake of Ochratoxin A by the population of EU Member States. Report of SCOOP task 3.2.7 - January. Available http://europaeu.int/comm/food/fs/scoop/3.2.7en.pdf 13. Pacin, A., Resnik, S., Vega, M., Saelzer, R., Bovier, E Ciancio, Ríos, G. and Martinez, N. (2005) Occurrence of ochratoxin A in wines in the Argentinean and Chilean markets. ARKIVOC 12 , pp. 214-223. 14. Rosa, CAR, Magnoli, CE, Fraga, ME, Dalcero, AM and Santana, DMN (2004) Occurrence of ochratoxin A in wine and grape juice marketed in Rio de Janeiro, Brazil. Food Additives and Contaminants 21 , pp. 358-364. 15. Institute, SAS (1999) SAS OnlineDoc®, Version 8 SAS Institute Inc. , Cary, NC Available: http://www.csc.fi/cschelp/sovellukset/stat/sas/sasdoc/sashtml/insight/chap38/sect17.ht m 16. Scott, PM (2005) Biomarkers of human exposure to ochratoxin A. Food Additives and Contaminants 22:Suppl. 1 , pp. 99-107. 17. Scott, PM, Kanhere, SR, Lau, BP, Lewis, DA, Hayward, S., Ryan, JJ and Kuiper-Goodman, T. (1998) Survey of Canadian human blood plasma for ochratoxin A. Food Additives and Contaminant 15 , pp. 555-562. 18. Soleas, GJ, Yan, J. and Goldberg, DM (2001) Assay of ochratoxin A in wine and beer by high-pressure liquid chromatography photodiode array and gas chromatography mass selective detection. Journal Agricultural Food Chemistry 49 , pp. 2733-2740. 19. Thuvander, A., Möller, T., Barbieri, HE, Jansson, A., Salomonsson, A-C and Olsen, M. (2001) Dietary intake of some important mycotoxins by the Swedish population. Food Additives and Contaminants 18 , pp. 696-706. 20. Thuvander, A., Paulsen, JE, Axberg, K., Johansson, N., Vidnes, A., EnghardtBarbieri, H., Tressou, J., Leblanc, J-Ch, Feinberg, M. and Bertail, P. (2004) Statistical methodology to evaluate food exposure to a contaminant and influence of sanitary limits: Application to Ochratoxin A. Regulatory Toxicology and Pharmacology 40 , pp. 252-263. 21. Walker, R. and Larsen, JC (2005) Ochratoxin A: Previous risk assessments and issues arising. Food Additives and Contaminants 22(Suppl. 1) , pp. 6-9. List of Figures [Enlarge Image] Figure 1. Control chart for ochratoxin A (OTA) (a) lower action limit (mean-3 sigma), and (b) upper action limit (mean+3 sigma). List of Tables Table I. Data of blood donors in Mar del Plata and General Rodríguez. Age (years) Weight (kg) Sample Female Male Female Male amount City Female Male Total Mean SD* Mean SD Mean SD Mean SD *Standard deviation. Mar del Plata 57 142 199 40 13 37 12 70 13 80 15 General 43 193 236 36 9 35 11 68 11 80 12 Rodríguez Table II. Frequency and level of ochratoxin A contamination in blood plasma samples from two areas of Buenos Aires province. General Rodriguez Mar del Plata -1 Ochratoxin A (ngml ) Male Female Male Female -1 n.d., not detected. Limit of detection (LOD)=0.012ngml ; limit of quantitation (LOQ)=0.019ngml-1. n.d. 71 18 51 21 LOD to <LOQ 0 0 1 0 LOQ to 0.2 16 9 39 17 >0.2 to 0.4 25 3 19 7 >0.4 to 0.6 17 5 11 3 >0.6 to 0.8 14 2 5 3 >0.8 to 1.0 5 0 4 0 >1.0 to 1.2 2 0 2 1 >1.2 to 1.4 8 2 2 2 >1.4 to 1.6 5 1 0 0 >1.6 to 1.8 1 0 0 0 >1.8 to 2.0 1 0 1 0 >2.0 to 2.2 2 0 0 0 >2.2 to 2.4 0 0 1 0 >2.4 to 2.6 0 0 1 0 >2.6 to 2.8 3 1 1 0 >2.8 to 3.0 0 0 1 1 >3.0 to 3.4 3 0 2 0 >3.4 to 3.8 3 0 0 0 >3.8 to 4.2 0 0 0 0 >4.2 to 4.6 3 1 0 0 >4.6 to 5.0 0 0 0 0 >5.0 to 6.0 3 0 0 0 >6.0 to 7.0 2 0 0 0 >7.0 to 8.0 0 0 0 1 >8.0 to 9.0 >9.0 to 10.0 >10.0 to 20.0 >20.0 to 30.0 >30.0 to 40.0 >40.0 to 50.0 >50.0 to 60.0 >60.0 to 70.0 >70.0 to 80.0 2 3 1 1 1 0 0 0 1 1 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0 0 Table III. Ochratoxin A (OTA) plasma concentrations (ngml-1) of the analysed population for the different age groups and comparison between the two areas of Buenos Aires province. Mar del General Plata Rodriguez Mean Population Mean* SD** n Mean SD n p differences *Winsorized mean. **Winsorized standard deviation. All 0.1537 0.1700 199 0.4319 0.4919 236 -0.2783 <0.01 Age 210.0940 0.1130 47 0.4640 0.5251 70 -0.3701 <0.01 group 30 310.2506 0.2735 53 0.4231 0.4846 71 -0.1725 <0.05 40 410.1627 0.1700 58 0.2675 0.3054 56 -0.1048 <0.05 50 510.1201 0.1311 24 1.7420 2.6191 19 -1.6219 <0.05 60 Table IV. Ochratoxin A (OTA) plasma Winsorized mean concentrations (ng ml-1): Comparison between males and females in two areas of the Buenos Aires province. Females Males Mean Population Mean* SD** n Mean SD n p differences *Winsorized mean. **Winsorized standard deviation. General 0.2059 0.2260 43 0.4752 0.5217 193 0.2693 <0.001 Rodriguez Mar del Plata 0.1524 0.1595 57 0.1644 0.1707 142 -0.1198 0.64 Privacy Policy | Terms & Conditions | Accessibility | RSS FAQs in: English . Français . Español . © 2008 Informa plc