International Journal of Agricultural Science and Research (IJASR) ISSN 2250-0057 Vol.

3, Issue 1, Mar 2013, 63-88 TJPRC Pvt. Ltd.

INTERACTION BETWEEN SALINITY AND CROP CYCLE: EFFECT ON YIELD AND QUALITY OF LETTUCE
IDA DI MOLA, MAURO MORI & FABRIZIO QUAGLIETTA CHIARANDA Department of Agricultural Engineering and Agronomy, University of Naples Federico II Agricultural faculty via Universita 100 80055 Portici (NA)

ABSTRACT
The 55% of groundwater is saline, particularly in arid and semi-arid regions where it is needed most for irrigation. The aim of this research was to simulate two real conditions: 1) the recent intrusion of sea water into the fresh water table and 2) the prolonged use of saline water on already salinized soil. We investigated the effects of salinity on yield, nitrate content, lypophilic, hydrophilic and Vitamin C antioxidant activity of lettuce. The lettuce Cambria was irrigated with water at five levels of salinity (0.7, 0.9, 1.8, 3.6 and 7.2 dS m-1 ECw) and two growing cycle were completed (Winter Transplant and Spring Transplant) for evaluating the possible interaction with salinity. Salinity reduced yield, fresh weight, and heads diameter and leaf area with more evident damages in winter cycle. The nitrate content was never higher than European limits and it decreased at increasing salt concentration. The salt stress didnt cause an increase of antioxidant activity, but this was higher in the spring cycle, probably due to photo-oxidative strtyess (high light intensity). It seems that irrigation with water at EC not greater than 1.8 dS m-1 reduces leaves nitrate content, without to compromise excessively the yield and nutritional quality of lettuce.

KEYWORDS: Lettuce, Antioxidant Activity, Nitrate Content, Salt-Tolerance INTRODUCTION

Change models suggest an increase in global average temperature, shifts in rainfall pattern and an increase in local climatic extremes. This also means changes in evaporative demand, irradiance, ultraviolet irradiance, and secondary factors such as tropospheric ozone concentration, as well as in the prime causative agent, carbon dioxide itself (Yeo, 1998). Also, the increasing world population and urbanization have forced farmers to utilize marginal lands with saline water (Bie et al., 2004). Large sectors of the current population now live in areas in which rainfall is already inadequate to support the food crops they need and consequently they depend upon irrigation. Demand for water already exceeds the renewable supply of freshwater given the geographic and temporal patchiness of both demand and supply. Irrigation demand is often met with poor quality water containing dissolved salts which can then accumulate in the soil. Irrigation and clearance of native perennial vegetation has generally caused water tables to rise eventually they may rise into the root zone and waterlog the land. More than half of all groundwater is naturally saline, particularly in arid and semi-arid regions of the world, so rising water tables bring saline water into the root zone.

64

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

Of the water applied in irrigation an average of 4045% is used by the crop, the rest is lost as leakage from the storage/distribution network or as deep percolation below the root zone (). Groundwater is a major water resource, but 55% of groundwater is saline, particularly in arid and semi-arid regions where it is needed most for irrigation (Ghassemi et al., 1995). In particular, in Mediterranean areas water shortages often occur during spring-summer and available supplies are rich in salts, due both to the nature of the sources and to intense evaporation (Postiglione et al., 1995). Crop responses to soil salinity or irrigation with saline water imply a whole array of biochemical and biophysical phenomena at different levels, which are integrated in different physiological and productive behaviors according to environmental and management variables (Li et al, 2001). The crop response to salinity varies also with the cultivars within the species and with the phenophase within the cultivar; so, due the extreme variability of crop response, it is needed to continue to investigate the effect of soil and water salinity on agronomic behavior and yield of crops. Its known that the first evident effect of salinity is the reduction of plant growth and yield; in fact we also found this response to salt stress on other crops, such as tomato (Mori et al. 2008) and snap bean (Mori et al. 2011). The productive performances of lettuce are considered as moderately sensitive to salinity. Flagella et al. (1999) indicate that the threshold above which yield starts to decline is 1.3 dSm1 and at 9.0 dSm1 the yield losses are 100% in the Maas and Hoffman model (Maas and Hoffman, 1977). Many experiments were carried out on the effects of salt on growth and yield of crops, but not many studies were conducted on the effects of salinity on yield quality. Instead, today, the qualitative characteristics of food are an important aspect for the choice of consumers. The quality of food is constituted by physical factors as: form, color, size, consistency, which can evaluate without laboratory analysis, and chemical factors as: content of antioxidants, nitrate, vitamins, etc., which can to be only determined by laboratory analysis. Some chemical characteristics are more important for some crops than the other ones; for example leafy vegetables, such as lettuce and spinach, are especially prone to nitrate accumulation (Siomos et al., 2002). Numerous studies have studied the effects of nitrate intake on human health. Although earlier reports implicating nitrate in the occurrence of cancer are largely unsubstantiated, other nitrate-induced syndromes, such as methaemoglobinaemia in infants (Blue baby syndrome) have been confirmed (Addiscott and Benjamin, 2004; Fewtrell, 2004). Moreover, the reduction of nitrates to nitrites and the subsequent formation of carcinogenic nitrosamines within the gastroenteric system have been reported and this observation has contributed to the adoption of maximum acceptable nitrate levels in lettuce and spinach by the European Union (Siomos et al., 2002). Nitrate uptake and accumulation in plants primarily relates to the amount (Chen et al., 2004; Mantovani et al., 2005) and form of N-fertilizer applied to the crop (Abu-Rayyan et al., 2004; Breimer, 1982; Lairon et al., 1984; Premuzic et al., 2002). Other factors inurning this process include the plant genotype, nitrate reductase activity, plant age and the cultivation season and environment (Boroujerdnia et al., 2007; Drews, 1996; Drews et al., 1997).

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

65

Nitrate accumulation tends to be usually higher in the winter than in other seasons (Burns et al., 2004; Ysart et al., 1999) due to the reduction of nitrate reductase activity within the leaves under low light intensity and low temperatures (Riens and Heldt, 1992). Lettuce is the more common leafy vegetable, mainly consumed fresh in salads (Altunkaya et al., 2009), which are consumed in increasing amounts due to their perception as being healthier foods (Dupont et al., 2000); besides, the lettuce is an important source of dietary antioxidants especially considering its high peroxyl radical scavenging activity (Caldwell, 2003). The antioxidants can be divided into three general classes including: lipid-soluble and membrane-associated atocopherol and b-carotene; water-soluble reductants, ascorbate and glutathione; enzymes such as superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), ascorbate peroxidase (APX) and glutathione reductase (GR) (Sudhakar et al., 2001). The intake of these bioactive compounds in food is an important health-protecting factor. Therefore, plant-based diets are widely suggested to contribute to reducing the risk of development of chronic diseases such as cancer, atherosclerosis, cardiac dysfunctions, diabetes, hypertension and neurodegenerative disorders (Bahorun et al., 2004; Bramley, 2000; Sies, 1991). These healthy properties of lettuce are attributed to a large supply of antioxidant compounds, mainly vitamin C (Martn-Diana et al., 2007) and polyphenols (Heimler et al., 2007). Vitamin C is a universal constituent of all green leaf plants, its minimum intake must be of 60mg day for adults. In crop plants, vitamin C content is influenced by seasonal factors such as sowing time and harvesting date and is often used as a marker for postharvest deterioration (Sood Mand Malhotra, 2001). Ascorbate occurs in the cytosol, chloroplasts, vacuoles, mitochondria and cell wall (Anderson, et al., 1983; Rautenkranz et al., 1994). The concentration in chloroplasts can be high and is probably related to its central role in photosynthesis (Foyer, 1993). In fact, it develops an important role in photosynthesis and photo-protection, in defense against ozone and other oxidative stresses (Smirnoff, 1996). In particular, in plants, L -ascorbate has been implicated in processes including growth (Pignocchi and Foyer, 2003), programmed cell death (de Pinto et al., 2006), pathogen responses (Barth et al., 2004), hormone responses, owering and senescence (Barth et al., 2006), as well as protection against environmental stresses including ozone (Conklin and Barth, 2004), UV radiation (Gao and Zhang, 2008), high temperatures (Larkindale et al., 2005) and high light intensity (Muller-Moul et al., 2004). The antioxidant compounds are susceptible to variation among varieties, growing practices, processing and storage conditions on the biologically active compounds (Baur et al., 2004; Dupont et al., 2000; Lee & Kader, 2000). The antioxidant content of plants varies considerably depending also on their growing and management conditions (Lee and Kader, 2000; Zhao et al., 2006). It has been demonstrated that antioxidants in plants are part of a complex defence mechanism against a wide range of stresses and thus, accumulate in response to these stresses (Blokhina et al., 2003; Dixon and Paiva, 1995). The effects of various environmental stresses in plants are known to be mediated, at least in part, by an enhanced generation of activated oxygen species (AOS; O2, H2O2 andOH) (Alscher et al., 1997; Hernndez et al. 1993, 1995; Mittler & Zilinskas 1994; Noctor & Foyer 1998). Although a wide range of genetic adaptations to saline conditions has been observed and a number of significant physiological responses have been associated with tolerance, the underlying

66

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

mechanisms of salt-tolerance in plants are still poorly understood (Hernndez et al. 2001). However, one determinant of salt tolerance could be how well stressed plants deal with the accompanying oxidative damage to sub cellular compartments (Gueta-Dahan et al., 1997; Hernndez et al. 1993 , 1995; Lpez et al. 1996; Van Camp et al. 1996). To mitigate and repair damage initiated by AOS, plants have developed a complex antioxidant system. The primary components of this system include carotenoids, ascorbate, glutathione and tocopherols, and some enzymes (Hernndez et al. 2001). Maggio et al. (2007) found that in tomato fruits salinity didnt affect the antioxidant activity of ascorbic acid and tocopherols but only one of carotenoids (they increase at salinity increased) and total phenolic compounds (they decrease with saline stress). The mechanisms of defense of plants to the stresses arent already completely known, besides each crop can have a different response in function of numerous variables: type of stress, crop genetic background, climatic conditions, agronomic practices, etc. The aim of this research was to simulate two real conditions: 1) the use of increasing salt concentration of water irrigation on non saline soil, that simulate the discovery of a saline wedge and, therefore, the intrusion of sea water into the fresh water table; 2) the long term use of increasing salt concentration of water irrigation with corresponding soil salinization. So, we wanted to appraise the different effects of salt stress on yield and quality of lettuce, in function of salinity is short term (not salinized soil) or long term (salinized soil).

MATERIAL AND METHODS

The trials were made in Portici, at experimental field of Naples Faculty of Agriculture (N 40 48.870; E 14 20.821; 70m slm) in 2005-2006. The shelter was a 116 m2 (14.5m x 8m) permanent metal structure, open on the sides and with a clear PVC top. The lettuce (Lactuca sativa L., var. capitata), cv Cambria was transplanted in lysimeters of 0.38 m2 (diameter of 0.7 m) and 0.60 m deep, placed on bricks at 0.2 m from the soil surface. Lysimeters were filled with 0.10 m of gravel for drainage and 0.45 m of loamy sandy soil (ISSS classification, tab. 1). For these trials the loamy sand soil was chosen for avoiding salt indirect effects on soil physical properties like: swelling, reduction of hydraulic conductivity and related phenomena visible in clayey soils, as reported by Rhoades et al. (1992) e De Pascale and Barbieri (2000). Two tests were carried out and for both, each treatment was replicated 6 times. In the first one, lettuce was irrigated with 5 levels of saline water irrigation and it was cultivated on not salinized (NS) soil: 1. 2. 3. 4. 5. 6. No saline stress: irrigation with freshwater (ECw 0.7 dS m-1) (NS1); Low saline stress: water at 0.9 dS m-1 electrical conductivity (NS2). Moderate saline stress: water at 1.8 dS m-1 electrical conductivity (NS3). Severe saline stress: water at 3.6 dS m-1 electrical conductivity (NS4). Very severe saline stress: water at 7.2 dS m-1 electrical conductivity (NS5). At the begin of trial, the average EC soil was 0.34 dS m-1. In the second test, the same 5 levels of saline irrigation water were tested, but lettuce was cultivated on saline soil (SS) with a specific initial EC corresponding to each treatment: 1. 2. Non saline soil: 0.32 dS m-1 (SS1) and fresh water irrigation Low saline stress soil: 0.71 dS m-1 (SS2) and irrigation water at 0.9 dS m-1

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

67

3. 4. 5.

Moderate saline stress soil: 1.08 dS m-1 (SS3) and irrigation water at 1.8 dS m-1 Severe saline stress soil: 1.38 dS m-1 (SS4) and irrigation water at 3.6 dS m-1. Very severe saline stress soil: 1.51 dS m-1 (SS5) and irrigation water at 7.2 dS m-1. Table 1: Soil Physical and Chemical Characteristics Texture - Coarse sand _ Fine sand - Silt - Clay Ntotal (Kjeldahl method) P assimilable (Olsen method) K assimilable (ammonium acetate method) Organic matter (bichromate method) Bulk density

% % % % % ppm ppm % kg dm-3

47.0 36.6 11.9 4.5 0.54 6.7 107.0 0.72 1.32

All lysimeters were well watered restoring the 100% of water uptake (ETc). Crop evapotraspiration (ETc) was estimated by the agro-meteorological approach based on Hargreaves formula (to calculate ETo) and the crop coefficient always fixed at 1 to allow leaching from the root zone. Saline water for the stressed treatments was obtained by diluting marine salt in freshwater. Each level of ECw was reached by adding salt according to the relation: Salt = 0,64 x EC The electrical conductivity of the solution (ECw) was checked with a conductimeter (Basic 30 CRISON). For evaluating also the interaction between salinity and climatic condition, two growth cycles for every trial were made: the first one with winter transplant (WT) on first December week and harvest on 03 and 09 March; the second one with spring transplant (ST) on 10 March and harvest in three times from 03 to 15 May. The winter cycles were long 92 days and the spring cycles were long 66 days. In both trials, three plants per lysimeter were transplanted at a distance of 0.30 m between them (8 plants m-2). Before transplant, 26.0 g of superphosphate and 11.4 g of potassium sulphate were applied in each lysimeter, corresponding at 130 kg ha-1 of P2O5 and 150 kg ha-1 of K2O. The nitrogen, as ammonium nitrate (26% N), was given at the rate of 15.0 grams per lysimeter, corresponding to 100 kg ha-1: 1/3 at transplant, 1/3 after 35 days and 1/3 after further 25 days, in the winter cycles, and 1/3 at transplant, 1/3 after 30 days and 1/3 after further 20 days, in the spring cycles. At transplant and in three successive dates, 9 litres of freshwater were applied to each lysimeter in order to help plant establishment. Starting from day 35 after transplant (DAT) in the winter cycles and at 28 DAT in the spring cycles, lysimeters were irrigated with saline water. In the WT cycles the waterings were about once a week, in the ST cycles about twice a week. For the WT cycles saline waterings were 9 with an average volume of 2.0 l per lysimeter; in total 10.6, 21.2, 42.5 and 85.0 g salt per lysimeter were given respectively for treatments ECw 0.9, ECw 1.8, ECw 3.6 and ECw 7.2.

68

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

For the ST cycles saline waterings were 11 with an average volume of 3.2 l per lysimeter; in total 22.1, 44.2, 88.4 and 176.8 g salt per lysimeter were given respectively for treatments ECw 0.9, ECw 1.8, ECw 3.6 and ECw 7.2. At harvest the following measurements were made: number and area (measured with a Li-Cor 3000 Leaf area meter, Li-Cor Lincoln NE) of leaves, fresh weight and diameter of head, content of nitrate with spectrophotometer (DR 2000, Hach Co, Loveland, CO) on three samples of green leaves dry matter (the nitrate values were reported to the fresh weight), lypophilic antioxidant activity with ABTS (2,2-azinobis (3-ethylbenzothiazoline-6-sulphonic acid) assay (Miller et al., 1995; Pellegrini et al., 1999), hydrophilic antioxidant activity and the vitamin C antioxidant activity (Abushita et al., 1997) with DMPD (N, N-dimetilanilina diidrocloruro) assay (De Pascale et al., 2001; Fogliano et al., 1999), on three samples of green leaves fresh matter. Both methods base on a valuation of absorption reduction, at the presence of antioxidant compounds, at fixed wave length: 734 nm for the ABTS radical cation and 505 nm for the DMPD radical cation. Therefore, the antioxidant activity is expressed as percentage decrease of absorbance or inhibition percentage, according equation: Abs (%) = (1-Absc/Absb)*100 Where: Absc is the cation absorbance after the addition of antioxidant extracted by sample Absb is the radical cation absorbance with distilled water. The samples made for determination of lypophilic, hydrophilic and vitamin C antioxidant activity were read by spectrophotometer (Hach DR 4000 U). The hydrophilic antioxidant activity and the Vit C antioxidant activity were expressed in mmol of ascorbic acid (AA) per 100 g of fresh leaves. The lypophilic antioxidant activity was expressed as Trolox Equivalent Antioxidant Capacity (TEAC) in mmol of trolox per 100 g of fresh leaves (Re et al., 1999). All data were analyzed with MSTAT software (Crop and Soil Science Department, Michigan State University, Version 2.0).

RESULTS
Soil Salinity Irrigation with saline water resulted in increased values of average soil salinity throughout the plant cycles. About Test 1, soil salinity always linearly increased. In the winter cycle (fig. 1A), EC soil increased at rate of 0.044 dS m-1 per each dS m-1 of saline water; instead in the spring cycle (fig. 1B), the increase rate was higher: 0.127 dS m-1 (almost three times more than winter cycle) per each dS m-1 of saline water.
1.5 1.2 dS m 0.9 0.6 0.3 0 0.0 2.0 4.0 ECw (dS m -1 ) 6.0 8.0
EC soil Linear (EC soil) y = 0.0441x + 0.4526 R2 = 0.7514
-1

Figure 1: A. Soil EC (1 to 5 Method) of Test 1 Lysimeters Vs EC Water in the Winter Cycle

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

69

1.5 1.2 dS m 0.9 0.6 0.3 0 0.0 2.0 4.0 ECw (dS m -1 ) 6.0 8.0
EC soil

y = 0.1269x + 0.5327 R2 = 0.9669

-1

Linear (EC soil)

Figure 1: B. Soil EC (1 to 5 Method) of Test 1 Lysimeters Vs EC Water in the Spring Cycle About Test 2, the electrical conductivity (dS m1) of water irrigation determined an increase of soil EC more marked for the two lower levels of salinity and less evident for the other two ones. In both cycles, this behaviour is best tted with a logarithmic model. In the winter cycle (fig. 2A), the marginal increase in EC soil ranged from about 0.34 dS m-1 at low salinity water (until to 2.0 dS m-1 of EC water) to about 0.12 dS m-1 for each dS m-1 at high salinity water (until to 7.0 dS m-1 of EC water ). Instead, in the spring cycle (fig. 2B), the marginal increase in EC soil was more marked and ranged from about 0.48 dS m-1 at low salinity water (until to 2.0 dS m-1 of EC water) to about 0.17 dS m-1 for each dS m-1 at high salinity water (until to 7.0 dS m-1 of EC water). During the two cycles of the two tests, the pH showed a low increase but without significant differences between the saline treatments and between lysimeters; the average increase was 0.55.
A

2.8 2.4 2.0 1.6 1.2 0.8 0.4 0.0 0.0 2.0 4.0 ECw (dS m -1 ) 6.0 8.0

dS m

y = 0.4889Ln(x) + 0.9383 R2 = 0.9035

-1

EC soil Log. (EC soil)

Figure 2: A. Soil EC (1 to 5 Method) of Test 2 Lysimeters vs EC Water in the Winter Cycle

B

2.8 2.4 2.0 1.6 1.2 0.8 0.4 0.0 0.0 2.0 4.0 ECw (dS m -1 ) 6.0 8.0

dS m

y = 0.686Ln(x) + 1.1264 R2 = 0.9785

-1

EC soil Log. (EC soil)

Figure 2: B. Soil EC (1 to 5 Method) of Test 2 Lysimeters vs EC Water in the Spring Cycle

70

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

Temperatures The external temperature trend was that typical of Mediterranean environments (fig. 3), only the minimum temperatures of winter months were lightly lower.
25 C 20
T min 2005-06 T max 2005-06

15

T min 1921-50 T max 1921-50

10

0 Dec Jan Feb M ar Apr M ay

Figure 3: Min and Max Temperatures During the two Crop Cycles Compared to 30 Years Mean Values The temperatures under the shelter (fig. 4) were always higher than external temperatures; in particular, they have been in average 4C higher than those external for maximum temperatures and 3C for minimum temperatures.

35.0 30.0 25.0 20.0 15.0 10.0 5.0 0.0 I II III I II III I II III I II III I II III I II III Dic Jan Feb Mar Apr May
WT ST
Average Minimum Maximum

Figure 4: Temperatures Pathway in the Shelter During the Two Cycles During the winter cycles, the daily thermic excursion was 8.3C on the average with the minimum temperatures that were from 7.5 to 11.3C and the maximum ones from 16.2 to 19.0C; instead, during the spring cycles, the average daily thermic excursion was 9.8C with the minimum temperatures that were from 9.0 to 16.2C and the maximum ones from 16.9 to 26.8C.

TEST 1
Yield The main effect of salinity and crop cycle was significant for yield, head fresh weight and diameter, green leaves number and leaf area (tab. 2).

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

71

Table 2: Analysis of Variance of Test 1: Significance of Main Factors and Interaction (P <0.05=*; P<0.01=**) Main Factors Crop Cycle Salinity Significativity DMS Significativity DMS Head fresh weight Yield Diameter Green leaves number Succulence Specific leaf weight Leaf area Hydrophilic antioxidant activity Vitamin C activity Lypophilic antioxidant activity Nitrate content * * ** 42.8 0.33 1.83 ** ** ** 67.68 0.52 2.89 Interaction Crop Cycle x Salinity Significativity DMS n.s. n.s. n.s. -

* n.s. n.s. **

5.51 295.05

** n.s. n.s. **

8.71 466.51

n.s. n.s. n.s. n.s.

* **

0.13 -

** **

0.21 -

n.s. **

0.05

* *

0.25 -

** **

0.39 -

n.s. *

829.46

The yield linearly decreases at the increase of soil salinity (fig. 5A and 5B). In the winter cycle, the rate of yield decrease was about 185.3% vs. the control per each dS m-1 of soil salinity. In the spring cycle, instead, it was lower: 77.2% vs. the control per each dS m-1 of soil salinity.
A 120.0 % 100.0 80.0 60.0 40.0 20.0 0.0 0 0.3 0.6 0.9 1.2 1.5 EC soil (dS m -1 ) y = -185.28x + 169.25 R = 0.7677
2

Figure 5: A. Test 1: Percentage Yield of Winter Cycle Lettuce Vs the Control in Function of EC Soil
B 120.0 % 100.0 y = -77.206x + 138.58 80.0 60.0 40.0 20.0 0.0 0 0.3 0.6 0.9 1.2 1.5 EC soil (dS m -1 ) R = 0.8944
2

Figure 5: B. Test 1: Percentage Yield of Spring Cycle Lettuce Vs the Control in Function of EC Soil

72

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

The yield reduction was proportional to the head fresh weight reduction (tab. 3). The control was always different from the all other treatments with the most stressed treatment (NS5) it was different also from the other saline treatments. The yield decreased also because the number of plants m-2 decreased. In fact, the density of lettuces of high salt concentration treatment (NS5) was 5.1 vs. an average value of 6.3 plants m-2 of the other three treatments (data not showed). The diameter of lettuce heads (tab. 3) was less influenced by salt: the control was statically different only from the two more stressed treatments. The effect of salinity on number of green leaves was still less evident (tab. 3); in fact, only the NS5 treatment was different from the all other ones. Table 3: Main Effect (Salinity and Crop Cycle) on Yield, Weight, Diameter and Green Leaves Number of Lettuce Head of Test 1. The Same Letters Show Not-Significant Differences Per P0.05 Yield kg m-2 2.35 a 1.79 b 1.70 b 1.52 b 0.50 c 1.21 b 1.93 a Weight g head-1 304.78 a 232.68 b 221.22 b 196.72 b 64.54 c 157.07 b 250.91 a Diameter cm 29.25 a 27.00 ab 26.68 ab 25.50 b 17.28 c 22.80 b 27.48 a Green Leaves Number n 50.25 a 43.00 a 45.25 a 42.25 a 25.25 b 36.20 b 46.20 a

Treatments NS1 NS2 NS3 NS4 NS5 Winter Spring

The leaf area linearly decreased at the increase of salt stress (fig. 6); the decrease rate was 289.0 cm2 per plant per each dS m-1 of water salinity, with the NS5 it was only 24% of control.
a 3000.0 2500.0 b cm plant 2000.0 1500.0 1000.0 500.0 0.0 0.0 2.0 4.0 EC water (dS m -1 ) 6.0 8.0 d
-1

y = -289x + 2781.7 R = 0.9526 bc c

leaf area Linear (Leaf area)
2

Figure 6: Leaf Area (Cm2 Plant-1) Vs EC Water of Test 1 Lettuce Besides, the leaf area of spring lettuce was significantly higher than the one of winter lettuce (fig. 7); it was more than double.
3000.0 2500.0
-1

2000.0

cm plant

b
1500.0 1000.0 500.0 0.0 Winter Spring

Figure 7: Effect of Crop Cycle on Leaf Area (Cm2 Plant-1) of Test 1 Lettuce

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

73

Also for the all other parameters (tab. 3), the spring cycle always showed the higher values, due probably to a greater percentage survival of seedlings and a greater development of heads. Yield Quality The quality of yield was investigated and the statistic analysis showed that the interaction salinity and crop cycle affected nitrate content and vitamin C antioxidant activity of lettuce heads, while the main effects were significant for lypophilic and hydrophilic antioxidant activity (tab. 2). The hydrophilic antioxidant activity (HAA) decreased with the increase of salt level (fig. 8); particularly, it decreased at rate of 0,11 mM ascorbic acid/100g of leaves fresh weight per dS m-1 of water irrigation, with NS5 (water at EC 7.2) it was 61% of the control and it was statistically different from the all other treatments. Besides, the HAA values of spring cycle (fig. 9) were higher than those of winter cycle, about more than 20%.
y = -0.1145x + 2.3006 R2 = 0.8375

3.0
HAA (mmol AA on 100 g leaves FW)

2.5 2.0 1.5 1.0 0.5 0.0 0.0

a b b c b

HAA Linear (HAA)

2.0

4.0 6.0 ECw (dS m-1 )

8.0

Figure 8: Effect of Electrical Conductivity of Water (ECw) on Hydrophilic Antioxidant Activity (HAA) of Test 1 Lettuce
HAA (mmol AA on 100 g levaes FW)

3.0 2.5 2.0 1.5 1.0 0.5 0.0 Winter Spring b a

Figure 9: Effect of Crop Cycle on Hydrophilic Antioxidant Activity (HAA) of Test 1 Lettuce The effect of saline water on LAA was greater (fig. 10) with a decreasing rate of 0,29 mM trolox/100g of leaves fresh weight per dS m-1 of water irrigation; NS5 was 55% of the control and, like NS4, it was statically different from the other two saline treatments and from the control.
LAA(mmol trolx on 100 g leaves FW)

6.0 5.0 4.0 3.0 2.0 1.0 0.0 0.0

a b b c d
LAA Linear (LAA) y = -0.2898x + 4.9253 R2 = 0.8025

2.0

4.0 ECw (dS m -1 )

6.0

8.0

Figure 10: Effect of Electrical Conductivity of Water (ECw) on Lypophilic Antioxidant Activity (LAA) of Test 1 Lettuce

74

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

Also the LAA showed the highest value in the spring cycle; it was about 31% more than the winter value (fig. 11).
LA A (mmol trolox on 100 g leaves FW )

6.0 5.0 4.0 3.0 2.0 1.0 0.0 Winter b

Spring

Figure 11: Effect of Crop Cycle on Lypophilic Antioxidant Activity (LAA) of Test 1 Lettuce The Vitamin C antioxidant activity (fig. 12) showed a decreasing linear trend; it was more marked in the winter cycle, with a rate of decrease was 0.10 mM ascorbic acid/100g of leaves fresh weight per dS m-1 of water irrigation. Instead, in the spring cycle, the rate was 0,04 mM ascorbic acid/100g of leaves fresh weight per dS m-1.

Vit C (mmol AA on 100 g leaves FW)

2.4 2.0 1.6 1.2 0.8 0.4 0.0 0.0

y = -0.1053x + 1.9769 R = 0.8889

2

DMS=0.05

Winter y = -0.0439x + 1.7271 R2 = 0.5331 Spring Linear (Winter) Linear (Spring)

2.0

4.0

6.0

8.0

Figure 12: Interaction of Salinity and Crop Cycle on Vitamin C Antioxidant Activity of Test 1 Lettuce Also the nitrate content (fig. 13) linearly decreased at the increase of salt concentration. In the winter cycle, the nitrate content decreased at rate of 341.5 ppm per each dS m-1 of water salinity; instead, in the spring cycle, the values of all treatments were lower, like the rate of decrease, which was 97.7 ppm per each dS m-1 of water salinity. The all saline treatments of spring cycle were statistically not different from the 2 more stressed treatments of winter cycle.
4500 4000
NO3 (ppm on leaves FW)

DMS=829.46
y = -341.5x + 3392.2 R2 = 0.7558 Winter Spring Linear (Winter) Linear (Spring) y = -97.737x + 1755.7 R2 = 0.6401

3500 3000 2500 2000 1500 1000 500 0 0.0

2.0

4.0

6.0

8.0

Figure 13: Interaction of Salinity and Crop Cycle on Nitrate Content of Test 1 Lettuce

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

75

TEST 2
Yield For all parameters of yield (head fresh weight, diameter, number of green leaves and leaf area) the interaction between the two main factors (crop cycle and salinity) was significant (tab. 4). Table 4: Analysis of Variance of Test 2: Significance of Main Factors and Interaction (P <0.05=*; P<0.01=**) Main Factors Interaction Crop Cycle Salinity Crop Cycle x Salinity Significativity DMS Significativity DMS Significativity DMS Head fresh weight Yield Diameter Green leaves number Succulence Specific leaf weight Leaf area Hydrophilic antioxidant activity Vitamin C activity Lypophilic antioxidant activity Nitrate content ** ** * * n.s. * ** ** ** ** ** ** ** ** ** ** * * ** * ** 56.74 0.44 2.68 8.12 11.06 0.98 532.50

** **

** **

** **

0.23 0.05

** n.s.

0.58 -

** **

0.92 -

n.s. **

350.92

The yield linearly decreased at the increase of soil salinity (fig. 14A and 14B). In the winter cycle, the rate of yield decrease vs. the control was 59.9% per each dS m-1 of soil salinity and in the spring cycle it was just lower, about 51.3%.

120.0 100.0 80.0 % 60.0 40.0 20.0 0.0 0.0 0.5 1.0 1.5
)

y = -59.886x + 106.41 R = 0.7418

2

2.0

2.5

EC soil (dS m -1

Figure 14: A. Test 2: Percentage Yield of Winter Cycle Lettuce Vs the Control in Function of EC Soil

76

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

120.0 100.0 80.0 % 60.0 40.0 20.0 0.0 0.0 0.5 1.0 1.5
-1 )

y = -51.293x + 123.97 R = 0.9801

2

2.0

2.5

EC soil (dS m

Figure 14: B. Test 2: Percentage Yield of Spring Cycle Lettuce Vs the Control in Function of EC Soil The yield values of spring cycle (tab. 5) were higher, except those of the two more stressed treatments; they were statistically not different from the all saline treatments of winter cycle. Also the head fresh weight decreased at the increase of salinity (tab. 5) but, like for the Test 1, the yield also decreased because the number of plants m-2 decreased. In fact, the density of lettuces decreased of the 30% vs. the control, for the SS2, SS3 and SS4 of winter cycle and, even, of the 44% for the SS5. In the spring cycle, the reduction of number of plants m-2 vs. the control was about 28% for the SS2 and SS3, 55.5% for the SS4 and 66.6% for the SS5 (data not showed).The salinity also had a depressing effect on heads diameter (tab.5), with the winter cycle it had lower values than those of spring cycle but, the percentage of diameter decrease was higher in the spring cycle. The number of green leaves decreased at the increase of salt stress (tab. 5); in fact the values of the two most stressed treatments of both cycles were less of the half of the value of the corresponding control. The values of the spring cycle treatments were always greater except for the two more stressed treatments; they were not different from the corresponding treatments of winter cycle. Table 5: Interaction between Salinity and Crop Cycle on Yield, Weight, Diameter and Green Leaves Number of Lettuce Head of Test 2. The Same Letters Show Not-Significant Differences Per P0.05 Yield kg m-2 Winter Spring 1.94 b 0.38 d 0.34 d 0.30 d 0.17 d 2.76 a 1.72 b 1.16 c 0.39 d 0.15 d Weight g head-1 Winter Spring 251.2 b 50.4 d 44.4 d 38.6 d 22.3 d 358.4 a 222.4 b 151.0 c 50.8 d 19.7 d Diameter cm Winter Spring 26.2 b 16.8 d 16.4 d 13.2 e 11.5 e 32.2 a 24.6 b 21.4 c 16.4 d 12.0 e Green leaves n head-1 Winter Spring 45.5 b 27.5 c 24.5 c 22.5 cd 20.0 cd 55.0 a 40.0 b 40.0 b 24.0 c 15.0 d

Cycle Salinity SS1 SS2 SS3 SS4 SS5

The heads of spring cycle showed also the highest values of leaf area (fig. 15), but however the leaf expansion was greatly depressed by salt stress. In fact, the leaf area of SS5 (water at EC 7.2 dS m-1) of winter cycle was 14.3% of the corresponding control but, even, the leaf area of SS5 of spring cycle was 5.1% of the corresponding control. The LA decrease was more marked for the two lower levels of salinity and less evident for the other two ones. In both cycles, this behaviour is best tted with a power model. In the winter cycle, the marginal decrease of leaf area ranged from about 305.1 cm2 plant-1 for each dS m-1 at low salinity water (until to 2.0 dS m-1 of EC water) to about 61.7 cm2 plant-1 for each dS

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

77

m-1 at high salinity water (from 3.0 dS m-1 of EC water). Instead, in the spring cycle, the marginal decrease of leaf area was more marked and ranged from about 1150.5 cm2 plant-1 for each dS m-1 at low salinity water (until to 2.0 dS m-1 of EC water) to about 149.1 cm2 plant-1 for each dS m-1 at high salinity water (from 3.0 dS m-1 of EC water).
DM S = 532.50

5000 4500 4000 3500 3000 2500 2000 1500 1000 500 0 0.0

y = 2142.9x
2

-1.1106

R = 0.9709
Winter Spring Power (Spring) Power (Winter)

cm plant

-1

y = 878.91x
2

-0.6151

R = 0.8463

2.0

4.0 EC water (dS m )

-1

6.0

8.0

Figure 15: Interaction of Salinity and Crop Cycle on Leaf Area (Cm2 Plant-1) of Test 2 Lettuce Yield Quality The statistic analysis found that the interaction saline water irrigation and crop cycle affected the nitrate content, the vitamin C and the hydrophilic antioxidant activity, while the main effects of two factors were significant for the lypophilic antioxidant activity (tab. 4). Like for the test 1, the hydrophilic antioxidant activity decreased with the increase of salt level (fig.16); besides, the HAA values of spring lettuces were always higher than ones of winter lettuces. The HAA decrease rate was 0,22 and 0.28 mM ascorbic acid/100g of leaves fresh weight per each dS m-1, for spring and winter lettuces respectively.
DM S= 0.23
y = -0.221x + 2.745 R2 = 0.8588 Winter

2.8 HAA (mmol AA on 100 g leaves FW) 2.4 2.0 1.6 1.2 0.8 0.4 0.0 0 1 2 3 4 5 6
y = -0.281x + 2.135 R2 = 0.743

Spring Linear (Winter) Linear (Spring)

Figure 16: Interaction of Salinity and Crop Cycle on Hydrophilic Antioxidant Activity (HAA) of Test 2 Lettuce The Vitamin C antioxidant activity (fig.17) also linearly decreased at the increase of salt stress; particularly, the rate of decrease was 0.03 and 0.12 mM ascorbic acid/100g of leaves fresh weight per each dS m-1 of saline water, for spring and winter lettuces respectively.

78

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

2.4
Vit C (mmol AA on 100 g leaves FW)

DM S= 0.05 y = -0.0295x + 1.6645 R2 = 0.4089 Winter Spring Linear (Winter) y = -0.1175x + 1.595 R = 0.4676
2

2.0 1.6 1.2 0.8 0.4 0.0 0.0 2.0

Linear (Spring)

4.0

6.0

8.0

Figure 17: Interaction of Salinity and Crop Cycle on Vitamin C Antioxidant Activity of Test 2 Lettuce The lypophilic antioxidant activity (fig. 18) decreased at a rate of 0.272 mmol Trolox/100 g of leaves fresh weight per each dS m-1 of water irrigation, but only the control was statistically different from the all saline treatments.
6.0
LAA(mmol trolox on 100 g leaves FW) y = -0.272x + 4.2717 R2 = 0.4821

5.0 4.0 3.0

b b
LAA Linear

2.0 1.0 0.0 0.0 2.0

b b

4.0 ECw (dS m -1 )

6.0

8.0

Figure 18: Effect of Electrical Conductivity of Water (ECw) on Lypophilic Antioxidant Activity (LAA) of Test 2 Lettuce Besides, as said about HAA, the values of spring cycle were significantly greater than the ones of winter cycle (Fig. 19).
LAA(mmol trolox on 100 g leaves FW)

6.0 5.0 4.0 3.0 2.0 1.0 0.0 Winter Spring

LAA

Figure 19: Effect of Crop Cycle on Lypophilic Antioxidant Activity (LAA) of Test 2 Lettuce The nitrate content (fig. 20) of lettuce leaves decreased with the increase of salt stress. The average rate of decrease, for both cycles, was higher at low level of salt stress (until 2.0 dS m-1), and it was 1472.59 ppm of NO3 and

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

79

1315.20 ppm of NO3 for winter and spring lettuces respectively; while at high level of salt stress it was 288.17 ppm of NO3 and 591.90 ppm of NO3 for winter and spring lettuces respectively.
DM S= 350.92

4500 4000
ppm NO3 on leaves FW

3500 3000 2500 2000 1500 1000 500 0 -500 0.0 2.0 4.0 6.0 8.0
y = -592.95Ln(x) + 1520.7 R2 = 0.9489 y = -970.97Ln(x) + 1809.1 R2 = 0.4624 Winter Spring Log. (Winter) Log. (Spring)

Figure 20: Interaction of Salinity and Crop Cycle on Nitrate Content of Lettuce Head of Test 2

DISCUSSIONS
At the end of the two trials (May), the irrigation with saline water determined a salinity increase in the top 30 cm soil: the average increase of EC due to saline waterings was 181.6% and 54.5% for the Test 1 and Test 2 lysimeters respectively. The rate of EC soil increase was always higher in the spring cycles. This trend of salt accumulation probably depend by the higher temperatures of spring, those caused also higher water evaporation from the soil surface with consequent salts concentration. This phenomenon is more evident in a soil as that of our test (sandy loam); in fact, also Barbieri et al., (1994) reported that, during soil drying, the increase speed of salt concentration is lower in the clayey soil than in sandy soil. The salinity increase of soil affected the development of lettuce heads and yield; in fact, the effects were strongest on lettuces of test 2. We fixed the threshold of marketable weight at 150 g per head and it was even more abridging than the fixed one by Reg. (CE) n. 1543/01 of 27/07/01: it is 100 g for lettuce (var. capitata) grown in protected environment (greenhouse, tunnel, etc.) and 150g for lettuce grown in open field. In particular, in the Test 1 the heads reached the marketable weight and the lettuces irrigated with water at 7.2 dS m-1 only showed the lowest values. Instead, in the Test 2 the all saline treatments didnt reach 150 grams per head, except the two less stressed treatments of spring cycle. This result is according with the results of other authors: Eraslan et al. (2007) found that the weight of lettuce treated by 40 mM of NaCl was less of half of the weight of control (about 82 g per plant vs. almost 187 g per plant). We found that the lower fresh weight of lettuce was due partly to a lower number of green leaves, but greatly to a lower leaf expansion, especially in the winter cycle. In fact, like De Pascale and Barbieri (1995), we also found that leaf area was strongly affected by salinity. On the other hand, also Bie et al. (2004) found that the lettuce leaf area decrease to increase of salt stress, whether the stress is due to Na2SO4 or whether it is due to NaHCO3. Munns and Termaat (1986) suggest that the LA decrease is a short-time response to salinity, governed by the response of root to the low water potential in the soil, and that this response is communicated to the shoot via growth regulators.

80

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

In fact, the first effect of salinization is a lowering of water potential of soil, so for the plants the water uptake is more difficult and they show the same symptoms of water stress, as: limited growth, reduced leaf area and, finally, also yield decrease. In effect, when the salinity increase and soil water potential decrease, also root water potential and leaf water potential decrease. In particular, turgor potential decrease and many authors think that it is principal variable to check stomatal behaviour (Millar et al., 1971; Turner, 1974), therefore stomatal conductance also decrease and consequently photosynthesis decrease. So, it is possible that the leaf area reduction, in addition to photosynthesis decrease (data also not published), caused yield losses in the two tests. Munns et al. (1995) proposed a model of biphasic response of the plants to salinity. The plants growth is initially reduced, because the roots absorb less water, due to low water potential of soil. In this phase plant suffers osmotic stress in function only of salt concentration and osmotic pressure, but not of the salt type. Then, the further inhibition of plant growth is due to toxic stress, initially shown in the old leaves; those die because increasing salt concentration. When the abscission of old leaves is greater than neosynthesis, the availability of assimilated decreases such as plant growth. The effects of salinity were more marked in the winter cycle; in fact, the winter yield loss was almost 37% for Test 1 and it was about 49% for Test 2. In effect, these differences among the two cycles probably are due to the better environmental condition of spring cycle. The higher temperature, but especially higher radiation, probably favoured the development of heads and allowed to the plants to satisfy the greater energetic demands needed for correcting the ionic unbalance metabolism (Helal and Mengel 1981). In fact, Dalton et al. (2000) found that high levels of CO2 and light radiation are able to reduce salt damages. The yield reduction, observed in the two tests, may be attributed, in addition to osmotic and toxic stress caused by sodium and chloride concentrations in the soil solution, also to the mineral nutrition disorders. In fact, a high presence of sodium in the soil solution can determine a competition with other cations both on soil exchange complex and at level of root absorption sites. Van Hoorn et al. (1993) showed that, at high level of Na in soil solution, the sodium concentration in biomass increase while Ca and Mg concentration decrease. In tomato, De Pascale et al. (2001) also found there is a reduction of P, K, Mg e nitrate uptake. On the other hand, we also found, in both tests, that N uptake (NO3 content of leaves) reduced. The nitrate levels in leafy vegetables can be of considerable importance for their quality owing to its potential harmful effects on human health by leading to methaemoglobinaemia or forming nitrosamines and nitrosamides by reaction with amines and amides after nitrate reduction (Bianco et al., 1998; Marschner, 1997; Santamaria et al., 2002; Walker, 1990). The acceptable daily intake of nitrate was set at 03.65 mg/kg body weight by the European Communities Scientific Committee for Food in 1992. In both tests, the nitrate content of leaves was always much lower than the European legal threshold, that is 4500 mg NO3/kg (or ppm) of fresh weight for lettuce harvested from 1 October and 31 March and 3500 ppm for lettuce harvested from 1 April and 30 September (REG EC N1822/2005). In effect, this low nitrate concentration in leaves was due to the higher light radiation of Mediterranean areas, which induce the nitrate reductase activity (NRA) (Crawford, 1995). In fact, in the test 1 we found that nitrate concentration into the leaves of spring cycle was always lower than that of winter cycle, probably because the lower light intensity and temperatures of winter season determine a reduction of nitrate reductase activity within the leaves and that is according also with other authors (Burns et al., 2004; Riens and Heldt, 1992; Ysart et al., 1999).

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

81

Realty, the NRA is also strongly influenced by other factors, among those the cultivation conditions and, therefore, also the salinity. In fact, many authors (Abdel Baki et al., 2000; Flores et al., 2000; Gouia et al., 1994; Meloni et al., 2004; Rao and Gnaham, 1990) found that the nitrate reductase activity (NRA) of leaves decreases in many plants under salt stress. The primary cause of a reduction of NRA in the leaves is a specific effect associated with the presence of Cl- in the external medium. This effect of Cl- seems to be due to a reduction in NO2- uptake and consequently a lower NO3 concentration in the leaves, although a direct effect of Cl- on the activity of the enzyme cannot be discarded (Cram, 1973; Flores et al., 2000; Smith, 1973). We also found, in both cycles of two tests, that the nitrate concentration of leaves decreased at the increase of salt stress, but with a different behaviour. In fact, in the not salinized soils (test 1) we hypothesize that the effect of salt stress on nitrate content was less strong than the effect of environmental conditions on NRA and, therefore, on nitrate concentration. In fact, the control of spring cycle (irrigated with fresh water) already was almost the half of winter one, as well as the two less stressed treatments. Instead, in the salinized soils (test 2) the effect of salt stress on nitrate content prevail; in fact the greatest effect of salt stress is demonstrated by the average nitrate concentration, that was about 748 ppm in the NaCl treated leaves of test 2, while it was even 1711 ppm in the NaCl treated leaves of test 1. On the other hand, a number of laboratory and greenhouse studies have shown that salinity can reduce N accumulation in plants (Al-Rawahy et al., 1992; Cram, 1973; Feigin et al., 1991; Pessarakli, 1991; Pessarakli and Tucker, 1988). This is not surprising that an increase in Cl- uptake and accumulation is often accompanied by a decrease in shootNO3- concentration. Examples of such effect have been found in cucumber (Martinez and Cerd, 1989), eggplant (Savvas and Lenz, 1996), melon (Feigin et al., 1987) and tomato (Martinez and Cerd, 1989; Feigin et al., 1987; Kafkafi et al., 1982). Many attributed this reduction to Cl- antagonism of NO3- uptake (Bar et al., 1997; Feigin et al., 1987; Kafkafi et al., 1982) while others attributed the response to salinity effect on reduced water uptake (Lea-Cox and Syvertsen, 1993). The quality and so the nutritional value of a vegetable is also determined by the antioxidant activity (Rice-Evans, Miller, & Paganga, 1996); besides its measurement allows the evaluation of this nutritional variable without an analysis of each antioxidant compound (Pellegrini et al., 1999; Scalfi et al., 2000). The antioxidant content of plants varies considerably depending on their growing and management conditions (Lee and Kader, 2000; Zhao et al., 2006). A major part of the antioxidant activity in lettuce arises from a number of phenolic compounds, the dominant ones being caffeic acid derivatives and avonols (Hermann, 1976; Nicolle, 2004). In addition to phenolic compounds, vitamins such as vitamin C and vitamin E also contribute to the antioxidant activity in lettuce (Nicolle, 2004). It has been demonstrated that antioxidants in plants are part of a complex defense mechanism against a wide range of stresses and thus, accumulate in response to these stresses (Blokhina et al., 2003; Dixon e Paiva, 1995). In fact, due to metabolic disturbance under stress conditions, reactive oxygen species (ROS) are produced and the ROS production increases under abiotic stresses including salinity (Gomez et al., 1999; Hernandez et al., 2001; Smirnoff, 1993). The plant response to ROS is the production of antioxidant molecules, as ascorbate, glutathione, -tocopherol and carotenoids, but also of detoxifying enzymes, as superoxide dismutase, catalase and enzymes of the ascorbate-glutathione cycle. The overproduction of these compounds and enzymes, and then the antioxidant activity, is not equal in all species

82

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

and it can vary under different types of stress. In fact, in tomato grown under salt stress condition, De Pascale et al. (2001) found that the antioxidant activity increased; also Maggio et al. (2007), on tomato grown under salt and ozone stress (in open top chambers), found that the salinity determined an increase of the antioxidant activity of the carotenoids extract, while didnt observe any response to salt stress with respect to ascorbate and tocopherols contents. In our research, we found that hydrophilic and lypophilic antioxidant activity decreased at the increase of salt stress in both tests. Also, Chisari et al. (2010) found that the antioxidant capacity of baby Romaine lettuce decreased from 2.8 to 3.8 dS m-1 at the harvest. The salt could to have had both a direct effect on antioxidant activity and an indirect effect, due to the different availability of nitrogen. In fact, Flores et al. (2004) found that the red pepper, grown at high NO3 conditions, showed an increase of LAA; they related this increase to the lycopene and -carotene increases produced by NO3-treatments. Then, it is possible that the lettuce also has the same behaviour. We already said that the soil salinity caused a reduced uptake of nitrogen (low NO3 content in the leaves), more evident in the test 2. In fact, the lettuces of test 2 (grown on salinized soils) always had a lower average value of hydrophilic and lypophilic antioxidant activity than lettuces of test 1. Instead, both the HAA and the LAA were higher in the spring cycles of the two tests. It is possible that the higher light conditions of spring season respect of winter one determined a stronger oxidative stress and so the plants responded with the increase of antioxidant activity. On the other hand, also Oh et al. (2009) found that the total phenol content and the antioxidant capacity of lettuce increase in high light conditions. In effect, this behaviour could be due to the Northern origin of lettuce. We hypothesize that, during its evolution, the lettuce is adapted to grow at low light conditions, therefore a variation from this condition (high radiation) cause an abiotic stress and, specifically, an oxidative stress, to which the plant genetically responses with an increase of antioxidant activity (hydrophilic and lypophilic). Instead, the lettuce had not the need to develop similar defence mechanisms against salt stress, since the native lands of this crop are not interesting by this stress and, therefore, the lettuce has not been exposed to soil salinity during its evolution. The reduction of antioxidant activity of vitamin C also is a further confirmation of the scarce capacity of lettuce to activate the antioxidant defence mechanism under salt stress. In effect, also Dumas et al., 2003 reported that the ascorbic acid decrease to salinity. Due to the involvement of ascorbic acid in several metabolic processes, its synthesis under stress may also depend on the interaction with other factors such as nitrogen availability (De Pascale et al., 2001). Besides, the ascorbic acid is produced from carbohydrates, therefore if photosynthesis decreases, as happened for plants under saline stress (Fagnano and Quaglietta Chiarand, 2004), there are few substances for beginning the synthesis process of Vit C. In fact, the value of Vit C antioxidant activity are, however, greater in the spring cycle, that is characterized by climatic conditions (high radiation and temperatures) more favourable to photosynthesis, that was more high (data also not published).

CONCLUSIONS
Starting from the aim of the two tests, that wanted to simulate two real conditions, 1) the recent intrusion of sea water into the fresh water table and, therefore, the use of this salinized water and 2) the long term use of saline water on soil already salinized, we can report the sequent conclusions. The use of saline water always determines an increase of soil

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

83

salinity, but it is more marked (three times more) in the spring cycle, because climatic condition favour evapotraspiration and, so, the salt accumulation in the top soil. The salt stress always affect the yield, fresh weight, heads diameter, number of leaves and their expansion; still, the crop salinity tolerance decrease if the lettuce is cultivated on already salinized soil (2 hypothesis), because in these soils, critical conditions for water absorption and nutrients uptake arise. Besides, the crop response to salinity varies with the seasons: in the winter cycle the salt effect on lettuce heads and yield have been more evident. Probably, in the spring cycle, better climate conditions have mitigated the effect of salt stress; in fact the higher radiation, probably allowed to the plants to satisfy the greater energetic demands needed for correcting the ionic unbalance metabolism. Instead the salinity doesnt affect greatly the lettuce quality. In fact, the salt stress decrease the nitrate content of leaves, with a notable advantage for the farmers and, especially, for the consumers; also if the nitrate content of leaves was always low, so it confirm that the cultivation of lettuce in Mediterranean area allow to contain the problem of nitrate accumulation in the leaves. About antioxidant activity, although is undeniable that salt stress determines a general oxidative stress, still it seems that the lettuce is not able to response to this stress with an increase of antioxidant compounds, probably it is due to a specific evolution of this specie. Overall, it seems that saline water irrigation with an ECw not greater than 1.8 dS m-1 reduces the nitrate content of leaves, without to compromise excessively the yield the nutritional quality of lettuce.

ACKNOWLEDGEMENTS
We thank the Prof. Massimo Fagnano (University of Naples, Italy) for the interesting suggestions for writing this paper.

REFERENCES
1. Abdel Baki, G.K., Siefritz, F., Man, H.M., Welner, H., Kaldenhoff, R., Kaiser, W.M. (2000). Nitrate reductase in Zea mays L. under salinity. Plant Cell Environ. 23, 15521. 2. Abu-Rayyan, A., Kharawish, B.H., Al-Ismail, K. (2004). Nitrate content in lettuce (Lactuca sativa L.) heads in relation to plant spacing, nitrogen form and irrigation level. J. Sci. Food Agric. 84 (9), 931936. 3. Abushita, A.A., Hebshi, E.A., Daood, H.G., Biacs, P.A. (1997). Determination of antioxidant vitamins in

tomatoes. Food Chem. 60 (2), 207-212. 4. 5. Addiscott, T.M., Benjamin, N. (2004). Nitrate and human health. Soil Use Manag. 20 (2), 98104. Al-Rawahy, S.A., Stroehlein, J.L., Pessarakli, M. (1992). Dry-matter yield and 15N, Na+, Cl- and K+ content of tomatoes under sodium chloride stress. J. Plant Nutr. 15, 341-358. 6. Alscher, R.G., Donahue, J.L., Cramer, C.L. (1997). Reactive oxygen species and antioxidants: relationships in green cells. Physiol. Plant. 100, 224 223. 7. Altunkaya, A., Miquel Becker, E., Gkmen, V., Skibsted, L. (2009). Antioxidant activity of lettuce extract (Lactuca sativa) and synergism with added phenolic antioxidants. Food Chem., 115, 163168. 8. Anderson, J.W., Foyer, C.H., Walker, D.A. (1983). Light-dependent reduction of dehydroascorbate and uptake of exogenous ascorbate by spinach chloroplasts. Planta 158, 442-450. 9. Bahorun, T., Luximon-Ramma, A., Crozier, A., Aruoma, O. I. (2004). Total phenol, flavonoid, proanthocyanidin and vitamin C levels and antioxidant activities of Mauritian vegetables. J. Sci. Food Agric. 84, 15531561. 10. Barbieri, G., De Pascale, S., Sifola, M.I. (1994). Effetti della frequenza di irrigazione sulle funzioni di risposta produttiva della melanzana (Solanum melongena L.) alla salinit. Riv. Agron. 3, 235-246.

84

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

11. Barth, C., De Tullio, M., Conklin, P.L. (2006). The role of ascorbic acid in the control of owering time and the onset of senescence. J. Exp. Bot. 57, 16571665. 12. Baur, S., Klaiber, R. G., Koblo, A., Carle, R. (2004). Effect of different washing procedures on phenolic metabolism of shredded, packaged iceberg lettuce during storage. J. Agric. and Food Chem. 52, 70177025. 13. Bianco, V.V., Santamaria, P., Elia, A. (1998). A nutritional value and nitrate content in edible wild species used in southern Italy. Acta Hortic. 467, 7187. 14. Bie, Z., Ito, T., Shinohara, Y. (2004). Effects of sodium sulphate and sodium bicarbonate on the growth, gas exchange and mineral composition of lettuce. Sci. Hortic. 99, 215224. 15. Blokhina, O., Virolainen, E., Fagerstedt, K.V. (2003). Antioxidants, oxidative damage and oxygen deprivation stress. Ann. Bot. 91, 179194. 16. Boroujerdnia, M., Ansari, N.A., Dehcordie, F.S. (2007). Effect of cultivars, harvesting time and level of nitrogen fertilizer on nitrate and nitrite content, yield in romaine lettuce. Asian J. Plant Sci. 6 (3), 550553. 17. Bramley, P.M. (2000). Is lycopene benecial to human health? Phytochem. 54, 233236. 18. Breimer, T. (1982). Environmental factors and cultural measures affecting the nitrate content in spinach. Nutr. Cycl. Agroekosyst. 3 (3), 191292. 19. Burns, I.G., Lee, A., Escobar-Gutierrez, A.J. (2004). Nitrate accumulation in protected lettuce. Acta Hortic. 633, 271278. 20. Caldwell, C.R. (2003). Alkylperoxyl radical scavenging activity of red leaf lettuce (Lactuca sativa L.) phenolics. J. Agric. Food Chem. 51, 45894595. 21. Chen, B.M., Wang, Z.W., Li, S.X., Wang, G.X., Song, H.X., Wang, X.N. (2004). Effect of nitrate supply on plant growth, nitrate accumulation, metabolic nitrate concentration and nitrate reductase activity in three leafy vegetables. Plant Sci. 167 (3), 635643. 22. Chisari, M., Todaro, A., Barbagallo, R.N., Spagna G. (2010). Salinity effects on enzymatic browning and antioxidant capacity of fresh-cut baby Romaine lettuce (Lactuca sativa L. cv. Duende). Food Chem. 119, 1502 1506. 23. Conklin, P.L. and Barth, C. (2004). Ascorbic acid, a familiar small molecule intertwined in the response of plants to ozone, pathogens, and the onset of senescence. Plant Cell Environ. 27, 959970. 24. Cram, W.J. (1973). Internal factors regulating nitrate and chloride influx in plant cells. J. Exp. Bot., 24, 328341. 25. Crawford, N.M. (1995). Nitrate: Nutrient and signal for plant growth. Plant Cell, 7, 85968. 26. Dalton F.N., Maggio A., Piccinni G. (2000). Simulation of shoot chloride accumulation, separation of physical and biochemical processes governing plant salt tolerance. Plant Soil 219, 1-11. 27. De Pascale, S., Maggio, A., Fagnano, V., Ambrosino, P., Ritieni A. (2001). Irrigation with saline water improves carotenoids content and antioxidant activity of tomato. J. Hortic. Sci. and Biotecnol. 76, 447-453. 28. De Pascale, S., Barbieri, G. (2000). Yield and quality of carrot as affected by soil salinity from long-term irrigation with saline water. Acta Hortic. 537, 621628. 29. De Pascale, S., Barbieri, G. (1995). Effects of soil salinity from long-term irrigation with saline-sodic water on yield and quality of winter vegetable crops. Sci. Hortic. 64, 145-157. 30. de Pinto, M.C., Paradiso, A., Leonetti, P., De Gara, L. (2006). Hydrogen peroxide, nitric oxide and cytosolic ascorbate peroxidase at the crossroad between defence and cell death. Plant J. 48, 784795. 31. Dixon, R.A., Paiva, N. (1995). Stress-induced phenylpropanoid metabolism. Plant Cell 7, 10851097.

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

85

32. Drews, M., Schonhof, I., Krumbein, A. (1997). Content of minerals, vitamins, and sugars in iceberg lettuce (Lactuca sativa var. capitata L.) grown in the greenhouse dependent on cultivar and development stage. Gartenbauwissenschaft 62 (2), 6572. 33. Drews, M. (1996). Nitrate, vitamin C, and sugar content of Lactuca sativa depending on cultivar and head development. Gartenbauwissenschaft 61 (3), 122129. 34. Dumas, Y., Dadomo, M., Di Lucca, G., Grolier, P. (2003). Effects of environmental factors and agricultural techniques on antioxidant content of tomatoes. J. Sci. Food Agric. 83, 369-382. 35. Dupont, S., Mondi, Z., Willamson, G., Price, K. (2000). Effect of variety, processing, and storage on the flavonoid glycoside and composition of lettuce and chicory. J. Agric. and Food Chem., 48, 39573964. 36. Eraslan, F., Inal, A., Savasturk, O., Gunes, A. (2007). Changes in antioxidative system and membrane damage of lettuce in response to salinity and boron toxicity. Sci. Hortic. 114, 510 37. European Communities (2001). Norme di qualit delle lattughe cappucce, romane e iceberg (Lactuca sativa var. capitata, longifolia e crispa). Reg. (CE) n. 1543/01 del 27/07/01. 38. Fagnano, M. and Quaglietta Chiarand, F. (2004). Relazione tra qualit dellambiente e produzione agricola. Riv. Agron. 38, 33-84. 39. Feigin, A., Pressman, E., Imas, P., Miltau, O. (1991). Combined effects of KNO and salinity on yield and chemical composition of lettuce and chinese cabbage. Irrig. Sci. 12, 223-230. 40. Feigin A., Rylski, I., Meiri, A., Shalevet, J. (1987). Response of melon and tomato plants to chloride-nitrate ratios in saline nutrient solutions. J. Plant Nutr. 10, 1787-1794. 41. Fewtrell, L. (2004). Drinking-water nitrate, methemoglobinemia and global burden of disease: a discussion. Environ. Health Perspect. 112 (14), 13711374. 42. Flagella, Z., Cantore, V., Boari, F., Volpe, D., De Caro, A. (1999). Tolleranza allo stress salino delle specie coltivate in relazione agli aspetti fisiologici, produttivi e qualitativi. ww.inea.it-otris. 43. Flores, P., Navarro, J.M., Garrido, C., Rubio, J.S., Martnez, V. (2004). Inuence of Ca2+, K+ and NO3 fertilisation on nutritional quality of pepper. J. Sci. Food Agric. 84, 569574. 44. Flores, P., Botella, M.A., Martinez, V., Cerda, A. (2000). Ionic and osmotic effects on nitrate reductase activity in tomato seedlings. J. Plant Physiol. 156, 552557. 45. Fogliano, V., Verde, V., Randazzo, G., Ritieni, A. (1999). Method for measuring antioxidant activity and its application to monitoring the antioxidant capacity of wines. J. Agric. and Food Chem. 47, 1035-1040. 46. Foyer C.H. (1993). Ascorbic acid. In: Alscher R.G., Hess J.L. (Eds.), Antioxidants in Higher Plants. CRC Press, Boca Raton, 31-58. 47. Gao, Q. and Zhang, L. (2008). Ultraviolet-B-induced oxidative stress and antioxidant defense system responses in ascorbate-decient vtc1 mutants of Arabidopsis thaliana. J. Plant Physiol. 165, 138148. 48. Ghassemi, F., Jakeman, A.J., Nix, H.A. (1995). Salinization of land and water resources. CAB International, Wallingford. 49. Gmez, J.M., Hernndez, J.A., Jimnez, A., Del Ro, L.A., Sevilla, F. (1999). Differential response of antioxidative enzymes of chloroplasts and mitochondria to long-term NaCl stress of pea plants. Free Rad. Res. 31, S11S18 50. Gouia, H., Ghorbal, M.H., Touraine B. (1994). Effects of NaCl on flows of N and mineral ions and NO3reductase rate within whole plants of salt-sensitive bean and salt tolerant cotton. Plant Physiol., 105, 1407-1418.

86

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

51. Gueta-Dahan, Y., Yaniv, Z., Zilinskas, B.A., Ben-Hayyim, G. (1997). Salt and oxidative stress: similar and specific responses and their relation to salt tolerance in Citrus. Planta 203, 460 469. DOI: 10.1007/s004250050215. 52. Heimler, D., Isolani, L., Vignolini, P., Tombelli, S., Romani, A. (2007). Polyphenol content and antioxidant activity some species of freshly consumed salads. J. Agric. and Food Chem. 55(5), 17241729. 53. Helal, M., Mengel, K. (1981). Interaction between light intensity and NaCl salinity and their effects on growth, CO2 assimilation, and photosynthate conversion in young broad beans. Plant Physiol. 67, 999-1002. 54. Hermann, K. (1976). Flavonols and avones in food plants: a review. J. Food Technol. 11, 433448. 55. Hernndez, J. A., Jimnez, A., Mullineaux, P., Sevilia, F. (2001). Tolerance of pea (Pisum sativum L.) to longterm salt stress is associated with induction of antioxidant defences. Plant, Cell & Environ. 23, 853862. 56. Hernndez, J.A., Olmos, E., Corpas, F.J., Sevilla, F., Del Ro, L.A. (1995). Salt-induced oxidative stress in chloroplast of pea plants. Plant Sci. 105, 151 167. 57. Hernndez, J.A., Corpas, F.J., Gmez, M., Del Ro, L.A., Sevilla, F. (1993). Salt-induced oxidative stress mediated by activated oxygen species in pea leaf mitochondria. Physiol. Plant. 89, 103-110. 58. Kafkafi, U., Valoras, N., Letey, J. (1982). Chloride interaction with nitrate and phosphate nutrition in tomato (Lycopersicon esculentum L.). J. Plant Nutr. 5, 1369-85. 59. Lairon, D., Spitz, N., Termine, E., Ribaud, P., Lafont, H., Hauton, J. (1984). Effect of organic and mineral nitrogen fertilization on yield and nutritive value of butter-head lettuce. Plant Food Hum. Nutr. 34 (2), 97108. 60. Larkindale, J., Hall, J.D., Knight, M.R., Vierling, E. (2005). Heat stress phenotypes of Arabidopsis mutants implicate multiple signaling pathways in the acquisition of thermotolerance. Plant Physiol. 138, 882897. 61. Lea-Cox, J.D. and Syvertsen, J.P. (1993). Salinity reduces water use and nitrate-N-use efficiency of citrus. Ann. Bot. 72, 47-54. 62. Lee K., Kader A. (2000). Preharvest and postharvest factors influencing vitamin C content of horticultural crops. Postharvest Biol. and Technol., 20, 207220. 63. Lpez, F., Vansuyt, G., Casse-Delbart, F., Fourcroy, P. (1996). Ascorbate peroxidase activity, not the mRNA level, is enhanced in salt-stressed Raphanus sativus plants. Physiol. Plant. 97, 13 20. 64. Maas, E. V. and Hoffman, G. J. (1977). Crop salt tolerance. J. Irrig. Drain. Div. ASCE 103, 115-134. 65. Maggio, A., De Pascale, S., Fagnano, M., Barbieri, G. (2007). Can salt stress-induced physiological responses protect tomato crops from ozone damages in Mediterranean environments? Eur. J. Agron. 26, 454-461. 66. Mantovani, J.R., Ferreira, M.E., Da Cruz, M.C.P. (2005). Lettuce growth and nitrate accumulation in relation to nitrogen fertilization. Hortic. Bras. 23 (3), 758762. 67. Marschner, H. (1997). Functions of Mineral Nutrients, Macronutrients: Mineral Nutrition of Higher Plants. Academic Press, San Diego, CA, 231255. 68. Martn-Diana Ana, B., Rico, D., Barry-Ryan, C., Fras, J., Henehan Gary, T. M. (2007). Efficiency of steamer jetinjection as alternative to chlorine in fresh-cut lettuce. Postharvest Biol. and Technol. 45(1), 97107. 69. Martinez, V., Cerd, A. (1989). Influence of N source on rate of Cl, N, Na, K uptake by cucumber seedlings grown in saline conditions. J. Plant Nutr. 12, 971-983. 70. Meloni, D.A., Gulotta, M.R., Martnez, C.A., Oliva, M.A. (2004). The effects of salt stress on growth, nitrate reduction and proline and glycinebetaine accumulation in Prosopis alba. Braz. J. Plant Physiol. 16, 39-46. 71. Millar, A.A., Gardner, W.R., Goltz, J.M. (1971). Internal water status and water transport in seed onion plants. Agron. J., 63, 770-784.

Interaction between Salinity and Crop Cycle: Effect on Yield and Quality of Lettuce

87

72. Miller, N. J., Paganga, G., Wiseman S., Vannielen W., Tijburg L., Chowienczyk P., Rice-Evans C. A. (1995). Total antioxidant activity of low-density lypoproteins and the relationship with alpha-tocopherol status. Febs Letters 365, 164-166. 73. Mittler, R. & Zilinskas, B.A. (1994). Regulation of pea cytosolic ascorbate peroxidase and other antioxidant enzymes during the progression of drought stress and following recovery from drought. Plant J. 5, 397-405. 74. Mori, M., Di Mola, I., Quaglietta Chiarand, F. (2011). Salt stress and transplant time in snap bean: growth and productive behaviour. Int. J. Plant Prod. 5 (1), pp. 49-64. 75. Mori, M. Amato, M., Di Mola, I., Caputo, R., Quaglietta Chiarand, F., Di Tommaso, T. (2008). Productive behaviour of cherry-type tomato irrigated with saline water in relation to nitrogen fertilisation. Eur. J. Agron. 29/2-3 pp. 135-143. 76. Muller-Moul, P., Golan, T., Niyogi, K.K. (2004). Ascorbate-decient mutants of Arabidopsis grow in high light despite chronic photo oxidative stress. Plant Physiol. 134, 11631172. 77. Munns, R., Schachtman, D.P., Condon, A.G. (1995). The significance of a two-phase growth response to salinity in wheat and barley. Aust. J. Plant Physiol. 22, 561-569. 78. Nicolle, C., Carnat, A., Fraisse, D., Lamaison, J., Rock, E., Michel, H., Amouroux, P., Remesy, C. (2004). Characterisation and variation of antioxidant micronutrients in lettuce (Lactuca sativa folium). J. Sci. Food Agric. 84, 20612069. 79. Noctor, G. & Foyer, C. (1998). Ascorbate and glutathione: keeping active oxygen under control. Ann. Review Plant Physiol. and Plant Mol. Biol. 49, 249-279. 80. Oh, M.-M., Trick, H.N., Rajashekar, C.B. (2009). Secondary metabolism and antioxidants are involved in environmental adaptation and stress tolerance in lettuce. J. Plant Physiol. 166, 180191. 81. Pellegrini N., Re, R., Yang, M., Rice-Evans, C. (1999). Screening of dietary carotenoids and carotenoid-rich-fruit extracts for antioxidant activities applying 2,2-azinobis (3-ethylenebenzothiazoline-6-sulfonic acid) radical cation decoloration assay. Methods Enzymol. 299, 379384. 82. Pessarakli, M. (1991). Dry-matter yield, N-15 absorption, and water-uptake by green bean under sodium-chloride stress. Crop Sci. 31, 16331640. 83. Pessarkli, M., Tucker, T. C. (1988). Dry matter yield and nitrogen-15 uptake by tomatoes under sodium chloride stress. Soil Sci. Soc. Am. J. 52, 698-700. 84. Pignocchi, C. and Foyer, C.H. (2003). Apoplastic ascorbate metabolism and its role in the regulation of cell signalling. Curr. Opin. Plant Biol. 6, 379389 85. Postiglione, L., Barbieri, G., Tedeschi, A. (1995). Long-term effects of irrigation with saline water on some characteristics of a clay loam soil. Riv. Agron. 29 (1), 24-30. 86. Premuzic, Z., Garate, A., Bonilla, I. (2002). Production of lettuce under different fertilization treatments, yield and quality. Acta Hortic. 571, 6572. 87. Rao, K.R., Gnaham, A. (1990). Inhibition of nitrate and nitrate reductase activity by salinity stress in Sorghum vulgare. Phytochem. 29, 1047-1049. 88. Rautenkranz, A.A.F., Li, L., Machler, F., Martinoia, E., Oertli, J.J. (1994). Transport of ascorbic and dehydroascorbic acids across protoplast and vacuole membranes isolated from barley (Hordeum vulgare L.cv Gerbel) leaves. Plant Physiol. 106, 187-193. 89. Re, R., Pellegrini, N., Proteggente, A., Pannala, A., Yang, M., Rice-Evans, C. (1999). Antioxidant activity applying an improved ABTS radical cation decolorization assay. Free Radic. Biol. Med. 26, 12311237.

88

Ida Di Mola, Mauro Mori & Fabrizio Quaglietta Chiaranda

90. Rhoades, J.D., Kandiah, A., Mashali, A.M. (1992). The use of saline waters for crop production. Irrig. and Drain. Paper No. 48. FAO, Rome. 91. Rice-Evans, C. A., Miller, J. N., & Paganga, G. (1996). Structure-antioxidant activity relationships of avonoids and phenolic acids. Free Radic. Biol. and Med. 20, 933956. 92. Riens, B., Heldt, H.W. (1992). Decrease of nitrate reductase activity in spinach leaves during a lightdark transition. Plant Physiol. 98 (2), 573577. 93. Santamaria, P., Elia, A., Parente, A., Serio, F. (2002). Effect of solution nitrogen concentration on yield, leaf element content, and water and nitrogen use efficiency of three hydroponically grown rocket salad genotypes. J. Plant Nutr. 25, 245258. 94. Savvas, D., Lenz, F. (1996). Influence of NaCl concentration in the nutrient solution on mineral composition of eggplants grown in sand culture. Angewandte Botanik 70, 124-127. 95. Scal, L., Fogliano, V., Pentangelo, A., Graziani, G., Giordano, I., & Ritieni, A. (2000). Antioxidant activity and general fruit characteristics in dierent ecotypes of Corbarini small tomatoes. J. Agric. and Food Chem. 48, 1363 1366. 96. Sies, H. (1991). Oxidative stress: Oxidant and antioxidant. Academic press, London. 97. Siomos, S.A., Papadopoulou, P.P., Niklis, D.N., Dogras, C.C. (2002). Quality of romaine and leaf lettuce at harvest and during storage. Acta Hortic. 579, 641646. 98. Smirnoff, N. (1996). The function and metabolism of ascorbic acid in plants. Ann. Bot. 78, 661-669. 99. Smirnoff, N. (1993). The role of active oxygen in the response of plant to water decit and desiccation. New Phytol. 125, 2758. 100. 100) Smith, F.A. (1973). The internal control of nitrate uptake into excised barley roots with differing salt contents. New Phytol. 72, 769782. 101. Sood Mand Malhotra, S.R. (2001). Effects of processing and cooking on ascorbic acid content of chickpea (Cicer arietinum L.) varieties. J. Sci. Food Agric. 82, 6568. 102. Sudhakar, C., Lakshmi, A., Giridarakumar, S. (2001). Changes in the antioxidant enzyme efficacy in two high yielding genotypes of mulberry (Morus alba L.) under NaCl salinity. Plant Sci. 141, 613619. 103. Turner, N.C. (1974). Stomatal response to light and water under filed conditions. Mechanisms of regulation of plant growth. Royal Soc. N.S. Bull. 12, 423-432. 104. Van Camp, W., Capiau, K., Van Montagu, M., Inz, D., Slooten, L. (1996). Enhancement of oxidative stress tolerance in transgenic tobacco plants overproducing Fe-superoxide dismutase in chloroplasts. Plant Physiol. 112, 1703-1714. 105. Van Hoorn, J.W., Katerji, N., Hamdy, A., Mastrorilli, M. (1993). Effect of saline water on soil salinity and on water stress, growth and yield of wheat and potatoes. Agric. Water Manag. 23, 247-265. 106. Walker, R. (1990). Nitrates, nitrites and N-nitroso compounds: a review of the occurrence in food and diet and the toxicological implications. Food Addit. Contam. 7, 717768. 107. Yeo, A. (1998). Predicting the interaction between the effects of salinity and climate change on crop plants. Sci. Hortic. 78, 159-174. 108. Ysart, G., Clifford, R., Harrison, N. (1999). Monitoring for nitrate in UK grown lettuce and spinach. Food Addit. Contam. 16 (7), 301306. 109. Zhao, X., Carey, E.E., Wang, W., Rajashekar, C.B. (2006). Does organic production enhance phytochemical content of vegetables and fruit? Current knowledge and prospects for research. Hort. Technol. 16, 18.