The effect of shading on nitrate metabolism in stalks and blades of celery leaves (Apium graveolens L. var. dulce)

The results of two year studies (2003 − 2004) were presented, concerning the effect of various manners of ‘Tango’ celery shading on NR activity and the contents of NO3 and NH4 ions, free amino acids, soluble sugars and dry matter, all of which were determined in the stalks and blades of leaves on three different dates, including the harvest time. Three different shading types were used: shading of leaf stalks only with black film fixed on leaf blade level, whole plant shading and shading by double density planting. Film shading started 16 days before the harvest. In comparison with the control plants, shading, particularly of whole plants, inhibited NR activity in leaf stalks and blades. Nitrate content in celery stalks decreased in the final two weeks of vegetation regardless of the method of Renata Wojciechowska, Piotr Siwek 26 cultivation. At harvest time, least nitrates were found in leaf stalks of double densed plants. NO3 level was significantly higher in the blades of leaves wholly shaded with black film, in which the lowest content of NH4 ions was also found. The significant increase of free amino acids in leaf stalks during vegetation occurred regardless of the method of cultivation, with highest amount of this compound presented in plants with shaded stalks as well as wholly shaded plants. Highest soluble sugar content was found in stalks and blades of the celeries cultivated in double density. Dry matter content in both parts of celery leaves was highest in double density treatment and in the control plants. INTRODUCTION Modification of environmental conditions which accompanies horticultural plants cultivation and its effect on the size and quality of the yield is a common knowledge. For example, direct covering of plants and soil mulching significantly improve the growth and yield of several varieties of Chinese cabbage (Kalisz and Cebula 2001). Studies conducted by Siwek (2002) showed that shading can effectively improve the commercial value of some vegetables in respect to their appearance and taste. In the cited study, the author employed an innovative manner of blanching leaf celery stalks using black film, which resulted in obtaining lighter and more delicate stalks, more valued on the market than the unbleached ones. However, reducing access to light can modify many physiological processes in growing plants, in particular photosynthesis. For instance, celery plants shading was found to have a significant effect on assimilation pigment content in stalks (Siwek et al. 2006b). However, available literature on the subject lacks data on nitrate metabolism in celery stalks depending on the applied shading method. Some works indicate that celery plants may accumulate considerable amounts of nitrates in overground parts (Martignon et al. 1994). The main factors which affect the degree of NO3 bioaccumulation in plants, apart from nitrogen fertilization, include the weather conditions which accompany field cultivation of vegetables, such as light, rainfall and temperature (Rożek 2000). Nitrate ion content in plants is closely connected with the rate of their reduction. Nitrate reductase (NR) is responsible for the first stage of this process. Although the induction of this enzyme occurs solely in the presence of the substrate, in this case NO3 ions, maximal induction, apart from nitrates, requires light (Lillo et al. 2004). Low light intensity, which is connected with low photosynthesis intensity, has an effect on nitrate content growth in plants, the fact that was proved to occur, for instance, in spinach leaves (Kaiser and Brendle-Benisch 1991). In the present study, the effect of various methods of limiting light access to stalks of the ‘Tango’ celery on NR activity as well as the contents of NO3 and Nitrate metabolism in celery leaves 27 NH4 ions, free amino acids, soluble sugars and dry matter was determined, separately for leaf stalks and blades, during 16-day vegetation until the harvest date. MATERIAL AND METHODS The study was carried out in 2003 and 2004 at the Experimental Station of Agricultural University in Kraków and concerned Tango celery cultivar. Seedlings were prepared in greenhouse conditions from seeds sown to containers on 21.01.2003 and 6.02.2004. At the 4-leaf phase, on 23.04 and 21.04 respectively, seedlings were placed directly in field soil in 30 × 25 cm spacing. Based on the chemical analysis of the soil and on the nutritional needs of celery, the following mineral fertilization was determined: 70 kg N ha (N-NO3), 100 kg P2O5 ha, and 200 kg K2O ha. Half of the nitrogen dose was introduced before seedling placement into the soil, and the other half as a top-dressing. The experiment was carried out in a randomized block design in four replications. Each treatment of the experiment included 112 plants. Treatments were made by various manners of shading: 1. without shading (control plants), 2. stalk shading with PP 50 g m black film (stalks were shaded by fixing the film which was earlier used as mulch, at the level of leaf blades), 3. whole plant shading with PP 50 g m film (on 1 m high tunnel construction), 4. double plant density i.e. sowing plants in 15 × 25 cm spacing. The properties of applied films were presented by Siwek (2002). Shading started on 17.06.2003 and 18.06.2004 and was continued until the harvest time which took place on 3 and 5 July respectively. The evaluation of the studied indices in celery stalks and blades was carried out in three different times, namely: I – 19 June 2003 and 21 June 2004; II – 26 and 28 June respectively and III (harvest time) – 3 and 5 July respectively. Nitrate reductase activity (abbr. NR) was determined in accordance with the method described by Jaworski with the modifications of Rożek (1982). NO3 and NH4 ion content in plant material was determined using ORION ion-selective electrodes working together with UNICAM-9460 ionometer (samples were 0.02M Al2(SO4)3 18H2O extracted). Free amino acid content was determined in accordance with the procedure described by Korenman (1973). Soluble sugar content was determined by the photometric method with antrone reagent, described by Yemm and Wills (1954). In order to determine dry matter content, samples of plant material were dried at 105°C. All the investigations were statistically evaluated using Duncan’s test at significance level p = 0.05. Renata Wojciechowska, Piotr Siwek 28 RESULTS AND DISCUSSION The results obtained show that regardless of the applied method of cultivation (Fig. 1) NR activity in stalks was many times lower than in blades. The highest activity of this enzyme was found in the blades of the control plants in the second investigation time, when a significant increase of reductase activity in the blades of leaves harvested from all combinations occurred, except plants which were wholly shaded for the period of 16 days until the harvest date. The latter manifested a drastic nitrate reductase decrease, lasting until the harvest time. Same dependence was shown for leaf stalks of the same plants. The present results confirm the ones obtained by other authors who claim that light is the main external factor which modifies NR activity in leaves on post-translation level, as in result of rapid plant shading the activity of this enzyme quickly decreases (Huber et al. 1992, Lillo 1994). The results obtained by Siwek et al. (2006b) show that in comparison with the control plants shading with each type of film resulted in the decrease of NR activity in celery stalks, with black film having the most inhibitive effect. However, the effect of the used film on nitrate content in celery stalks was not explicitly shown in their work.