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Investigation of sampling and Sources of Variation in Nitrate Levels in Lettuce and Spinach

Project Code: C03030

31/03/2004

ADAS Consulting Ltd, Groesfaen
Weightman, R ; Buxton, J; Dyer, C;
Direct Laboratories Ltd
Farrington, D

This study was commissioned by the Food Standards Agency in order to investigate variation in tissue nitrate concentration (TNC) in spinach and lettuce, grown on farm. In four separate experimental series, on 14 different commercial sites and one experimental site, the response of TNC to (i) short-term fluctuation in light prior to harvest, (ii) varietal variation, and variability in soil mineral nitrogen, (iii) time of day harvested and (iv) within crop variation, were studied.

On the basis of these results, a second objective was to advise the FSA on best practice, with regards to sampling for nitrate in lettuce or spinach (in the field or glasshouse), and to comment of the suitability and robustness of the current sampling Directive (2002/36/EC) when applied to sampling of lettuce and spinach for nitrate analysis.

Prior to experimental work, an initial literature review was carried out to establish the basis of current sampling protocols. It was found that there was no scientific basis for the choice of 10 plants (or 1 kg by weight) to be harvested. Current field sampling recommendations for soil are based on work carried out in the 1940's and 1950's and its relevance to current field sampling protocols is discussed.

Results of the field experiments are summarised as follows:

The effects on TNC of short term fluctuations in light level (e.g. shading for 24-48 h prior to harvest) in the present study were relatively small, compared to the large differences known to occur following dull periods in the 10-14 days prior to harvest, or for example between winter and summer grown crops;
Variety accounted for 16-37% of the variance in TNC within a species, and is a significant factor which needs to be taken into account when drafting sampling protocols;
Time of day at which plants were harvested accounted for 20-34% of the variability in TNC measured within the day, although there was no evidence for a consistent pattern of diurnal variation in TNC, which could be used to advise harvesting strategy in order to minimise nitrate;
Estimates of mean TNC at two commercial lettuce sites (60 plants sampled at each), based on groups of 10 plants selected at random, using either W or X shaped sampling patterns, were not significantly different from the grand mean, suggesting that these sampling patterns were acceptable. In contrast, significant differences in mean TNC were obtained between different groups of ten lettuces sampled contiguously along the row;
A sample size of 10 appears to be an acceptable for lettuce in order to obtain an unbiased estimate of the mean, assuming an appropriate sampling plan is used (equivalent data not collected for spinach);
When data were pooled from all experiments, coefficients of variation ranged from 14-63% for lettuce and 29-92% for spinach. When means and standard deviations for all datasets were pooled, an average CV of 37% was estimated;
Doubling the sample size from 10 to 20 plants would reduce the standard error from 16 to 13% of the mean in the case of lettuce, and from 22 to 17% for spinach. Further increasing sample size to 40 plants would reduce standard errors to 12 and 14% of the mean for lettuce and spinach respectively. Further increases would have little additional impact because of the underlying contribution from analytical variability, which would remain constant regardless of sample size.

On the basis of the literature review and the results from field studies, comments are presented on the current sampling directive (2002/63/EC), and improved guidance to samplers is discussed. The findings of this project will inform the design of improved protocols for field sampling for nitrate in lettuce and spinach.

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