Agricultural and Food Science, Vol. 15 (2006): –11. � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Vol. 15 (2006): �–11. © Agricultural and Food Science Manuscript received December 2005 Cadmium content of Finnish linseed and estimated  consumer intake Hanna-Riitta Kymäläinen and Anna-Maija Sjöberg Department of Agrotechnology, PO Box 28, FI-00014 University of Helsinki, Finland, e-mail: hanna-riitta.kymalainen@helsinki.fi The aim of this study was to screen the cadmium (Cd) contents of seed and seed crush of Finnish linseed and to evaluate the acceptable daily dose. The Cd content of Finnish linseed is of interest, since some prop- erties of linseed as the content of fatty acids are different in northern growth locations compared with other growing areas. Seed samples (N = 85) were collected from the experimental farm of Helsinki University and from commercial fields in different locations in southern Finland and Ostrobothnia. In addition, 15 samples of linseed crush were examined. The cadmium contents of the linseed samples, determined with atomic absorption spectroscopy, varied between 0.27 and 1.3 mg kg-1 (dry weight) (mean 0.61 mg kg-1). The Cd contents of the crush samples were between 0.47 and 1.5 mg kg-1 (mean 0.85 mg kg-1). There were dif- ferences between the Cd contents of the three examined varieties and years at the experimental farm. In- crease in nitrogen fertilization somewhat increased the Cd uptake of linseed. The permitted intake of lin- seed depends on body weight and on the Cd content of the product and is in most cases higher than the di- etary recommendation (24–30 g daily dose of linseed). Key words: linseed, Linum usitatissimum, cadmium, fertilizers, varieties, intake Introduction Linseed (Linum usitatissimum L.) has several pos- itive effects on human health, e.g. favourable fatty acids, proteins and dietary fibre (Morris 2003). It has industrial interest as a functional food or as an ingredient of functional foods e.g. in the form of oil and crush (Oomah 2001). However, despite its numerous advantageous components promoting human health, linseed also contains some com- pounds not beneficial to health, including cadmi- um taken up from soil. In the form of the divalent ion Cd2+, cadmium is readily taken up by linseed even at low external concentrations (Chakravarty and Srivastava 1997). However, a wide variation � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Kymäläinen, H.-R. & Sjöberg, A.-M. Cadmium content of linseed of Cd content in the grain of linseed has been measured in various international studies (Table 1). Published results concerning the Cd content of Finnish linseed were not available. They, however, are of interest, since some properties of linseed as the content of fatty acids are different in northern growth locations compared with other growing ar- eas (Kanta-Oksa 1992). According to Codex Alimentarius Commission (2001), the maximum Cd content for cereals is 0.1 mg kg-1 (dw, dry weight,), which corresponds to 0.11 mg kg-1 (dw) (moisture content of the seed between 5 and 13%, wet basis). In Finland there is no official limit value of Cd for the food products (Tarpila et al. 2005). The German guide value for the Cd content of linseed is 0.3 mg kg-1 (fw, fresh weight) (Li et al. 1997). The maximum permitted concentration for confectionery linseed traded on the international market is 0.25 mg kg-1 in Aus- tralia (Hocking and McLaughlin 2000). Similar values are used in various European countries (Liukkonen-Lilja and Penttilä 1992). For non- smokers food is the main source of Cd (WHO 2000). However, in areas without heavy industrial pollution, cadmium exposure is mainly influenced by personal smoking habits (Heinzow et al. 1991). The Finnish estimated daily intake (EDI) is 10.8 μg per person according to Tahvonen (1995). In a study by Leblanc et al. (2000), the average French EDI for Cd was 17 µg per person. An average di- etary intake of Cd in some polluted Polish towns was 40 µg per person (Skibniewska 2003), which was considered alarming. The aim of this study was to screen the cadmi- um contents of seed and seed crush of Finnish lin- seed and to evaluate their acceptable daily intakes for humans. In addition, the differences between varieties and the effect of nitrogen fertilization on the cadmium contents were investigated. Material and methods A total of 24 seed samples were collected from the experimental farm of the University of Helsinki, located in Siuntio in southern Finland. As shown in Table 2, the samples were collected from differ- ent varieties and nitrogen fertilization levels with three replicates. In addition, 61 single seed sam- ples were collected from commercial fields in dif- ferent locations of southern Finland and Ostro- bothnia. A total of 15 samples of linseed crush, produced from some of the commercial linseed samples were also examined. The crush samples were the residue after pressing oil from the seeds. In order to examine the possible relation of various parameters to the Cd levels, the following back- ground data of the samples were collected: type and amount of fertilization, soil type, preceding crop and use of pesticides. Unfortunately this background data was not available for 13 commer- cial samples and six sowing seed samples, as is shown in Table 2. The Cd contents of the samples were analysed by the City of Vantaa Food and Environmental Table 1. Studies concerning cadmium content of linseed. Examined linseed samples Cd content, mg kg-1 Reference Yellow mean 0.23 Klein & Weigert 1987 Brown mean 0.38 Klein & Weigert 1987 118 seed samples and 16 genotypes 0.10–1.70 (dw) Marquard et al. 1990 14 commercial varieties 0.02–0.10 (fw) Li et al. 1997 60 plant introduction lines 0.14–1.37 (fw) Li et al. 1997 Cultivars grown under experimental conditions 0.23 and 0.55 (dw), no differences between the brown and yellow seeded varieties Hocking & McLaughlin 2000 dw = dry weight, fw = fresh weight � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Vol. 15 (2006): �–11. Table 2. Linseed samples, areal origin and year of harvesting. Areal origin of the samples Year of harvesting 2002 2003 2002 or 2003* 2004 Experimental farm of Helsinki University, Siuntio (southern Finland) 3 seed samples: 3 replicates of var. Laser 9 seed samples: 3 replicates of varieties Helmi, Laser and the Bor breeding line – 12 seed samples: 9 replicates of var. Laser: 3 replicates for each of the N fertilisation levels of 40, 60 and 80 kg ha-1. 3 replicates of var. Helmi: N fertilisation level of 60 kg ha-1. Southern Finland, commercial farms 3 seed samples 1 crush sample 25 seed samples 9 crush samples 8 seed samples 5 crush samples – Ostrobothnia, commercial farms 8 seed samples 17 seed samples – – Unknown areal origin of sowing seed samples – – 6 seed samples: 3 replicates of sowing seed of Helmi and the Bor breeding line – – No samples * exact year not available Laboratory, an accredited laboratory. The method was the standard method NMKL 161, based on atomic absorption spectroscopy. Samples were ho- mogenized and combusted in a wet combustion apparatus (Milestone Ethos Plus) with nitric acid and hydrogen peroxide. Also zero tests without sample were carried out for each combustion. The samples were analyzed with atomic absorption spectroscopy in a graphite oven (Perkin Elmer Si- maa 6100) with background correction (Zeeman). A reference material BCR No 279 (Sea Lettuce) was used. The uncertainty of the measurements was ±18% (95% confidence interval) in the range between 0.3 mg kg-1 and 1.5 mg kg-1. Variance analysis (One-way ANOVA, Tukeys’s post hoc test) of the SPSS statistical tool was used to examine the possible differences in cadmium content of the samples between the growing areas southern Finland and Ostrobothnia. Bivariate cor- relation analysis of the SPSS tool was used to ex- amine the correlation between the Cd content and nitrogen fertilization level of the seed samples. Results Cadmium content of linseed The Cd content of the linseed samples varied be- tween 0.27 and 1.3 mg kg-1 (dw), with a mean value of 0.62 mg kg-1. The Cd contents of the crush samples were between 0.47 and 1.5 mg kg-1 (dw), with a mean of 0.85 mg kg-1. As shown in Fig. 1, the Cd contents of seeds grown in the Ostrobothnia area were somewhat lower (mean 0.48 mg kg-1) than those of seeds grown in southern Finland (mean 0.67 mg kg-1). Despite the internal variation within the two areas, seen as a variation range of 0.27–1.3 mg kg-1 for the southern samples and 0.27–0.96 mg kg-1 for the northern samples, the difference between the two major areas was statis- tically significant (P < 0.005). However, because the samples from Siuntio had a higher Cd content than all the other southern samples (Fig. 1), the ar- eal differences were analyzed further. The variance � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Kymäläinen, H.-R. & Sjöberg, A.-M. Cadmium content of linseed analysis showed statistically significant differenc- es between Siuntio and Ostrobothnia but also be- tween Siuntio and the other southern areas (P value in the Tukey’s test <0.001 in both cases). When Siuntio was excluded from the southern samples, there was no statistically significant difference be- tween Ostrobothnia and southern Finland (P > 0.05). The cadmium content of the crush samples, produced from other southern samples than those from Siuntio, was at the same level as that of the seed samples of Siuntio (Fig. 1). However, as shown in Fig. 2, the Cd contents of the all crush samples were higher than those of the correspond- ing seed samples from the commercial farms. The increase in the Cd content in the seed residue after pressing of oil was 56% on average, varying from 12% (Sample 9) to 138% (Sample 15). Figure 3 provides an indication of the differ- ences between the Cd contents of the varieties and years. In the year 2003, the Bor line had the high- est and ‘Laser’ the lowest Cd content. However, in 2004 there was no difference between ‘Laser’ and ‘Helmi’. As presented in Fig. 3, at the experimen- tal farm of Siuntio the increase in nitrogen fertili- zation level somewhat increased the Cd intake of 0.00 0.10 0.20 0.30 0.40 0.50 0.60 0.70 0.80 0.90 1.00 Ostrobothnia, seed Excluding Siuntio, seed and Ostrobothnia, Seed All, seed Crush Siuntio, seed S ou th Cd content mg kg-1 (dw) Fig. 1. Cadmium contents of lin- seed samples from different growing areas (dw = dry weight). Columns are means and bars in columns standard errors of the means of the replicates. Number of replicates is presented in Table 2. Fig. 2. Comparison of the cadmi- um contents of the grouts and of the corresponding seed samples from commercial farms. Results are single measurements of the samples. 0 0.2 0.4 0.6 0.8 1 1.2 1.4 1.6 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 sample nr Cd content -1 mg kg (dw) seed crush � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Vol. 15 (2006): �–11. ‘Laser’ in 2004. Only a weak general correlation between Cd content and the nitrogen fertilization level was found when all seed samples with a known nitrogen fertilization background data (N = 72) were analyzed (Pearson’s correlation coeffi- cient 0.41, P < 0.01). The types of fertilization, soil types, preceding crops and use of pesticides were extremely variable and could not be used for ex- plaining the Cd contents of the samples. Use of organic fertilization (manure or similar) was not associated with high Cd values, but the number of samples was too low to allow definite conclu- sions. Estimated intake In a clinical study by Tarpila et al. (2004), the blood cadmium of patients remained unchanged after six months of linseed treatment, and ground flaxseed was considered safe at a daily dose of up to 24 g. According to Morris (2003), a daily dose of 30 g of linseed is optimal for utilization of func- tional components as fatty acids. These recom- mended values for daily dose of linseed were used as references for intake estimations. By comparing the Cd values from the present study (seed 0.27– 1.3 mg kg-1, crush 0.47–1.5 mg kg-1) with the 24 g reference values, the estimated daily intake of Cd is 6.5–31 µg Cd (seed) and 11–36 µg Cd (crush). These values are 9.3–44% (seed) and 14–51% (crush) of the provisional tolerable daily intake (PTDI) value (70 μg d-1) presented by WHO and FAO (1993). By comparing the Cd values from the present study with the 30 g reference values, the EDI of Cd is 8.1–39 µg (seed) and 14–45 µg (crush). These values are 11–55% (seed) and 20– 63% (crush) of the 70 μg d-1 PTDI value. If all the Cd consumed by an individual origi- nated from the linseed studied here, the maximum daily dose related to the PTDI value of WHO would be 55–263 g linseed seed or 47–151 g lin- seed crush for a person with a 70 kg body weight, depending on the Cd content of linseed. However, because linseed and linseed products are consumed only in small amounts, the Cd intake should be evaluated accordingly. Furthermore, the body weights of different people vary considerably, leading to a need for personal recommendations of intakes e.g. in health advisory institutions. There- fore the body weights 50 kg, 70 kg and 90 kg are used in the following. The Finnish maximum EDI 0.00 0.20 0.40 0.60 0.80 1.00 1.20 1.40 Laser 2002 (60) Laser 2003 (60) Laser 2004 (40) Laser 2004 (60) Laser 2004 (80) Helmi 2003 (60) Helmi 2004 (60) Bor line 2003 (60) Cd content mg kg-1 (dw) Fig. 3. Cadmium contents of vari- eties Laser, Helmi and the line Bor grown in Siuntio, southern Finland. Columns are means and bars in columns the variation range (min-max) of the three rep- licates. The N fertilization levels of 40 kg ha-1, 60 kg ha-1 and 80 kg ha-1 are given in parentheses. � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Kymäläinen, H.-R. & Sjöberg, A.-M. Cadmium content of linseed of 11 μg in food is equivalent to 15% of PTDI (Tahvonen 1995), and the French EDI of 17 µg (Leblanc et al. 2000) corresponds to 24% of PTDI (70 µg, WHO). Similarly, a high EDI of 40 µg (Skibniewska 2003) corresponds to 57% of PTDI. If we injest Cd from linseed in addition to this ba- sal EDI (11–40 µg), the possible additional daily intake of Cd would be 30–60 µg. As can be seen in Figure 4, the additional permitted intake of linseed strongly depends on body weight. In most cases the permitted intake is even higher than that asso- ciated with the dietary recommendation of 24–30 g linseed, but in the case of a relatively low body weight (50 kg) the limit is rather close if linseed with a high Cd content is used. Discussion The Cd contents of most seed samples were be- tween 0.30 mg kg-1 and 0.90 mg kg-1 (dw). A few samples had a Cd content below 0.30 mg kg-1, which is the German limit value for linseed. All values were above the Codex Alimentarius Com- mission limit of 0.1 mg kg-1, which however is set for cereals that are consumed in large amounts compared with linseed. Cadmium accumulation in the seed of linseed may be influenced by a number of factors, e.g. fer- tilizer application (Cd content of the fertilizer), use of sewage sludge, soil type and pH, and agronomic practices (Guo et al. 1995, Hocking and McLaugh- lin 2000). In a study by Jiao et al. (2004), zinc ad- dition at 20 mg Zn kg-1 soil with phosphorus de- creased seed Cd concentration by 42%. Low soil pH can increase soil Cd solubility (Eriksson et al. 1996). In a study of Matthäus and Zubr (2000), the combined effect of climatic and soil conditions af- fected the accumulation of heavy metals in cameli- na seed. In a study of oats by Eurola et al. (2003), the samples with highest Cd values had grown in dry, warm conditions leading to low biomass and a relatively high concentration of Cd in the plant tis- sues. In the present study, the warm growing sea- son of the year 2003 led to a higher Cd content in the variety Helmi than the wet growing season of 2004 at the experimental farm of Siuntio, whereas there was no corresponding difference in the vari- ety Laser. In the present study, the linseed grown in the northern province of Ostrobothnia had a somewhat lower Cd content on average than linseed grown in southern Finland. However, the Cd contents of the samples from Siuntio were significantly higher than those of the samples from the other southern locations. Reason for that should be investigated. In the study by Eurola et al. (2003), the lowest Cd values of oats were measured in the northern trial locations in Finland. The Cd content of Finnish soils is higher in southern than in northern parts of the country (Sippola and Mäkelä-Kurtto 1986, Louekari et al. 2000), which affects the Cd content of cereals accordingly (Liukkonen-Lilja and Pent- a) seed 0 20 40 60 80 100 120 140 50 70 90 body weight (kg) Additional daily intake of linseed (g) b) crush 0 20 40 60 80 100 120 140 50 70 90 body weight (kg) Additional daily intake of linseed (g) EDI 11 EDI 17 EDI 40 Fig. 4. Estimated average intake of seed and crush of lin- seed for persons with different body masses, when linseed is considered as an additional source of cadmium. The es- timates are calculated from means of the cadmium con- tents of (a) seed and (b) crush for estimated daily intake values of 11 μg (Finnish), 17 μg (French value) and 40 μg (polluted Polish area). � A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Vol. 15 (2006): �–11. tilä 1992). In the present study this result was not confirmed for all southern samples. Flax used for food applications should not be grown in polluted areas (Marquard et al. 1990, Grant et al. 2000, An- gelova et al. 2003). In the study by Angelova et al. (2003), cadmium contents as high as 2.3 mg kg-1 were measured in flaxseed at a distance of 0.5 km from an industrially polluted area, whereas 15 km from the polluted area the flax seed accumulated only 0.52 mg kg-1. Industrial emission sources of Cd include e.g. the mining, steel and fertilizer in- dustries and energy production (Liukkonen-Lilja and Penttilä 1992). This should direct the selection of growing areas for linseed, although in practice a consumer will probably not consistently eat lin- seed with the highest contaminant level. As was seen in the present study and in other studies (Marquard et al. 1990, Li et al. 1997), cad- mium uptake of linseed from the soil is clearly cul- tivar dependent. The results indicate that it is pos- sible to select for low Cd uptake genotypes in fu- ture breeding programs. Cadmium interacts with e.g. zinc ions, and Chakravarty and Srivastava (1997) showed that by adding Zn2+ to a growth medium at a non-toxic level it is possible to regen- erate linseed plants tolerant to Cd2+. Hocking and McLaughlin (2000) also suggested utilization of genotypic differences in reducing Cd concentra- tions in linseed, because many of the soil and site factors are difficult for producers to control. How- ever, more investigations are needed for develop- ing varieties with this feature. The Cd content of the seed residue after sepa- ration of oil was higher than that of the whole seed. In a study by Hocking and McLaughlin (2000), the cadmium contents of the de-seeded capsules and the seeds of linseed varied only slightly compared to other oilseed plants, and ineffective barriers dis- criminating against the transport of Cd to seed were proposed. In a study of Jiao et al. (2004), in- effective barriers discriminating against the trans- port of Cd from the root to the shoot, and restric- tion of Cd translocation to the seed via the phloem were found. Li et al. (1997) suggested that there is no relationship between the oil content of a crop and its patterns of Cd accumulation. Cadmium did not penetrate markedly into the pressed oil of camelina (Matthäus and Zubr 2000). In accord- ance with this result, Lei et al. (2003) showed that the major Cd-binding fraction in the seed of lin- seed is associated with one part of the proteins, and proposed that it could be possible to isolate the major storage protein of flaxseed with a low Cd content. However, an industrially feasible separa- tion method would be necessary (Oomah and Mazza 1993). Several limit values are used for evaluation of the maximum human intake of harmful chemical substances. The acceptable daily intake (ADI) val- ue relates to lifelong use. Short-term intakes can be accepted at exposure levels exceeding the ADI. Some contaminants have tolerable daily intake and provisional tolerable weekly intake (PTWI) values (Nordberg 1999). In the cited study, exposures to 200 µg daily intakes for months or even years were estimated to be tolerated without obvious gastroin- testinal symptoms or other indications. On the other hand, in a study of Meberg et al. (1979), the daily amount of Cd which can cause accumulation in the body was considered to be 200–300 µg. Ac- cording to WHO and FAO (1993), the present PTWI for Cd is 500 μg, often expressed as a provi- sional tolerable daily intake of 70 μg or 1 μg kg-1 body weight. Conclusion Cadmium content of the Finnish linseed was in general similar than what has been measured in previous international studies, although there was marked variation both in the present and previous studies. Samples taken from Siuntio were signifi- cantly higher than those of the samples from the other southern locations, and the reason for that should be studied. In the case of a normal Finnish diet with a relatively low EDI (11 μg), the nutri- tionally recommended daily dose of linseed (24– 30 g) can safely be consumed if the present PTDI value set by WHO is applied. If the normal diet contains a large amount of Cd or the body weight of the person is low, the recommended daily doses 10 A G R I C U L T U R A L   A N D   F O O D   S C I E N C E Kymäläinen, H.-R. & Sjöberg, A.-M. Cadmium content of linseed should be lowered at least when using linseed with a high Cd content. Growing areas with a low Cd content should be favoured in the cultivation of linseed. There was some evidence in the present study of an increase of Cd content of linseed due to increased nitrogen fertilization. Furthermore, the differences between linseed varieties should be considered when selecting varieties for breeding and cultivation. Acknowledgements. We are grateful to the Agro Fibre Net- work project in the EMOTR/ALMA program for financial support. We warmly thank Aija Kortesmaa, Hanna Kin- nunen, Ari Klemola, Minna Nykter and Eero Lamminen for expertise, assistance and fruitful discussions during the study. Finnish linseed companies Elixi Oil Oy, Oy Linseed Protein Finland Ltd, Neomed Oy and Sini-Pellava Oy are acknowledged for cooperation and for providing the sam- ples. References Angelova, V., Ivanova, R., Delibaltova, V. & Ivanov, K. 2003. Bio-accumulation and distribution of heavy metals in fibre crops (flax, cotton and hemp). Industrial Crops and Products 19: 197–205. Chakravarty, B. & Srivastava, S. 1997. Effect of cadmium and zinc interaction on metal uptake and regeneration of tolerant plants in linseed. Agriculture, Ecosystems and Environment 61: 45–50. 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Diet monitoring for assessment of human exposure to environmental pollutants. Environ- ment International 28: 703–709. Tahvonen, R. 1995. Contents of lead and cadmium in foods in Finland. University of Turku, Department of Biochem- istry and Food Chemistry, Turku, Finland. AcademicAcademic dissertation. 115 p. Tarpila, S., Tarpila, A., Gröhn, P., Silvennoinen, T. & Lind- berg, L. 2004. Efficacy of ground flaxseed on constipa- tion in patients with irritable bowel syndrome. Current Topics in Nutraceutical Research 2: 119–125. SELOSTUS Suomalaisen öljypellavan kadmiumpitoisuus ja arvioitu saanti kuluttajille Hanna-Riitta Kymäläinen ja Anna-Maija Sjöberg Helsingin yliopisto Tutkimuksen tavoitteena oli selvittää suomalaisen öljy- pellavan siemenen ja rouheen kadmiumpitoisuus ja ar- vioida hyväksyttävää päiväsaantia. Siemennäytteet (N = 85) kerättiin Helsingin yliopiston koetilalta sekä kaupal- lisilta viljelmiltä Etelä-Suomesta ja Pohjanmaalta. Li- säksi tutkittiin 15 pellavarouhenäytettä. Öljypellava- näytteiden kadmiumpitoisuus määritettiin atomiabsorp- tiospektrometrisesti, ja se vaihteli 0,27 ja 1,3 mg/kg vä- lillä (keskiarvo 0,61 mg/kg); pitoisuudet on ilmoitettu kuivamassaa kohti. Rouhenäytteiden Cd-pitoisuudet oli- vat 0,47–1,5 mg/kg (keskiarvo 0,85 mg/kg). Koetilalla viljeltyjen kolmen öljypellavalajikkeen kadmiumpitoi- suus vaihteli lajikkeittain ja myös eri vuosina. Typpilan- noituksen lisäys lisäsi pellavan maasta ottamaa Cd-mää- rää jonkin verran. Sallittu kadmiumin saanti riippuu ke- hon painosta ja tuotteen Cd-pitoisuudesta, ja se on useimmissa tapauksissa suurempi kuin suositeltu syönti- määrä (24–30 g päivittäinen annos öljypellavaa). Cadmium content of Finnish linseed and estimated consumer intake Introduction Material and methods Results Discussion Conclusion References SELOSTUS