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Content of vitamin C, carotenoids, chlorophylls and polyphenols in green parts of dill (Anethum graveolens L.) depending on plant height
ARTICLE IN PRESS JOURNAL OF FOOD COMPOSITION AND ANALYSIS Journal of Food Composition and Analysis 19 (2006) 134–140 www.elsevier.com/locate/jfca
Original Article
Content of vitamin C, carotenoids, chlorophylls and polyphenols in green parts of dill (Anethum graveolens L.) depending on plant height Zofia Lisiewska, Waldemar Kmiecik, Anna Korus Department of Raw Materials and Processing of Fruit and Vegetables, Agricultural University, 122 Balicka Street, 30-149 Krakow, Poland Received 6 October 2004; received in revised form 14 April 2005; accepted 16 April 2005
Abstract The aim of the present investigation was to determine the level of vitamin C, carotenoids, beta-carotene, chlorophylls and polyphenols in dill plants 20, 30, 40, 50 and 60 cm in height. Analyses included the leaf blades, petioles, whole leaves (leaf blades with petioles), stems and whole plants (whole leaf with the stem). In all parts of dill plants, with the exception of petioles, the content of vitamin C decreased with plant growth. The content of carotenoids and beta-carotene tended to increase with growth, with the exception of stems and whole plants. The proportion of beta-carotene in the content of carotenoids varied within the range of 9–17%, the highest being noted in leaf blades. The taller the plants, the higher the level of chlorophylls and polyphenols in the analysed parts of dill, except in whole plants and—in the case of polyphenols—also in stems. The taller the plants, the lower the ratio of chlorophyll a to b. Of all the constituents analysed, the highest content was found in the leaf blade and the lowest in the stem and the petiole. r 2005 Elsevier Inc. All rights reserved. Keywords: Dill; Plant height; Vitamin C; Carotenoids; Beta-carotene; Chlorophylls; Polyphenols
1. Introduction In recent years the demand for seasoning vegetables has grown, reflecting the increase in their consumption (Sloan, 2004). The variety of dill classed as a seasoning vegetable is being applied more and more widely in modelling the flavour of numerous food products. It can be used as an ingredient in dried seasoning mixtures; in the production of cheeses, fish, and vegetarian dishes; as an admixture in ‘‘ready-to-eat’’ and—more recently— ‘‘do-it-for-me’’ dishes (Pszczola, 2001; Sloan, 2004); and also as a basic constituent of soups and sauces which are very popular in Central and Eastern Europe. The use of dill for various culinary purposes depends to a great extent on its vegetative development as measured by the plant height. The value of dill for processing also Corresponding author. Tel.: +48 12 662 47 56; fax: +48 12 662 47 57. E-mail address: [email protected] (Z. Lisiewska).
0889-1575/$ - see front matter r 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.jfca.2005.04.009
depends on the stage of growth. Young delicate plants can be used for drying, the older ones for freezing, while those at a more advanced stage of growth can be used for the preparation of stock or extracts. The intensity of colour and its shade—and hence the attractiveness of the raw material—depend on the level of chlorophyll pigments and their proportions (Lisiewska et al., 2001). In green plants the chlorophyll pigments are accompanied by carotenoids, which affect the colour of the raw material and of products obtained from it and also enhance their vitamin content. The last statement chiefly concerns beta-carotene (Bhaskarachary et al., 1995). Like vitamin C and polyphenol compounds, carotenoids are also classified among the basic constituents of the antioxidative effect (Duthie et al., 2003; Kidmose et al., 2001). Green vegetables, including dill, are a rich source of these substances (Agte et al., 2000). The aim of the present study is to determine the level of compounds on which the colour of dill depends and
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the level of compounds with an antioxidative effect. The objects were dill plants measuring 20, 30, 40, 50 and 60 cm in height; their separate parts, i.e. leaf blades, petioles, whole leaves (leaf blade with petioles), stems and whole plants (all the leaves with the stem) were analysed.
2. Materials and methods 2.1. Condition of plant growth The material investigated was the various usable parts of dill plants of the cultivar Amat, harvested at the height of 20, 30, 40, 50 and 60 cm. Dill was cultivated in the experimental field of the Department of Raw Materials and Processing of Fruit and Vegetables, situated in the western outskirts of Krakow, on brown soil developed from loess formations with the mechanical composition of silt loam. The soil, of good horticultural quality, was characterized by an almost neutral pH value of 6.5 in H2O; a moderate content of humus (0.93%); and a high content of phosphorus (70 mg/dm3), potassium (175 mg/dm3) and calcium (1460 mg/dm3). Dill was grown in the third year after manure fertilization with cucumbers as the preceding crop. Taking into account the soil fertility and the nutritional requirements of the species, the following doses of mineral fertilizers were applied: nitrogen before sowing 30 kg N/ha in the form of ammonium nitrate; phosphorus before sowing 15 kg P2O5/ha in the form of triple super phosphate; and potassium before sowing 30 kg K2O/ha in the form of 60% potassium chloride. 2.2. Harvest of the dill Dill was harvested when plants attained the height determined by the accepted method. This occurred after 50, 54, 59, 63, and 69 days of sowing, respectively. The number of leaves per plant increased from range 3–5 at a height of 20 cm to 6–9 at a height of 60 cm. The experiment was ended when umbels began to appear on the plants. The selection of plant heights depended on the use of dill, i.e. at a height of 20 and 30 cm whole plants could be used especially for soups and sauces; at a height of about 40 and 50 cm whole leaves with petioles can be used for these purposes. However, from plants 60 cm in height, which are bolting in this stage of development, only the leafy part or the brew from whole plants can be used in practice. The harvest was carried out by cutting plant tops 1.5–2.0 cm above the soil. Directly after harvesting the plants were inspected; all those with yellowing leaves or single yellow leaves were discarded. The objects of the study were usable parts of dill: the leaf blade, petiole,
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whole leaf (the blade with petiole), the stem, and whole plants (the whole leaf with the stem). 2.3. Chemical analysis Analyses were started about 1 h after harvest. An average evaluated sample was about 1000 g in weight and sampled so as to be representative of the given investigated material. Chemical analyses were carried out in four replications, each in two parallel samples. The content of vitamin C was determined using the spectrophotometrical method (ISO/6557-2, 1984). Oxalic acid solution (2%) was used for extraction of the ascorbic acid, after quantitative reduction of 2,6dichlorophenolindophenol dyestuff by ascorbic acid and extraction of the excess dyestuff using xylene. The excess was measured at 500 nm in a Shimadzu UV 160A spectrophotometer and compared with a vitamin C reference standard. Beta-carotene analysis consisted of the extraction procedures of pigment, followed by liquid/liquid partitioning with hexane, concentration and column chromatography (ISO/6558-2, 1992). The same extracts obtained for carotenoid estimation were used. Hexane extracts was filtered over anhydrous sodium sulphate on filter paper (Whatman No. 1 equivalent) and was made up to a known volume. The extract was concentrated 10fold by evaporation and loaded onto the column. Columns (150 10 mm) were packed with aluminium oxide to a length of 100 mm and covered with a 10 mm of anhydrous sodium sulphite, then were washed with hexane containing 1% acetone. The orange-coloured eluent containing beta-carotene was collected to a volumetric flask. The concentration of beta-carotene was measured at 450 nm in a Shimadzu UV 160A spectrophotometer and compared with a beta-carotene reference standard. Total carotenoids and chlorophylls were determined according to the methods of Wettstein (1957). The extraction procedures of pigment were carried out under dim light and in a glassware wrapped with aluminium foil. The method consisted of acetone extraction repeated until the obtention of a colourless residue with pestle and mortar and filtered over a cotton pad. The extracts were made up to 50 mL with acetone. The concentration of carotenoids was measured at 440.5 nm, chlorophyll a at 662 nm, chlorophyll b at 644 nm in a Shimadzu UV 160A double-beam spectrophotometer. Total phenolic compounds were determined using the Folin–Ciocalteau reagent according to Swain and Hillis (1959) procedure. Two grams of homogenized samples were boiled 20 min in a 80% ethanol under reflux. Five millilitres of 10-fold diluted extract and 0.5 mL two-fold diluted Folin–Ciocalteau reagent were added along with 1 mL of 25% sodium carbonate, and the contents were mixed thoroughly. The absorbance was measured at
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675 nm in a Shimadzu spectrophotometer after 60 min using catechin as a standard. 2.4. Statistical analysis To establish possible differences in the level of analysed indices of chemical composition between the parts of dill plants and also of differences resulting from the size of plants, an analysis of variance was carried out according to the Excel 5.0 program using the SnedecorF and Student-t-tests, while the least-significant difference (LSD) was calculated for the level of error probability of P ¼ 0:01 and 0:05 (ANOVA).
3. Results and discussion As Ba˜no et al. (2003) postulate, the distribution of constituents changes during plant growth and their concentration varies in different parts of the plant. Furthermore, these inter-relationships differ according to the plant species (Najda et al., 2003; Vallejo et al., 2003; Yamada et al., 2003). 3.1. Content of vitamin C Depending on the usable part, the recorded content of vitamin C varied within the range of 29 mg in 100 g dill stems 60 cm in height to 186 mg in 100 g leaves of dill plants 20 cm in height (Table 1). Oguchi et al. (1996) and G˛ebczyn´ski (1998, 1999) found that the leaves of spinach and of leaf beet contained more vitamin C than the petioles. In whole dill the smaller—i.e. the younger—the plants, the greater the content of vitamin C. A similar tendency was also observed in the analysed parts of dill, with the exception of petioles, where the level of vitamin C was uniform. In leaves of broccoli and spinach the concentration of vitamin C increased with the maturation of plants (Botero Omary et al., 2003; Yamada et al., 2003); however it decreased in leaves of iceberg and butterhead lettuce (Drews et al., 1995,
1997). In comparison with a whole plant of appropriate height where the content of vitamin C was assumed to be 100%, the leaf blades of the smallest plants contained 160% vitamin C, and the largest 295%, the petiole 33% and 67%, respectively, the whole leaf 119% and 216%, and the stem 34% and 53%. According to Ishida et al. (2000), the petioles and stems of sweet potatoes, respectively, contained 14–21% and 21–24% of the quantity of vitamin C found in leaves. 3.2. Content of carotenoids The direction of changes in the level of carotenoids depending on the plant height was not uniform in the analysed parts of dill plants (Table 2). In leaf blades and in whole leaves the content of carotenoids gradually increased with the growth of the plant, decreasing in whole plants at the same time. In petioles and stems the content decreased up to 40 cm of plant height, then increased again. Saric et al. (1990) found the greatest content of carotenoids in the oldest leaves of cabbage. Of the analysed parts of dill plants the greatest content of carotenoids was noted in leaf blades (Table 2). Jaworska et al. (2001) reported that in leaves of dill the content of carotenoids was 181% of that found in whole plants. Ishida et al. (2000) also recorded a greater content of carotenoids in leaves of two cultivars of sweet potato than in petioles and stems. If it is assumed that the content of these compounds in whole dill plants, for each height separately, was 100%, then the leaf blades contained 164–379%, the petiole 38–74%, whole leaf 123–275% and the stem 16–28%. In each case the percentages increased with the height (i.e. the age) of the plant. 3.3. Content of beta-carotene Of the analysed parts of dill the greatest content of beta-carotene was found in leaf blades, 4.48–5.62 mg of this compound in 100 g fresh matter, and in the whole leaf, 2.79–3.95 mg (Table 3). A distinctly smaller content
Table 1 Content of vitamin C in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0:01 LSD Po0:05 a
Mean (n ¼ 4).
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
18677 17375 17075 15975 16276 11.9 8.6
3873 3572 3972 3372 3772 4.4 3.2
13875 12274 12274 11673 11975 9.0 6.5
3972 3072 3372 2972 2971 3.8 2.8
11674 8973 7172 5872 5572 5.6 4.0
9.5 7.2 6.9 6.3 7.7
6.8 5.2 5.0 4.6 5.6
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Table 2 Content of carotenoids in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0:01 LSD Po0:05 a
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
27.871.0 29.171.1 28.971.0 30.672.0 34.971.2 2.75 1.99
6.470.2 6.170.3 6.070.3 6.270.3 6.870.3 n.s. 0.44
20.870.7 20.570.8 20.570.7 22.271.2 25.370.9 1.84 1.33
3.270.2 2.270.3 2.070.2 2.170.2 2.670.3 0.47 0.34
16.970.6 13.970.4 10.070.2 8.870.4 9.270.4 0.92 0.66
1.34 1.38 1.22 2.24 1.47
0.97 1.00 0.88 1.62 1.06
Mean (n ¼ 4).
Table 3 Content of b-carotene in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0.01 LSD Po0.05 a
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
4.4870.23 4.0770.22 4.0970.20 5.2270.18 5.6270.17 0.420 0.304
0.6670.03 0.6470.04 0.6570.03 0.7270.04 0.7370.04 0.075 0.054
3.2470.16 2.7970.14 2.8270.13 3.6770.11 3.9570.11 0.277 0.201
0.2470.02 0.2170.02 0.2070.01 0.2270.02 0.2570.02 n.s. 0.030
2.5770.13 1.8670.09 1.3470.05 1.3770.04 1.3270.05 0.164 0.118
0.291 0.261 0.230 0.202 0.201
0.211 0.189 0.167 0.146 0.145
Mean (n ¼ 4).
was found in the petiole, 0.64–0.73 mg, and especially in the stem, 0.20–0.25 mg in 100 g fresh matter. The dependence of beta-carotene content on plant height was similar to that given in the discussion of total carotenoids, since the proportion of this compound in total carotenoids did not vary. Depending on plant height, the content was 14–17% in leaf blades and whole plants, 10–12% in the petiole, 9–10% in the stem and 13–16% in whole plants. Contrary to the present results, Drews et al. (1995, 1997) observed a decreasing content of beta-carotene with the maturation of butterhead and iceberg lettuce. 3.4. Content of chlorophylls In the individual parts of larger plants a higher chlorophyll content was observed than in smaller plants, although the differences were statistically non-significant in many cases (Table 4). In whole plants the opposite tendency was observed. If the level of chlorophyll was 100% in the different parts of the youngest plants, the respective values for the oldest dill were: 153% in leaf blades, 137% in petioles, 149% in whole plants, and distinctly less—115%—in the stems. With the increasing height of dill plants, the proportions
between the above parts changed, that of the stem particularly increasing. Moreover, on the assumption that the whole plant contained 100% of chlorophyll, the stem contained only 15–26% of this amount, hence the content of chlorophylls in the whole plant consistently decreased as the plant grew. In comparison with the whole plant, the remaining parts of dill contained chlorophyll as follows: the leaf blade 166–387%; the petiole 37–78%; and the whole leaf 124–281%. The older the plants, the higher the percentage values. As in earlier studies, the authors also found considerable differences in the concentration of chlorophyll between leaves and whole dill plants (Kmiecik et al., 2001; Lisiewska et al., 2001). As in the present experiment, Saric et al. (1990) also reported a greater content of chlorophyll in older leaves of cabbage than in younger ones. As the height of the plants increased the ratio of chlorophyll a to chlorophyll b consistently decreased (Table 4). Levels were 4.48–2.86 for the leaf blade; 2.89–1.90 for the petiole; 4.27–2.74 for the whole leaf; 3.25–1.65 for the stem; and 4.21–2.47 for the whole plant, showing a distinctly faster increase in the content of chlorophyll b in comparison with chlorophyll a as the plant grew.
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Table 4 Content of chlorophylls in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
Chlorophyll (a+b) 20 30 40 50 60 LSD Po0:01 LSD Po0:05
95.372.8 125.175.2 140.675.2 141.073.5 146.276.1 9.79 7.08
21.471.2 21.870.9 22.070.8 25.470.6 29.375.8 5.68 4.11
71.372.1 86.673.2 96.973.2 101.272.5 106.374.4 6.61 4.78
8.570.5 8.670.3 8.970.4 9.170.4 9.870.5 0.87 0.63
57.371.8 58.672.1 47.171.7 39.970.8 37.871.5 3.35 2.42
3.83 6.07 5.95 4.09 8.96
2.77 4.39 4.31 2.96 6.48
Chlorophyll a 20 30 40 50 60 LSD Po0:01 LSD Po0:05
77.972.3 95.172.7 105.273.6 104.872.4 108.373.9 6.14 4.61
15.970.8 15.370.6 15.170.5 16.870.3 19.270.8 4.28 0.97
57.771.7 65.471.7 72.072.2 74.571.7 77.972.8 5.15 3.09
6.570.4 5.970.1 6.070.3 5.970.3 6.170.3 n.s. n.s.
46.371.4 44.071.1 34.671.2 28.870.5 26.971.0 4.45 1.60
4.70 4.74 5.09 4.62 5.30
2.22 2.31 2.99 2.03 3.35
Chlorophyll b 20 30 40 50 60 LSD Po0:01 LSD Po0:05
17.470.7 30.072.5 35.471.6 36.271.2 37.972.3 3.73 2.70
5.570.4 6.570.3 6.970.4 8.670.3 10.170.6 0.81 0.58
13.570.6 21.271.5 24.971.1 26.770.8 28.471.7 2.50 1.81
2.070.1 2.770.2 2.970.1 3.270.1 3.770.2 0.31 0.23
11.070.5 14.671.0 12.470.5 11.070.3 10.970.5 1.27 0.92
0.99 2.91 1.86 1.38 2.78
0.71 2.11 1.35 1.00 2.01
a
Mean (n ¼ 4).
Table 5 Content of polyphenols in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0:01 LSD Po0:05 a
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
19778 17378 214710 278711 331712 20.6 14.9
5573 5473 9277 8574 8774 9.5 6.9
15176 12976 16975 21278 24879 14.7 10.7
5373 4974 6674 5473 4973 6.3 4.6
12975 10073 11174 10773 10774 8.3 6.0
11.2 10.8 13.3 13.7 15.3
8.1 7.8 9.6 9.9 11.0
Mean (n ¼ 4).
3.5. Content of polyphenols With the increasing height of the plant, the content of polyphenols increased, with some exceptions, in leaf blades, petioles and whole leaves, while in general it showed a decreasing tendency in the stem and the whole plant (Table 5). Moreover, in the analysed parts of the plant differences in the content of these compounds according to height were not always significant. In
comparison with parts of the youngest plants where the level of polyphenols was assumed to be 100%, in the oldest plants the leaf blade contained 168%, the petiole 158%, whole leaf 164%, the stem 92%, and the whole plant 83%. Choi and Lee (1999) also recorded fewer polyphenols in younger leaves of Agastacha rugosa. However, according to Ba˜no et al. (2003) the greatest accumulation of polyphenol compounds occurred in early developmental stages of Rosmarinus officinalis.
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Contrary to the present study, Vallejo et al. (2003) wrote that in broccoli plants the content of polyphenol compounds showed a tendency to increase from the first stage to the last. If it was assumed that, separately for each height, the content of polyphenols in the whole plant was 100%, the leaf blade contained as much as 153–309% of these compounds, the petiole 43–93%, whole leaf 117–232%, and the stem 41–59%. With the exception of the petiole from a plant 60 cm in height and of the stem from plants 50 and 60 cm in height, steady growth occurred in the content of these compounds. Romani et al. (2003) also stress a high content of polyphenols in soy leaves compared with stems. However, Najda et al. (2003) demonstrated a smaller differentiation in the content of polyphenol compounds in lovage plants where the leaf blade contained 119% and the stem 38% compared with the whole plant.
4. Conclusions The highest content of all the analysed constituents was found in the leaf blade, and the lowest in the stem and the petiole. The content of vitamin C decreased with plant growth, with the exeption of petiols; the content of carotenoids and beta-carotene tended to increase with growth, with the exception of stems and whole plants. The higher the plants, the higher the level of chlorophylls and polyphenols in the analysed parts of dill, except in whole plants and, in the case of polyphenols, also in stems.
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Original Article
Content of vitamin C, carotenoids, chlorophylls and polyphenols in green parts of dill (Anethum graveolens L.) depending on plant height Zofia Lisiewska, Waldemar Kmiecik, Anna Korus Department of Raw Materials and Processing of Fruit and Vegetables, Agricultural University, 122 Balicka Street, 30-149 Krakow, Poland Received 6 October 2004; received in revised form 14 April 2005; accepted 16 April 2005
Abstract The aim of the present investigation was to determine the level of vitamin C, carotenoids, beta-carotene, chlorophylls and polyphenols in dill plants 20, 30, 40, 50 and 60 cm in height. Analyses included the leaf blades, petioles, whole leaves (leaf blades with petioles), stems and whole plants (whole leaf with the stem). In all parts of dill plants, with the exception of petioles, the content of vitamin C decreased with plant growth. The content of carotenoids and beta-carotene tended to increase with growth, with the exception of stems and whole plants. The proportion of beta-carotene in the content of carotenoids varied within the range of 9–17%, the highest being noted in leaf blades. The taller the plants, the higher the level of chlorophylls and polyphenols in the analysed parts of dill, except in whole plants and—in the case of polyphenols—also in stems. The taller the plants, the lower the ratio of chlorophyll a to b. Of all the constituents analysed, the highest content was found in the leaf blade and the lowest in the stem and the petiole. r 2005 Elsevier Inc. All rights reserved. Keywords: Dill; Plant height; Vitamin C; Carotenoids; Beta-carotene; Chlorophylls; Polyphenols
1. Introduction In recent years the demand for seasoning vegetables has grown, reflecting the increase in their consumption (Sloan, 2004). The variety of dill classed as a seasoning vegetable is being applied more and more widely in modelling the flavour of numerous food products. It can be used as an ingredient in dried seasoning mixtures; in the production of cheeses, fish, and vegetarian dishes; as an admixture in ‘‘ready-to-eat’’ and—more recently— ‘‘do-it-for-me’’ dishes (Pszczola, 2001; Sloan, 2004); and also as a basic constituent of soups and sauces which are very popular in Central and Eastern Europe. The use of dill for various culinary purposes depends to a great extent on its vegetative development as measured by the plant height. The value of dill for processing also Corresponding author. Tel.: +48 12 662 47 56; fax: +48 12 662 47 57. E-mail address: [email protected] (Z. Lisiewska).
0889-1575/$ - see front matter r 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.jfca.2005.04.009
depends on the stage of growth. Young delicate plants can be used for drying, the older ones for freezing, while those at a more advanced stage of growth can be used for the preparation of stock or extracts. The intensity of colour and its shade—and hence the attractiveness of the raw material—depend on the level of chlorophyll pigments and their proportions (Lisiewska et al., 2001). In green plants the chlorophyll pigments are accompanied by carotenoids, which affect the colour of the raw material and of products obtained from it and also enhance their vitamin content. The last statement chiefly concerns beta-carotene (Bhaskarachary et al., 1995). Like vitamin C and polyphenol compounds, carotenoids are also classified among the basic constituents of the antioxidative effect (Duthie et al., 2003; Kidmose et al., 2001). Green vegetables, including dill, are a rich source of these substances (Agte et al., 2000). The aim of the present study is to determine the level of compounds on which the colour of dill depends and
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the level of compounds with an antioxidative effect. The objects were dill plants measuring 20, 30, 40, 50 and 60 cm in height; their separate parts, i.e. leaf blades, petioles, whole leaves (leaf blade with petioles), stems and whole plants (all the leaves with the stem) were analysed.
2. Materials and methods 2.1. Condition of plant growth The material investigated was the various usable parts of dill plants of the cultivar Amat, harvested at the height of 20, 30, 40, 50 and 60 cm. Dill was cultivated in the experimental field of the Department of Raw Materials and Processing of Fruit and Vegetables, situated in the western outskirts of Krakow, on brown soil developed from loess formations with the mechanical composition of silt loam. The soil, of good horticultural quality, was characterized by an almost neutral pH value of 6.5 in H2O; a moderate content of humus (0.93%); and a high content of phosphorus (70 mg/dm3), potassium (175 mg/dm3) and calcium (1460 mg/dm3). Dill was grown in the third year after manure fertilization with cucumbers as the preceding crop. Taking into account the soil fertility and the nutritional requirements of the species, the following doses of mineral fertilizers were applied: nitrogen before sowing 30 kg N/ha in the form of ammonium nitrate; phosphorus before sowing 15 kg P2O5/ha in the form of triple super phosphate; and potassium before sowing 30 kg K2O/ha in the form of 60% potassium chloride. 2.2. Harvest of the dill Dill was harvested when plants attained the height determined by the accepted method. This occurred after 50, 54, 59, 63, and 69 days of sowing, respectively. The number of leaves per plant increased from range 3–5 at a height of 20 cm to 6–9 at a height of 60 cm. The experiment was ended when umbels began to appear on the plants. The selection of plant heights depended on the use of dill, i.e. at a height of 20 and 30 cm whole plants could be used especially for soups and sauces; at a height of about 40 and 50 cm whole leaves with petioles can be used for these purposes. However, from plants 60 cm in height, which are bolting in this stage of development, only the leafy part or the brew from whole plants can be used in practice. The harvest was carried out by cutting plant tops 1.5–2.0 cm above the soil. Directly after harvesting the plants were inspected; all those with yellowing leaves or single yellow leaves were discarded. The objects of the study were usable parts of dill: the leaf blade, petiole,
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whole leaf (the blade with petiole), the stem, and whole plants (the whole leaf with the stem). 2.3. Chemical analysis Analyses were started about 1 h after harvest. An average evaluated sample was about 1000 g in weight and sampled so as to be representative of the given investigated material. Chemical analyses were carried out in four replications, each in two parallel samples. The content of vitamin C was determined using the spectrophotometrical method (ISO/6557-2, 1984). Oxalic acid solution (2%) was used for extraction of the ascorbic acid, after quantitative reduction of 2,6dichlorophenolindophenol dyestuff by ascorbic acid and extraction of the excess dyestuff using xylene. The excess was measured at 500 nm in a Shimadzu UV 160A spectrophotometer and compared with a vitamin C reference standard. Beta-carotene analysis consisted of the extraction procedures of pigment, followed by liquid/liquid partitioning with hexane, concentration and column chromatography (ISO/6558-2, 1992). The same extracts obtained for carotenoid estimation were used. Hexane extracts was filtered over anhydrous sodium sulphate on filter paper (Whatman No. 1 equivalent) and was made up to a known volume. The extract was concentrated 10fold by evaporation and loaded onto the column. Columns (150 10 mm) were packed with aluminium oxide to a length of 100 mm and covered with a 10 mm of anhydrous sodium sulphite, then were washed with hexane containing 1% acetone. The orange-coloured eluent containing beta-carotene was collected to a volumetric flask. The concentration of beta-carotene was measured at 450 nm in a Shimadzu UV 160A spectrophotometer and compared with a beta-carotene reference standard. Total carotenoids and chlorophylls were determined according to the methods of Wettstein (1957). The extraction procedures of pigment were carried out under dim light and in a glassware wrapped with aluminium foil. The method consisted of acetone extraction repeated until the obtention of a colourless residue with pestle and mortar and filtered over a cotton pad. The extracts were made up to 50 mL with acetone. The concentration of carotenoids was measured at 440.5 nm, chlorophyll a at 662 nm, chlorophyll b at 644 nm in a Shimadzu UV 160A double-beam spectrophotometer. Total phenolic compounds were determined using the Folin–Ciocalteau reagent according to Swain and Hillis (1959) procedure. Two grams of homogenized samples were boiled 20 min in a 80% ethanol under reflux. Five millilitres of 10-fold diluted extract and 0.5 mL two-fold diluted Folin–Ciocalteau reagent were added along with 1 mL of 25% sodium carbonate, and the contents were mixed thoroughly. The absorbance was measured at
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675 nm in a Shimadzu spectrophotometer after 60 min using catechin as a standard. 2.4. Statistical analysis To establish possible differences in the level of analysed indices of chemical composition between the parts of dill plants and also of differences resulting from the size of plants, an analysis of variance was carried out according to the Excel 5.0 program using the SnedecorF and Student-t-tests, while the least-significant difference (LSD) was calculated for the level of error probability of P ¼ 0:01 and 0:05 (ANOVA).
3. Results and discussion As Ba˜no et al. (2003) postulate, the distribution of constituents changes during plant growth and their concentration varies in different parts of the plant. Furthermore, these inter-relationships differ according to the plant species (Najda et al., 2003; Vallejo et al., 2003; Yamada et al., 2003). 3.1. Content of vitamin C Depending on the usable part, the recorded content of vitamin C varied within the range of 29 mg in 100 g dill stems 60 cm in height to 186 mg in 100 g leaves of dill plants 20 cm in height (Table 1). Oguchi et al. (1996) and G˛ebczyn´ski (1998, 1999) found that the leaves of spinach and of leaf beet contained more vitamin C than the petioles. In whole dill the smaller—i.e. the younger—the plants, the greater the content of vitamin C. A similar tendency was also observed in the analysed parts of dill, with the exception of petioles, where the level of vitamin C was uniform. In leaves of broccoli and spinach the concentration of vitamin C increased with the maturation of plants (Botero Omary et al., 2003; Yamada et al., 2003); however it decreased in leaves of iceberg and butterhead lettuce (Drews et al., 1995,
1997). In comparison with a whole plant of appropriate height where the content of vitamin C was assumed to be 100%, the leaf blades of the smallest plants contained 160% vitamin C, and the largest 295%, the petiole 33% and 67%, respectively, the whole leaf 119% and 216%, and the stem 34% and 53%. According to Ishida et al. (2000), the petioles and stems of sweet potatoes, respectively, contained 14–21% and 21–24% of the quantity of vitamin C found in leaves. 3.2. Content of carotenoids The direction of changes in the level of carotenoids depending on the plant height was not uniform in the analysed parts of dill plants (Table 2). In leaf blades and in whole leaves the content of carotenoids gradually increased with the growth of the plant, decreasing in whole plants at the same time. In petioles and stems the content decreased up to 40 cm of plant height, then increased again. Saric et al. (1990) found the greatest content of carotenoids in the oldest leaves of cabbage. Of the analysed parts of dill plants the greatest content of carotenoids was noted in leaf blades (Table 2). Jaworska et al. (2001) reported that in leaves of dill the content of carotenoids was 181% of that found in whole plants. Ishida et al. (2000) also recorded a greater content of carotenoids in leaves of two cultivars of sweet potato than in petioles and stems. If it is assumed that the content of these compounds in whole dill plants, for each height separately, was 100%, then the leaf blades contained 164–379%, the petiole 38–74%, whole leaf 123–275% and the stem 16–28%. In each case the percentages increased with the height (i.e. the age) of the plant. 3.3. Content of beta-carotene Of the analysed parts of dill the greatest content of beta-carotene was found in leaf blades, 4.48–5.62 mg of this compound in 100 g fresh matter, and in the whole leaf, 2.79–3.95 mg (Table 3). A distinctly smaller content
Table 1 Content of vitamin C in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0:01 LSD Po0:05 a
Mean (n ¼ 4).
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
18677 17375 17075 15975 16276 11.9 8.6
3873 3572 3972 3372 3772 4.4 3.2
13875 12274 12274 11673 11975 9.0 6.5
3972 3072 3372 2972 2971 3.8 2.8
11674 8973 7172 5872 5572 5.6 4.0
9.5 7.2 6.9 6.3 7.7
6.8 5.2 5.0 4.6 5.6
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Table 2 Content of carotenoids in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0:01 LSD Po0:05 a
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
27.871.0 29.171.1 28.971.0 30.672.0 34.971.2 2.75 1.99
6.470.2 6.170.3 6.070.3 6.270.3 6.870.3 n.s. 0.44
20.870.7 20.570.8 20.570.7 22.271.2 25.370.9 1.84 1.33
3.270.2 2.270.3 2.070.2 2.170.2 2.670.3 0.47 0.34
16.970.6 13.970.4 10.070.2 8.870.4 9.270.4 0.92 0.66
1.34 1.38 1.22 2.24 1.47
0.97 1.00 0.88 1.62 1.06
Mean (n ¼ 4).
Table 3 Content of b-carotene in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0.01 LSD Po0.05 a
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
4.4870.23 4.0770.22 4.0970.20 5.2270.18 5.6270.17 0.420 0.304
0.6670.03 0.6470.04 0.6570.03 0.7270.04 0.7370.04 0.075 0.054
3.2470.16 2.7970.14 2.8270.13 3.6770.11 3.9570.11 0.277 0.201
0.2470.02 0.2170.02 0.2070.01 0.2270.02 0.2570.02 n.s. 0.030
2.5770.13 1.8670.09 1.3470.05 1.3770.04 1.3270.05 0.164 0.118
0.291 0.261 0.230 0.202 0.201
0.211 0.189 0.167 0.146 0.145
Mean (n ¼ 4).
was found in the petiole, 0.64–0.73 mg, and especially in the stem, 0.20–0.25 mg in 100 g fresh matter. The dependence of beta-carotene content on plant height was similar to that given in the discussion of total carotenoids, since the proportion of this compound in total carotenoids did not vary. Depending on plant height, the content was 14–17% in leaf blades and whole plants, 10–12% in the petiole, 9–10% in the stem and 13–16% in whole plants. Contrary to the present results, Drews et al. (1995, 1997) observed a decreasing content of beta-carotene with the maturation of butterhead and iceberg lettuce. 3.4. Content of chlorophylls In the individual parts of larger plants a higher chlorophyll content was observed than in smaller plants, although the differences were statistically non-significant in many cases (Table 4). In whole plants the opposite tendency was observed. If the level of chlorophyll was 100% in the different parts of the youngest plants, the respective values for the oldest dill were: 153% in leaf blades, 137% in petioles, 149% in whole plants, and distinctly less—115%—in the stems. With the increasing height of dill plants, the proportions
between the above parts changed, that of the stem particularly increasing. Moreover, on the assumption that the whole plant contained 100% of chlorophyll, the stem contained only 15–26% of this amount, hence the content of chlorophylls in the whole plant consistently decreased as the plant grew. In comparison with the whole plant, the remaining parts of dill contained chlorophyll as follows: the leaf blade 166–387%; the petiole 37–78%; and the whole leaf 124–281%. The older the plants, the higher the percentage values. As in earlier studies, the authors also found considerable differences in the concentration of chlorophyll between leaves and whole dill plants (Kmiecik et al., 2001; Lisiewska et al., 2001). As in the present experiment, Saric et al. (1990) also reported a greater content of chlorophyll in older leaves of cabbage than in younger ones. As the height of the plants increased the ratio of chlorophyll a to chlorophyll b consistently decreased (Table 4). Levels were 4.48–2.86 for the leaf blade; 2.89–1.90 for the petiole; 4.27–2.74 for the whole leaf; 3.25–1.65 for the stem; and 4.21–2.47 for the whole plant, showing a distinctly faster increase in the content of chlorophyll b in comparison with chlorophyll a as the plant grew.
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Table 4 Content of chlorophylls in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
Chlorophyll (a+b) 20 30 40 50 60 LSD Po0:01 LSD Po0:05
95.372.8 125.175.2 140.675.2 141.073.5 146.276.1 9.79 7.08
21.471.2 21.870.9 22.070.8 25.470.6 29.375.8 5.68 4.11
71.372.1 86.673.2 96.973.2 101.272.5 106.374.4 6.61 4.78
8.570.5 8.670.3 8.970.4 9.170.4 9.870.5 0.87 0.63
57.371.8 58.672.1 47.171.7 39.970.8 37.871.5 3.35 2.42
3.83 6.07 5.95 4.09 8.96
2.77 4.39 4.31 2.96 6.48
Chlorophyll a 20 30 40 50 60 LSD Po0:01 LSD Po0:05
77.972.3 95.172.7 105.273.6 104.872.4 108.373.9 6.14 4.61
15.970.8 15.370.6 15.170.5 16.870.3 19.270.8 4.28 0.97
57.771.7 65.471.7 72.072.2 74.571.7 77.972.8 5.15 3.09
6.570.4 5.970.1 6.070.3 5.970.3 6.170.3 n.s. n.s.
46.371.4 44.071.1 34.671.2 28.870.5 26.971.0 4.45 1.60
4.70 4.74 5.09 4.62 5.30
2.22 2.31 2.99 2.03 3.35
Chlorophyll b 20 30 40 50 60 LSD Po0:01 LSD Po0:05
17.470.7 30.072.5 35.471.6 36.271.2 37.972.3 3.73 2.70
5.570.4 6.570.3 6.970.4 8.670.3 10.170.6 0.81 0.58
13.570.6 21.271.5 24.971.1 26.770.8 28.471.7 2.50 1.81
2.070.1 2.770.2 2.970.1 3.270.1 3.770.2 0.31 0.23
11.070.5 14.671.0 12.470.5 11.070.3 10.970.5 1.27 0.92
0.99 2.91 1.86 1.38 2.78
0.71 2.11 1.35 1.00 2.01
a
Mean (n ¼ 4).
Table 5 Content of polyphenols in different usable parts of dill depending on height of plant, mg/100 g fresh mattera Height of plant (cm)
20 30 40 50 60 LSD Po0:01 LSD Po0:05 a
Usable part of dill
LSD
Leaf blade
Petiole
Whole leaf
Stem
Whole plant
Po0:01
Po0:05
19778 17378 214710 278711 331712 20.6 14.9
5573 5473 9277 8574 8774 9.5 6.9
15176 12976 16975 21278 24879 14.7 10.7
5373 4974 6674 5473 4973 6.3 4.6
12975 10073 11174 10773 10774 8.3 6.0
11.2 10.8 13.3 13.7 15.3
8.1 7.8 9.6 9.9 11.0
Mean (n ¼ 4).
3.5. Content of polyphenols With the increasing height of the plant, the content of polyphenols increased, with some exceptions, in leaf blades, petioles and whole leaves, while in general it showed a decreasing tendency in the stem and the whole plant (Table 5). Moreover, in the analysed parts of the plant differences in the content of these compounds according to height were not always significant. In
comparison with parts of the youngest plants where the level of polyphenols was assumed to be 100%, in the oldest plants the leaf blade contained 168%, the petiole 158%, whole leaf 164%, the stem 92%, and the whole plant 83%. Choi and Lee (1999) also recorded fewer polyphenols in younger leaves of Agastacha rugosa. However, according to Ba˜no et al. (2003) the greatest accumulation of polyphenol compounds occurred in early developmental stages of Rosmarinus officinalis.
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Contrary to the present study, Vallejo et al. (2003) wrote that in broccoli plants the content of polyphenol compounds showed a tendency to increase from the first stage to the last. If it was assumed that, separately for each height, the content of polyphenols in the whole plant was 100%, the leaf blade contained as much as 153–309% of these compounds, the petiole 43–93%, whole leaf 117–232%, and the stem 41–59%. With the exception of the petiole from a plant 60 cm in height and of the stem from plants 50 and 60 cm in height, steady growth occurred in the content of these compounds. Romani et al. (2003) also stress a high content of polyphenols in soy leaves compared with stems. However, Najda et al. (2003) demonstrated a smaller differentiation in the content of polyphenol compounds in lovage plants where the leaf blade contained 119% and the stem 38% compared with the whole plant.
4. Conclusions The highest content of all the analysed constituents was found in the leaf blade, and the lowest in the stem and the petiole. The content of vitamin C decreased with plant growth, with the exeption of petiols; the content of carotenoids and beta-carotene tended to increase with growth, with the exception of stems and whole plants. The higher the plants, the higher the level of chlorophylls and polyphenols in the analysed parts of dill, except in whole plants and, in the case of polyphenols, also in stems.
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