Scientia Horticulturae 111 (2007) 319325

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Effect of organic and inorganic fertilizers applied during successive

crop seasons on growth and nitrate accumulation in lettuce
Georgios C. Pavlou a,*, Constantinos D. Ehaliotis b, Victor A. Kavvadias a
a

National Agricultural Research Foundation (N.AG.RE.F.), Olive and Horticultural Crops Institute, Lakonikis 87, Kalamata 24100, Greece
b
Agricultural University of Athens, Soil Science and Agricultural Chemistry Lab., Iera Odos 75, Athens 11855, Greece
Received 23 December 2005; received in revised form 10 August 2006; accepted 6 November 2006

Abstract
A romaine-type lettuce (Lactuca sativa L.) cv. Corsica was cultivated during three successive crop seasons (late-spring, late-autumn and latewinter) in the same soil of an experimental greenhouse in S.W. Peloponnese, Greece. Seven long-term fertilization treatments were tested for their
effect on plant growth and nitrate concentration in the external lettuce leaves. Treatments included: three different doses of organic fertilization
(composted sheep manure) applied at the start of each crop season, three different doses of inorganic N fertilization applied via fertigation during
each crop season, and a control treatment in which no fertilizer was applied. A drip irrigation system was used to water all plants. The highest
nitrate levels were observed in the medium and maximum inorganic fertilization treatments (572664 mg kg1) in all crop seasons. They were
significantly higher compared to the respective organic fertilization treatments (253435 mg kg1) and all other fertilization treatments (148
435 mg kg1). Crop season affected lettuce growth more than nitrate accumulation in the lettuce leaves: lettuce biomass production was the
smallest and most uniform in the late-autumn season and did not respond to the fertilization treatments tested (ranging from 409 to 439 g plant1),
while in the late-spring season biomass production was the highest and most variable (561841 g plant1), it correlated with nitrate concentration
in the leaves and in the medium and maximum inorganic fertilizer doses it significantly exceeded production from all other fertilization treatments
(827841 g plant1). Following the three crop seasons the residual availability of N, P and K was clearly enhanced in the soil receiving the organic
compared to the inorganic fertilization. Nitrate concentration in lettuce leaves was far below the upper limits set by the European Commission in all
fertilization treatments throughout the three crop seasons, a result attributed mainly to the sufficient level of light intensity and duration throughout
the year in Southern Greece.
# 2006 Elsevier B.V. All rights reserved.
Keywords: Lettuce; Lactuca sativa L.; Fertilization; Organic fertilizers; Manure; Nitrates

1. Introduction
In the temperate zone regions lettuce is cultivated
throughout the year, but production and quality characteristics
benefit from cool weather and high light intensity. The
concentration of nitrates in the edible leaves of lettuce are
regulated by the European Commission Regulation No 563/
2002 which has set upper limits in order to protect consumers
from potential toxicological risks following the consumption of
nitrate-rich foods (Maynard et al., 1976; Walker, 1990;
Bruning-Fann and Kaneene, 1993). Recent studies however,
report on beneficial effects of nitrates mainly related to the

* Corresponding author. Tel.: +30 27210 29812; fax: +30 27210 27133.
E-mail address: gpavlou.kal@nagref.gr (G.C. Pavlou).
0304-4238/$ see front matter # 2006 Elsevier B.V. All rights reserved.
doi:10.1016/j.scienta.2006.11.003

control of the gut flora (Lundberg et al., 2004; Addiscott and

Benjamin, 2004).
Accumulation of nitrates results from an imbalance between
the uptake and translocation of nitrates by the xylem, and the
reduction of these nitrates to ammonia which is subsequently
rapidly incorporated into amino acids (Maynard et al., 1976).
However, the internal nitrate concentration in the plant seems to
be controlled by a self-regulatory mechanism exerted either by
negative feedback control on the net nitrate uptake rate
(Cardenas-Navarro et al., 1998) or by passive control on nitrate
efflux (Scaife, 1989). The viewpoint that non-structural
carbohydrates and nitrates have a complementary role in
maintaining cell tugor (Blom-Zandstra and Lampe, 1985; Behr
and Wiebe, 1988) offers a credible model for the plant nitrate
regulation mechanism suggesting the accumulation of nitrates
in the vacuole as an alternative osmoticum under low radiation

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G.C. Pavlou et al. / Scientia Horticulturae 111 (2007) 319325

conditions (Blom-Zandstra, 1989; Buwalda and Warmenhoven,

1999).
Accumulation of nitrates in lettuce has been shown to be
affected by the soil texture and the source of fertilizer-N (Scaife
et al., 1986; Gianquinto et al., 1992; Gunes et al., 1995), the
NH4-N-to-NO3-N fertilizer-N ratio (McCall and Willumsen,
1998; Demsar and Osvald, 2003), the timing of fertilizer-N
release (Tesi and Lenzi, 1998), the light intensity and duration
(Behr and Wiebe, 1992; Chadjaa et al., 1999; Drews et al.,
1995; Gaudreau et al., 1995), crop season (Gianquinto et al.,
1992), and lettuce type and cultivar (Blom-Zandstra and
Eenink, 1986; Siomos, 2000; Escobar-Gutierrez et al., 2002). In
most types of lettuce, including the romaine type, the highest
concentration of nitrates is normally observed in the external
leaves (Corre and Breimer, 1979; Santamaria et al., 1999; AbuRayyan et al., 2004).
In countries of the Mediterranean basin, sheep manures are
traditionally used as an organic fertilization source. Their
recycling into soils low in organic matter, which are widespread
in these regions, could benefit soil structure and long term
fertility, and is also an alternative to inorganic fertilizers in the
growing organic vegetable production business.
In this work we compare short and long term effects of
organic versus inorganic sources of N on the growth and the
accumulation of nitrates in the external leaves of a romaine
lettuce. Composted sheep manure is used for organic
fertilization whereas standard fertigation applied by farmers
in the area is used for the comparison to inorganic fertilization.
Fertilization from both sources is applied at a minimum, a
medium and a maximum dose during three successive crop
seasons in the same soil. Effects of crop season and long term
cumulative fertilization on plant and soil are studied.
2. Materials and methods
2.1. Location and design of experiments
The experiments were carried out during three successive
crop seasons: late spring (first, 6 April17 May 2000), late

autumn (second, 26 October5 December 2000), and latewinter (third, 6 February28 March 2001), at the N.AG.RE.F.Olive and Horticultural Crops Institute of Kalamata, S.W.
Peloponnese, Greece (378030 N, 228070 E, altitude 9 m). At the
start of each crop season, a romaine lettuce crop cv. Corsica
(Royal Sluis) was established in a polyethylene covered
greenhouse (8085% transparency to visible irradiation), with
60% side wall cover. The greenhouse soil was a sandy clay
loam (SCL) with chemical characteristics presented in
Table 1.
Seven long-term fertilization treatments were compared
including: (i) three different organic fertilization treatments
(O-min, O-mid, O-max) consisting of composted sheep
manure (Table 1) applied at the start of each cultivation
period, at doses of 13 l plant1 respectively for the first
and second crop seasons, and reduced by 33% for the third
crop season (0.67, 1.34, and 2.01 l plant1, respectively), (ii)
three different treatments of inorganic-N fertilization (Imin, I-mid, I-max) applied via drip irrigation during each
crop season at doses corresponding to 87, 174 and
261 mg N plant1 fertigation1, respectively (Table 2),
and (iii) a control treatment in which no fertilizer was
applied (C). The nutrient elements provided in total during
each crop season by the organic and inorganic fertilization
treatments are shown in Table 3. In all three inorganic
fertilization treatments (I-min, I-mid, I-max) the soluble
commercial fertilizers 153015 and 202020 plus micronutrients were applied, each at a rate of 0.25 g plant1
fertigation1. Extra N fertilization was applied for the I-mid
and I-max treatments with the inorganic fertilizer 34.500
(NH4NO3) at rates of 0.25 and 0.5 g plant1 fertigation1,
respectively.
Each experimental plot consisted of 54 lettuce plants
placed in three rows (30 cm apart) of 18 plants (25 cm
apart). The plants of the two outer rows and the three plants
in each end of the middle row were kept as guard plants.
There were five replicate plots per treatment and the
experiment was set up as a randomized complete block
design.

Table 1
Chemical characteristics of soil and sheep manure used in the experiments

Soil
Manurea
a

E.C.
(mS cm1)

pH

CaCO3 (%)

O.M. (%)

C.E.C.
(cmol(+) kg1)

NKjeldahl
(g kg1)

POlsen
(mg kg1)

Kexch
(mg kg1)

2.6
21.7

7.4
8.0

13.4
8.4

1.4
72.0

8.6
36.0

0.15
1.64

37.0
1.4 (% d.w.)

118.2
1.8 (% d.w.)

Manure dry weight = 201 g l1.

Table 2
Nutrient elements provided per fertigation by the inorganic fertilization treatments applied to lettuce
Fertilization
treatment

I-max
I-mid
I-min

Nutrient elements provided in each crop season

N (g plant1
fertigation1)

P (g plant1
fertigation1)

K (g plant1
fertigation1)

Mg (mg plant1
fertigation1)

Fe (mg plant1
fertigation1)

Mn (mg plant1
fertigation1)

Zn (mg plant1
fertigation1)

Cu

0.261
0.174
0.087

0.027
0.027
0.027

0.036
0.036
0.036

0
0
0

0.195
0.195
0.195

0.081
0.081
0.081

0.0163
0.0163
0.0163

0.0163
0.0163
0.0163

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321

Table 3
Nutrient elements provided per crop season by the fertilization treatments applied to lettuce
Crop season

Fertilization treatment

Macro-elements (g plant1)

Micro-elements (mg plant1)

Mg

Fe

Mn

Zn

Cu

First and second

O-max
O-mid
O-min

9.840
6.560
3.280

0.258
0.172
0.086

4.692
3.128
1.564

0.102
0.068
0.034

13.53
9.02
4.51

13.11
8.74
4.37

6.69
4.46
2.23

4.71
3.14
1.57

Third

O-max
O-mid
O-min

6.593
4.395
2.198

0.173
0.115
0.058

3.144
2.096
1.048

0.068
0.046
0.023

9.07
6.04
3.02

8.78
5.86
2.93

4.48
2.99
1.49

3.16
2.10
1.05

First

I-max
I-mid
I-min

3.380
2.260
1.140

0.356
0.356
0.356

0.473
0.473
0.473

0
0
0

2.54
2.54
2.54

1.06
1.06
1.06

0.21
0.21
0.21

0.20
0.20
0.20

Second

I-max
I-mid
I-min

2.080
1.390
0.700

0.218
0.218
0.218

0.291
0.291
0.291

0
0
0

1.56
1.56
1.56

0.63
0.63
0.63

0.13
0.13
0.13

0.13
0.13
0.13

Third

I-max
I-mid
I-min

2.860
1.910
0.960

0.301
0.301
0.301

0.398
0.398
0.398

0
0
0

2.15
2.15
2.15

0.90
0.90
0.90

0.18
0.18
0.18

0.18
0.18
0.18

2.2. Cultivation and irrigation practices

The outline of practices and climatic conditions are
presented in Table 4. Lettuce seedlings were pre-grown in
190 ml plastic pots filled with the organic substrate Terra
Nature (Holland). They were transplanted in the greenhouse at
the five to seven true-leaf stage. Organic fertilizer (sheep
manure) had already been incorporated into the soil of the
respective plots down to a depth of about 20 cm, 23 days
before transplantation using a milling machine.
Inorganic fertilizers were dissolved in 30 l water tanks and
injected by an electric pump at a pressure slightly greater than
1.5 atm separately in each of the three 30 m long primary pipes,
from where the plants of the three inorganic fertilization
treatments were fertigated by 5 m long secondary dripperpipes. Plants of control and organic fertilization treatments
were irrigated through separate pipes with water only. Every
plant was irrigated by a labyrinth type dripper (Netafim)
supplying about 3 l h1 at a pressure of 1 atm.
The irrigation water used had the following characteristics:
electrical conductivity 686 mS/cm (at 25 8C), pH 7.29, S.A.R.
2.7, HCO3 409 mg l1, Ca2+ 52.1 mg l1 and Mg2+
12.2 mg l1.

Until around 10 days after transplanting, plants of all

treatments were irrigated with water only (0.2
0.4 l plant1 irrigation1). Afterwards, when plants were well
established in the soil, fertigation was applied with water
quantities of 0.50.6 l plant1 fertigation1. The fertilizer
concentration in the irrigation water ranged between 0.5%
and 1%.
When plants had reached approximately their highest
commercial size they were harvested by cutting the stems at
the soil level. Twelve plants from each plot were harvested soon
after sunrise on a typical for each season day.
2.3. Analytical methods
Just after harvest the fresh weight (f.w.) of the plants was
determined; then four external leaves from each plant were
removed and their f.w. was also determined. The leaves were
then washed off with tap water and blotted dry on absorbing
paper. Their dry weight (d.w.) was subsequently determined
following drying in a drying chamber at 70 8C for 6070 h. Dry
leaves were ground to 40-mesh size by a centrifugal mill
(TecatorSweden). Homogenized leaf powder was reinserted
to the drying chamber at 70 8C for about 24 h, and 0.225 g were

Table 4
Cultivation and irrigation practices and climatic conditions during the three successive crop seasons: late-spring (first), late-autumn (second) and late-winter (third)

Lettuce transplanting
Date of harvesting
Crop duration (days)
Irrigation water (l plant1) total
Total number of irrigations
Number of fertigations
Sunshine duration (h) total
Average photoperiod (h)
Average air min temperature (8C)
Average air max temperature (8C)

First crop season

Second crop season

Third crop season

6 April 2000
17 May 2000
41
12.2
26
13
354
13.53
13.3
32.7

26 October 2000
5 December 2000
39
6.9
12
8
229
10.27
11.8
29.5

6 February 2001
28 March 2001
50
7.7
17
11
329
11.47
8.8
27.7

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G.C. Pavlou et al. / Scientia Horticulturae 111 (2007) 319325

Table 5
Yield of lettuce (fresh weight) grown under different organic (O) or inorganic (I) fertilization schemes during three successive crop seasons: late-spring (first), lateautumn (second) and late-winter (third)
Fertilization treatment

First crop season (g plant1)

Second crop season (g plant1)

Third crop season (g plant1)

I-max
I-mid
I-min
O-max
O-mid
O-min
Control

841
827
689
701
670
561
541

439
445
441
443
432
409
335

625
624
534
569
536
504
262

a
a
b
b
bc
cd
d

a
a
a
a
a
a
b

a
a
ab
ab
ab
b
c

Means in the same column (crop season) followed by different letters denote significant differences according to Duncans multiple range test ( p = 0.05).

used for colorimetric determination of nitrate-N (NO3-N) at

410 nm by a Hitachi U-2001 spectrophotometer following the
nitration by salycilic acid method described by Cataldo et al.
(1975). Five replicate subsamples were measured to obtain each
plot-sample value. Randomly selected samples were also crosschecked for NO3-N content with the copperized-Cd reduction
method (Keeney and Nelson, 1982) to confirm methodological
accuracy.
Length and width of the biggest leaf on each of the 12 plants
under harvest were measured the day before harvest.
Air temperature at plants height was recorded by a
Lambrecht thermograph, while sunshine duration was obtained
from the Kalamata Meteorological Station of the National
Meteorological Service. Photoperiod was calculated using the
approximate local sunrise and sunset hours.
Soil available P was estimated by extraction with NaHCO3
(Olsen-P) and measured colorimetrically at 880 nm (Hitachi U2001) following the ascorbic acid molybdate reduction method
(Olsen and Sommers, 1982). Soil available K (exchangeable
plus water soluble) was estimated following extraction with
NH4OAc (Thomas, 1982) and determined by flame photometry
(Coring 410). The anaerobic incubation index of soil-N
availability was estimated by measuring the NH4-N production
from soil samples incubated anaerobically for 1 week at 40 8C
according to Keeney (1982).
2.4. Statistical analysis
Analysis of variance was performed to evaluate differences
in leaf length, leaf width, plant fresh weight, and nitrate

concentration in the external leaves of lettuce. Treatments were

thereafter compared by Duncans multiple range test
( p = 0.05). Simple linear regression was also applied using
the SPSS v.10 for Windows OS.
3. Results
3.1. Lettuce plant growth
During the first crop season (late-spring) characterized by
the greatest temperatures, photoperiod, and sunshine duration
(Table 4) high yields were observed in all treatments (Table 5).
Lettuce biomass yield in the maximum and medium inorganic
fertilization treatments (I-max, I-mid) was significantly greater
than in all other treatments (Table 5). The respective organic
fertilization treatments (O-max, O-mid) did not differ
significantly from the minimum inorganic fertilization treatment (I-min) whereas the control resulted in the lowest yield
which was similar to the minimum organic fertilization (O-min)
treatment only.
During the second crop season (late-autumn), characterized
by intermediate temperatures and the shortest photoperiod and
sunshine duration (Table 4), reduced and uniform yields were
observed among treatments (Table 5). Similar lettuce biomass
yield was obtained in all treatments apart from the significantly
smaller yield of the control (Table 5).
During the third crop season (late-winter) characterized by
the lowest temperatures and intermediate photoperiod and
sunshine duration (Table 4) plant yields were also increased,
but did not reach the yields observed in the first (late-spring)

Table 6
Maximum leaf length and width of lettuce plants grown under different organic (O) or inorganic (I) fertilization schemes during three successive crop seasons: latespring (first), late-autumn (second) and late-winter (third)
Fertilization treatment

First crop season

Length (cm)

Width (cm)

Length (cm)

Width (cm)

Length (cm)

Width (cm)

I-max
I-mid
I-min
O-max
O-mid
O-min
Control

32.92
32.33
30.41
30.65
30.16
29.24
29.00

21.85
21.23
19.42
19.77
19.10
18.56
18.12

31.49
32.10
31.53
31.87
31.17
30.92
29.44

16.63
17.48
16.94
17.18
16.30
15.83
14.15

32.30
32.28
31.02
31.94
31.22
30.90
28.14

19.40
19.35
17.69
18.59
17.82
17.28
12.72

a
a
b
b
bc
cd
d

Second crop season

a
a
b
b
bc
cd
d

a
a
a
a
a
a
b

Third crop season

abc
a
ab
ab
bc
c
d

a
a
ab
ab
ab
b
c

a
a
bc
ab
bc
c
d

Means in the same column (length or width) followed by different letters denote significant differences according to Duncans multiple range test ( p = 0.05).

G.C. Pavlou et al. / Scientia Horticulturae 111 (2007) 319325

323

Table 7
Nitrate concentration, on a fresh weight basis, in the external leaves of lettuce plants grown under different organic (O) or inorganic (I) fertilization during three
successive crop seasons: late-spring (first), late-autumn (second) and late-winter (third)
Fertilization treatment

First crop season (mg kg1)

Second crop season (mg kg1)

Third crop season (mg kg1)

I-max
I-mid
I-min
O-max
O-mid
O-min
Control

623 a
572 a
314 b
311 b
253 bc
186 c
71 d

664 a
588 a
363 bc
435 b
412 b
282 c
92 d

662 a
623 a
372 b
295 b
277 b
148 c
25 d

Means in the same column (crop season) followed by different letters denote significant differences according to Duncans multiple range test ( p = 0.05).

season (Table 5). Similar lettuce biomass yield was obtained in

all treatments apart from the minimum organic fertilization (Omin) treatment which resulted in significantly lower yield
compared to the maximum and medium inorganic fertilization
treatments (I-max, I-mid) but greater yield compared to the
control (Table 5).
The width and length of the largest leaf in each lettuce plant
were measured, as indicators of plant growth. Statistical
differences in leaf length among treatments (Table 6) were
identical to the lettuce biomass differences (Table 5).
Differences in leaf width were analogous to the lettuce biomass
differences for the first crop season, but small differentiations
were observed in the following seasons (Table 6).
3.2. Nitrate accumulation in leaves
Nitrate concentration in the external leaves did not exceed
the 664 mg kg1 f.w. (Table 7) in all three cultivation periods,
even under the maximum fertilization treatments. Similar levels
and patterns of nitrate accumulation in the external leaves were
observed in all three crop seasons (Table 7). A remarkable
increase in nitrate accumulation was observed in the organic
fertilization treatments during the late autumn period,
characterized by short photoperiod and sunshine duration
(Table 4), but specific experimentation is needed to confirm this
trend. In all three crop seasons nitrate accumulation in the
external leaves of lettuce plants was significantly greater in the

maximum and medium inorganic fertilization treatments (Imax, I-mid) (Table 7); the two respective organic fertilization
treatments (O-max, O-mid) followed, together with the
minimum inorganic fertilization treatment (I-min). The
concentration of nitrates was even lower in the minimum
organic fertilization treatment (O-min) except for the O-mid
treatment in the first crop season and the I-min treatment in the
second crop season (Table 7). Finally the control (nonfertilized) treatment showed the lowest nitrate concentration
compared to all fertilization treatments in each season
(Table 7).
The same nitrate accumulation patterns and treatment
differences were obtained by expressing nitrate concentration
on a dry weight basis and the data were highly correlated to the
per fresh weight data (r2 = 0.892, p < 0.01) apparently due to
the small variability of the d.w.-to-f.w. ratio at harvest time,
indicating that both expressions were valid for analyzing nitrate
accumulation data.
3.3. Residual fertilization effects
Following the three successive crop seasons the residual
availability of N, P and K in the soil was estimated. Residual N,
P and K availability in the soil was generally higher and tended
to increase with organic fertilization treatment dose (O-min, Omid, O-max) compared to control (non-fertilized) plots, but did
not respond to inorganic fertilization dose (Fig. 1AC,

Fig. 1. Residual effect of three different doses of inorganic fertilization (I-max, I-mid, I-min) and organic fertilization (O-max, O-mid, O-min) applied to lettuce
plants in three successive crop seasons on the soil-N availability (A), on the soil-P availability (B) and on the soil-K availability (C). Bars with different letters denote
significant differences according to Duncans multiple range test, p = 0.05.

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G.C. Pavlou et al. / Scientia Horticulturae 111 (2007) 319325

respectively). Specifically, the O-max treatment resulted in a

significantly greater index of soil-N availability compared to all
the inorganic fertilization treatments (Fig. 1A), the treatments
O-mid and O-max resulted in a significantly higher P
availability compared to all the inorganic fertilization treatments (Fig. 1B), whereas all the organic fertilization treatments
significantly exceeded the exchangeable-K values observed in
the inorganic fertilization treatments (Fig. 1C).
4. Discussion
Comparisons between inorganic and organic fertilizer-N
sources are hard to perform since there is usually a dramatic
difference in N availability from these two sources of N.
Comparisons on the basis of similar amounts of total N applied
are therefore of limited relevance to agricultural practice,
whereas comparisons on the basis of similar N availability are
hindered by the lack of reliable nitrogen release estimates for
organic fertilizer sources derived from animal manures (Van
Kessel and Reeves, 2002). A general rule of thump assuming
35% of total-N mineralization in the first year following manure
application to soil is occasionally used (Klausner, 1997; Van
Kessel and Reeves, 2002), but deviations are large and manureN mineralization may also significantly depend on soil
temperatures and moisture (Agehara and Warncke, 2005).
Our main objective, however, was to compare nitrate
accumulation in lettuce cultivated in different seasons under
typical organic and inorganic N fertilization regimes. A
standard fertilization by inorganic fertilizers or sheep manure
used in the cultivation area was therefore chosen as the standard
fertilization treatment, increased by 50% in N content to make
the maximum dose treatment and decreased by 50% in N
content to make the minimum dose treatment. As sheep manure
had a standard elemental synthesis, all other nutrients applied
varied in the same order (+50% and 50% for the O-min and
O-max, respectively); however, treatment effects on lettuce
growth should be attributed mainly to N availability since other
nutrients in soil were adequately available and no deficiencies
were observed in randomly checked plants (data not presented).
General changes in lettuce growth between crop seasons
were in line with changes in photoperiod and sunshine duration.
This is expected, since photoperiod and sunshine duration
differed widely, whereas minimum temperatures differed only
slightly and were near optimum during all seasons. High lettuce
plant biomass production occurred mainly in the late-spring
season associated with long sunshine duration, which is in line
with the positive effects that have been observed on the biomass
of lettuce by supplementary light (McCall and Willumsen,
1999) and long photoperiod (Chadjaa et al., 1999). Fertilizer
effects on growth were most clearly differentiated in this spring
season; on the contrary a poor response of plants to either
organic or inorganic N fertilization was observed in the late
autumn season indicating that suboptimal environmental
conditions (short photoperiod and sunshine duration) limited
plant growth independent of N availability.
Seasonality affected plant growth more than nitrate
concentration in leaves. Apparently, Mediterranean climatic

conditions led to adequate photosynthetic activity and nitrate

reduction rates in planta during all seasons and therefore the
accumulation of nitrates in the lettuce leaves were far bellow
the maximum limits set by the E.C. 563/2002 Regulation for
lettuce fresh product (3500 and 4500 mg kg1 for the
cultivation periods April-to-September and October-to-March,
respectively). Indeed raised nitrate reductase activity has been
observed at increased light levels (Gaudreau et al., 1995) and a
close negative correlation between photosynthetic activity and
nitrate content has been reported for various lettuce cultivars
(Behr and Wiebe, 1992) and was used for modeling plant nitrate
accumulation (Scaife and Schloemer, 1994).
Nitrate accumulation in lettuce leaves was roughly doubled
in all crop seasons in the maximum and medium inorganic
fertilization treatments compared to the following treatments
(maximum and medium organic, minimum inorganic). This is
despite the higher total N in the respective sheep manure
treatments and is apparently explained by the higher N
availability in the inorganic fertilizer applications. However, a
significant growth advantage was obtained for those two
inorganic fertilization treatments in the late-spring crop season
only, with the highest sunlight intensity and duration (20% and
23% yield increase compared to the respective maximum and
medium organic fertilization treatments). Sheep manure or lowdose inorganic N fertilizer applications may therefore be
successfully used under suboptimal environmental conditions
to avoid unnecessary costs and nitrate accumulation in lettuce
leaves.
Reduced nitrate accumulation results have been reported for
farm yard manure fertilized lettuce for one crop season only
(Gianquinto et al., 1992; Stopes et al., 1989). Our results show
that composted sheep manure applications were particularly
safe in terms of nitrate accumulation in lettuce leaves in the
long-term. They remained significantly lower than in the
respective inorganic fertilization treatments, even after subsequent high level applications in different cultivation seasons
(for example, I-max caused 100%, 53% and 124% increase in
nitrate concentration compared to O-max in the respective first,
second and third crop season, but a significant yield benefit was
observed in the first season only). Moreover, sheep manure
applications contributed to the built up of soil fertility at the end
of the experiment, since they resulted in significantly increased
N, P and K availability, contrary to the inorganic fertilization
treatments that showed residual fertilizer effects in soil not
different from the control.
The effects of fertilization treatments on lettuce yield were
similar to their effects on lettuce growth indicators (largest leaf
length and width). However, leaf length showed limited
variability between treatments (2833 cm) and was not a
sensitive indicator of lettuce growth (r2 = 0.218, p = 0.03); on
the contrary leaf width varied widely (1322 cm) and was
strongly correlated to lettuce growth (r2 = 0.938, p < 0.01).
5. Conclusions
Nitrate concentration in romaine lettuce leaves depends on
fertilizer type and dose rather than on the crop season and may

G.C. Pavlou et al. / Scientia Horticulturae 111 (2007) 319325

remain in Southern Greece far below the upper limits set by

the European Commission Regulation, even under high and
repeated fertilization treatments, a result attributed mainly to
the sufficient sunlight intensity and duration throughout the
year in this region. High application rates of sheep manure
may be needed to reach the lettuce growth obtained by
inorganic fertilizers under optimal climatic conditions only.
Increased inorganic fertilizer doses should be avoided since,
especially under suboptimal light conditions, they might
result in significant rise in nitrate accumulation in leaves but
marginal yield increase; on the contrary composted sheep
manure may be safely applied even at high doses. High
residual availability of N, P and K may be obtained in the soil,
following the sheep manure applications, which relates to
manure dose; on the contrary no residual availability of N, P
and K was obtained by inorganic fertilization applied via
fertigation.
Acknowledgement
This work was financially supported by the National
Agricultural Research Foundation.
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