Effects of fertigation with purified urban wastewater on soil and pepper plant ( Capsicum annuum L . ) production , fruit quality and pollutant contents

The effects, in greenhouse conditions, of Purified Urban Wastewater (PW) from Almería (Spain), in the fertigation of pepper (Capsicum annuum L.) on sandy mulch soil, were evaluated. Primary, secondary (active sludges) and tertiary (Chlorination + ozonation) purification treatments were applied to wastewater. Irrigation treatments applied were PW, natural Ground Water (GW), Fertilizer PW (FPW) and Fertilizer GW (FGW). The vegetal biomass, yield and fruit quality were controlled. Heavy metals (Cr, Cd, Pb, Ni, Mn, Cu and Zn), arsenic (As) and Polycyclic Aromatic Hydrocarbons (PAH) in water, soil, leaf, and fruit were analysed. The PW presented heavy metal, As and PAH contents acceptable for its use in drip irrigation. In the soil, fertigated with PW, the concentration of heavy metals and As did not increase, whilst the PAH concentration decreased. The PW treatment supplied enough nutrients to obtain yield and fruit quality equal to that of GW with fertilization. A significant saving on N, P and K fertilizers (37%, 66% and 12% respectively) was achieved by using PW. The Cd, Pb and As contents of the fruit did not show risk for human consumption. The total PAH concentrations in the fruit were low, the highest of which was phenathrene, with no carcinogenic signification. Additional key words: fertilizer savings; food safety; heavy metals; irrigation; polycyclic aromatic hydrocarbons (PAH); treated wastewater.


Introduction
The search for new water supplies in the Mediterranean basin area is constant, due to its irrigable terrain: although it only represents 30% of cultivated land, it produces 75% of agricultural output and in many of these areas agriculture would not be possible without irrigation (Papadopoulos, 1995).Due to scarcity of available irrigation water, the use of urban wastewater has been implemented and used in Mediterranean countries (Massoud et al., 2003).In 2004, Spain reused 1 hm 3 day -1 of purified wastewater, which is estimated to be 7% (INE, 2008).
The use of Purified Wastewater for agricultural usage continues to expand due to the benefits it offers such as: a solution to irrigation water scarcity; the availability of large amounts throughout the year; the possibility to reserve better quality water for human consumption; the reduction of fertilizers needed due to the nutrients contained in this type of water; protection of the environment; the reduction of effluent waters in the surrounding area; an increase in the returns on investment made for the water's purification; regeneration of wet lands; protection of the water quality in waterbearing terrain by refilling the aquifers; avoiding marine intrusion in coastal areas and overexploitation.However, inadequate handling of fertilisation and irrigation with these types of water could supply the crop with higher quantities of nutrients and produce excessive accumulations within the plant and soil, negatively affecting the yield and production quality.
The main problems caused by the use of wastewater result from the presence of biological and chemical contaminants, most importantly those that have not been treated.These could harm the agricultural environment, as well as the health of farmers and consumers as they could cause a build-up of chemical contaminants in the soil, cause the mobilisation of contaminants from the soil to the crop due to cultivation, lead to soil salinization and cause diseases for both the farmers, who are in direct contact with the water, and for consumers if the crops have been colonised by pathogenic micro-organisms (Khan et al., 2008;Klay et al., 2010).Inorganic chemical contamination is basically due to heavy metals, As and Na.The concern over these elements is due to the fact that they are not biodegradable.They are absorbed by the crops and they can easily accumulate in different parts of the human body, even if they are present in low concentrations, as the body has no effective elimination mechanism (Arora et al., 2008).
Organic contaminants that appear in urban wastewater are from diverse origins.The majority are found in the remnants of soaps, detergents, general cleaning products, pesticide residues and organic material in the stages of decomposition.There are certain groups of contaminants that, due to their chemical properties, are not very soluble in water, and as a result they appear in wastewater in very low concentrations.This is the case with Polycyclic Aromatic Hydrocarbons (PAH), which are important contaminants because they are highly toxic, and have mutagenic, teratogenic and carcinogenic properties (IARC, 2005).
The main source of PAH in wastewater occurs by it being deposited on surfaces from the atmosphere and subsequently these compounds are transferred by rain water (Blanchard et al., 2001).In wastewater, PAH with the least molecular weight appear in the highest concentration since their solubility decreases as molecular weight increases, and PAH with greater molecular weight are adsorbed onto particles (Charalabaki et al., 2005).It has been noted that sewage sludge in urban areas can contain greater concentrations of PAH compared to that from industrial or rural areas (Sánchez- Brunete et al., 2007).
The correct purification treatment of wastewater, along with some control over its use and quality, minimizes the health and environmental problems, even preventing their occurrence (Lubello et al., 2004).Hence, in 2007 a Spanish regulation was created concerning the reuse of wastewater (RD 1620(RD /2007;;BOE, 2007), which makes primary, secondary and tertiary treatments of wastewater which is going to be reused for irrigating crops mandatory, and which limits the concentrations of chemical substances and micro-organisms present in purified wastewater.One of the possible tertiary treatments is ozonization which Fertigation with wastewater on soil, pepper plant production and pollutant contents achieves significant reductions in the microbiological populations of wastewater.Segura et al. (2001) used purified ozone disinfected wastewater for the cultivation of melon and produced fruit without any microbial contamination, even in the fruit which was in direct contact with the soil.
The objective of this paper is to evaluate the suitability of Purified Urban Wastewater (PW) from Almería (Spain) for use in the fertigation of greenhouse horticultural crops by studying the water's effect on the soil-plant system, the crop yield, fruit quality and the presence of inorganic chemical contamination (heavy metals and arsenic), and organic (Polycyclic Aromatic Hydrocarbons) as crop food safety parameters using pepper (Capsicum annuum L.) as the model plant.

Origin and treatment of urban wastewater
The PW came from the wastewater purification plant in the city of Almería (36º50'N, 2º27'W).It has been estimated that this plant treats 15 hm 3 yr -1 .In this purification plant the primary treatment of wastewater is carried out by decanting the solids and breaking down the fatty emulsions.The next stage is the secondary treatment (biological) by activated sludge.After these treatments, the water is sent to the tertiary treatment plant located 6 km away in Viator.In this second treatment plant, the water undergoes a chloration process with sodium hypochlorite followed by ozonation using a Tonozone T.E.F.ozonizor.The dosage used in both treatments varies depending on the microbiological contamination present in the purified wastewater to be treated.The disinfection process eliminates on average 99.96% of the total coliforms, faecal coliforms and faecal streptococcus present in the water.

Soil and growing conditions
Cultivation was carried out at the Research Station of the IFAPA (Andalusian Autonomous Government) in La Cañada (Almería, Spain), in a multi-tunnel greenhouse containing climatological equipment (air temperature and relative humidity sensor) to automatically control ventilation.The greenhouse covers an area of 800 m 2 , divided into 16 plots.In August 2007, pepper seedlings plants (cv.Aifos) were transplanted in the summer-autumn cycle (175 days).The planting density was set at 2 plants m -2 .The experimental design was random blocks with four treatments and four replications.
The soil used for cultivation is an artificial soil called "sandy mulch" and classified by the FAO as a cumilic anthrosol.This type of soil is made by adding a 10-15 cm layer of earth with variable texture (clay to sandy loam) on top of natural soil, adding 2 cm of manure, and then covering the manure with a layer of around 10 cm of sand.Before cultivation began, a characterization analysis of the soil was carried out.The layer of sand was not taken into account in this analysis.It was a basic soil with a pH of 8.6 and 4.5% carbonates.The electrical conductivity is 1.8 dS m -1 and it had low organic mineral content (0.9 %).
The application of the fertilizers was by drip irrigation using a dripper for each plant with a volume of flow of 3 L h -1 .Two different sources of water were used for the fertigation of the crop, Ground Water (GW) as a control, and PW from the Viator (Almería) ozonizator plant.The chemical characteristics of the waters studied are shown in Table 1.
Four irrigation treatments were carried out: GW, PW, Fertilized GW (FGW) and Fertilized PW (FPW).Composition and volumes of irrigation solutions and total nutrients applied are presented in Table 2. Fertilizers used were ammonium nitrate (NH 4 NO 3 ), potassium sulphate (K 2 SO 4 ) and phosphoric acid (H 3 PO 4 ).Pepper fertilization was carried out in accordance with crop extractions under local conditions (Contreras et al., 2006).The irrigation rates were estimated according to the ETc (Fernandez, 2000) and measures of soil matrix potential at 15 cm depth by tensiometers (Tensiometers Irrometer, Irrometer, USA) to maintain tension near 15 kPa.

Sampling and analysis of the water
Samples of GW and PW were collected every month in plastic containers to be analysed for inorganic contaminants (Cr, Ni, As, Cd and Pb), micronutrients (Mn, Cu, Zn), macronutrients (P, Ca, Mg, K) and Na.Measurements were made by Inductive Coupled Plasma-Mass Spectrometry (ICP-MS) Elan 6000 Perkin-Elmer Sciex equipped with auto-sampler AS 91 (Canada).A basic agronomic analysis was carried out to determine the pH, electrical conductivity (EC), bicarbonates, nitrates, sulphates, chlorides and ammonium in GW  , 1994); CO 32-and HCO 3 -were determined with a normalized solution of HCl (MAPA, 1994).
In parallel to this, PW and GW samples were collected in translucent 1000 mL glass bottles to determine the PAH.The PAH analyzed were: benzo[a]pyrene (BaP) and the sum of benzo[b]fluoranthene (BbF), benzo[ghi]perylene (BghiP), benzo[k]fluoranthene (BkF) and indene[1,2,3-cd]pyrene (IcdP) in accordance with the Spanish human water consumption regulation (RD 140/2003;BOE, 2003).The water samples were filtered, and a liquid-liquid extraction was made with hexane.Then the solvent was eliminated to 1 mL dis-solution.The samples were injected into GC-MS/MS (Varian 3800 GC fitted with a Varian Saturn 2200 mass detector with a 1079 PTV injector, USA).

Sampling and analysis of soil, plant and fruit
After the growing cycle (175 days), samples of the soil from each treatment and repetition at 15 cm from the plants were taken.The total content of heavy metals (Cr, Mn, Ni, Cu, Zn, Cd and Pb) and As were determined by digestion with HCl-HNO 3 (3:1) in a microwave oven (CEM MARS Xpress, USA).The dissolutions obtained were stored in polyethylene bottles at 4ºC for ICP-MS analysis.Soil and fruit PAH extraction was carried out using an accelerated solvent extractor (ASE350 Dionex) with dichloromethane:acetone 50:50 (v/v) at 100ºC, 1500 psi for 5 min.The solvent was eliminated by nitrogen flow and the residue was Table 1.Characterization of ground water (GW) and purified urban wastewater (PW) used in the experiments, concentrations of inorganic contaminants and maximum concentration allowed Whole plants, excluding the roots, were harvested to the end of experiment.Two plants per plot were randomly selected for destructive sampling and separated into different plant fractions: developing fruits, leaves and stem.Harvested fruits were randomly selected every harvesting, collecting 10 fruits per plot.Fresh samples for the different plant parts were dried at 70ºC to constant weight, and dry matter was determined in samples.
To analyse the heavy metals in the plant, for both micronutrients (Mn, Cu and Zn) and contaminants (As, Cd and Pb), the leaf and fruit samples were digested after the cultivation cycle in a microwave oven with a mixture of HNO 3 -H 2 O 2 .Analysis was carried out by ICP-MS.
The bioavailability of heavy metals and Arsenic was determined by the Concentration Factor (CF) of each element.The CF is defined as the relationship as a percentage between the concentration of the element in each plant organ and its concentration in the soil CF = 100 * C plant / C soil (Mohamed et al., 2003;Cui et al., 2005).
The production of mature fruit was quantified and classified by calibre categories according to the European Quality and Commercialization Norm (Commission Regulation (EC) No 2147No /2002;;OJ, 2002).The fruit was classified by its equatorial diameter as medium sized (50-70 mm), large (70-90 mm) and very large (> 90 mm).Fruit less than 50 mm was not taken into account.
The organoleptical quality variables of the fruit analysed were: pH, titratable acidity and total soluble solids (ºBrix) in the juice and hardness of the fruit.To determine fruit quality, pepper fruits were liquidized and turned into juice from each harvest.The content of total soluble solids from the pepper juice was determined with an Atago N1 manual refractometer, the results being expressed in ºBrix at 20ºC.The titratable acidity of the fruit was determined using a 0.1 M NaOH solution (AOAC, 1995) and expressed as mmolc of citric acid per litre of juice.The pH of the pepper juice was determined with a micro pH 2001 Crison pH-meter (Crison Instruments, Spain).The hardness of the fruit was determined with a Fruit Pressure Tester FT 327 (Facchini SRL; Italy) using a tip of 0.5 cm 2 (8 mm diam).

Data processing
The SPSS 16.0 software package was used to analyse the data in terms of descriptive statistics, standard error, univariant analysis and the Duncan's test with a 95% significance level (p ≤ 0.05).

Effects of the type of water and fertilization on the growth, production and quality of the pepper
The total biomass production of the crop is shown in Fig. 1.The production of leaves, stems and biomass, expressed as the sum of each was significantly greater in the PW treatment compared to the GW treatment, and was statistically not different to the FGW and FPW treatments.Purified Wastewater did not negatively affect crop development and achieved growth equal to the treatments with mineral fertilizers.
With regard to crop yield and fruit size (Fig. 2), the PW treatment achieved total production statistically equal to the treatments with mineral fertilization (FGW and FPW).The production of peppers classified as large exhibited significant differences.The treatments with fertilization (FPW and FGW) produced a more peppers classified as large (70-90 mm equatorial diameter) than treatments with PW and GW.In the very large category (> 90 mm equatorial diameter), the PW treatment exhibited pepper production significantly greater than with GW, and not different to FPW and FGW treatments.The production of medium sized pepper (50-70 mm equatorial diameter) did not exhibit significant differences amongst the treatments.The quality parameters pH, acidity, ºBrix in the juice and hardness of the fruit did not exhibit any differences among the applied treatments and were agree with the reference parameters (Urresterazu, 2004;Serrano & Ferna n dez-Trujillo, 2007) (Table 3).

Inorganic contamination (heavy metals and As) in water, soil, leaf and fruit
The results of the analysis for heavy metals and Arsenic in GW and PW are shown in Table 1.The micronutrients Mn, Cu and Zn appeared in higher concentration in GW, while the toxic elements Cr, Pb, Cd and As showed greater concentrations in PW.
In the soil (Fig. 3), none of the elements analysed (Cr, As, Cd, Pb, Ni, Mn, Cu and Zn) showed significant greater concentration between the irrigation treatments.Of the contaminants studied, Pb and As are found in greater concentrations, while Cd was in very low concentrations (< 0.30 mg kg -1 ).
The elements analysed (Mn, Cu, Zn, As, Cd and Pb) in fruit and leaf are shown in Table 4.In the leaf, the results for metals and As did not show significant differences for either the micronutrients or the contaminants between the applied treatments.The metals found in greater concentrations in the leaf were Mn, Cu and Zn.The FGW treatment showed greater concentration of Cu than the reference but were not significant higher than the other irrigation treatments, although no phytotoxicity symptoms or reductions in the biomass generated or in yield were observed.The three contaminants analysed (As, Cd and Pb), appeared in very low concentrations, and in the case of As were found to be below the detection limit (< 0.01 mg kg -1 d.m.).
The concentrations of heavy metals in fruit were lower than in leaf and were in the following order: Zn > Mn > Cu > Cd > Pb > As.No significant differences in the content of heavy metals in the pepper fruit were observed between the treatments for Mn, Cu, Zn, Cd and As.The CF order in leaf and fruit was: Cd  Cu > Zn > Mn > Pb  As (Table 5).The CF of Pb and As were very low (< 1%) therefore, although they were found in relative abundance in the soil, the crop did not uptake these elements.As and Pb were not bioavailable in the growing conditions.The Cd exhibited a high CF, but being found in low concentrations in the soil it did not represent risk and the final concentrations in leaf and fruit were low as has been shown.However, Cd was a highly mobile element and therefore crops which are developed under similar conditions to those in the present trial have to be controlled.

Organic contamination (PAH) in water, soils and fruits
The concentration of PAH in GW and PW was less than 0.005 µg L -1 for BaP and less than 0.10 µg L -1 for the sum of BbF, BghiP, BkF and IcdP.
As can be seen in Table 6, the total concentration of PAH (ΣPAH) in the soil before starting cultivation was significantly greater than the concentration of these compounds after cultivation for both GW and PW.Individually, the PAH which significantly reduced their concentration in the soil throughout cultivation in both the PW and GW treatments were Phe and Flu.The greatest concentrations of PAH in the soil before cultivation were probably due to the amounts of PAH in the manure used in making the sandy mulch.The largest group of PAH (Fig. 4) in both soil samples and manure corresponded to the 3 aromatic rings, mainly due to Phe.The most abundant PAHs in the soil were Phe, Flu, Fla and Py, whilst the lowest concentration showed the compounds with 5 aromatic rings, BaP, BkF and DBahA; of these, the last two exhibited concentrations in the soil lower than quantifiable limits.The main source of PAH in the soil at the beginning was manure, with PAH total concentration of 64.91 ng g -1 .
In the pepper fruit (Table 6) no significant differences were found in the total concentration of PAH between PW and GW treatments.As in the soil, the largest group found in the fruit corresponds to PAHs with 3 aromatic rings (Fig. 5), with Phe being found in the highest concentration in the two treatments, PW and GW.PAHs with 3 aromatic rings were found in significantly greater concentrations in pepper fruits fertigated with PW than GW (Fig. 5).In the PW treatment, Phe and BaA were found in significantly greater concentrations than in GW treatment.On the other hand, Fla and Flu were found in significantly greater concentrations in fruit with GW treatment.Naph, Ant, Ch, BbF, BkF, BaP and BghiP were not detected in pepper fruit in either of the two treatments.

Effects of the type of water and fertilization on the growth, production and quality of the pepper
The PW, FPW and FGW treatments did not exhibit differences in growth, yield or fruit quality.The better results from the PW treatment compared to the GW treatment are due to PW having greater concentrations of N-NH 4 + , P, K, S, Ca and Mg (Table 1).Because of this, a greater quantity of nutrients is carried to the plant throughout cultivation (Table 2).Mineral fertilization and water were not the only sources of nutrients; manure was also applied as an organic amendment (36% organic matter, 2.2% N, 0.8% P, 4.1% K).The mineralization of this organic material supplied nutrients to the crop slowly, which along with the nutrients contained in the PW were enough to equal in growth and production that plants treated with mineral fertilization (FGW and FPW).Manure on its own was not enough to obtain growth and yield equal to mineral fertilization, since the GW treatment produced lower yield and plant growth.Comparing the results obtained in yield with earlier studies under similar conditions (Contreras et al., 2006) it can be seen that production in this study is greater.The treatments without fertilization achieved a production level of 2768 g m -2 , well below that achieved with the PW treatment (Fig. 2).Table 6.Average PAH concentration (ng g -1 ) in the manure, initial soil, soil after purified urban wastewater (PW) or ground water (GW) irrigation and fruit of pepper irrigated with purified urban wastewater (PW) or ground water (GW) (n = 4)  The use of PW also brought about a great saving in mineral fertilizers.From the total N, P and K applied to the crop in the FPW treatment, 37% of N, 66% of P and 12% of K could already be found in the PW, whilst in GW the percentages decreased to 10%, 0% and 3% for N, P and K, respectively.

Inorganic contamination (heavy metals and As) in water, soil, leaf and fruit
The greater concentration of Cu, Zn and Mn in GW could be due to the continued addition throughout the years of quelated micronutrient fertilizers and their later lixiviation to the aquifer where the GW was taken from.
The low heavy metals concentrations in PW were due to a combination of several factors, the wastewater is mainly urban and the area has little industry, the effluent itself has a high pH which produces precipitation of the metallic cations (Karvelas et al., 2003), and the purifying system propels the heavy metals into the sewage sludge.The levels of metals and As observed in the two types of irrigation water were lower than those permitted by Spanish legislation concerning the use of regenerated wastewater for irrigating crops by localized irrigation (RD 1620(RD /2007;;BOE, 2007).
The lack of differences of soil heavy metals and As contents between irrigation treatments indicate that neither the accumulation of heavy metals nor As in the soil could be attributed to using PW.This result was to be expected due to the low concentrations of inorganic contaminants in PW.Mn and Zn were the main metals, and their presence in basic soils carries low risk as they are micronutrients for the plants and were not found in high concentrations.The results obtained do not concur with those found in the bibliography, since there are multiple studies that observe the accumulation of heavy metals and As in soils through use of wastewater and purified wastewater (Yadav et al., 2002;Kalavrouziotis et al., 2008).The fact that the concentration of metals and As in PW was very low, can be due to years of application of wastewater onto the soil are necessary to produce the accumulation of metals (Klay et al., 2010).According to the reference values for contaminated agricultural soils with pH ≥ 7 (Junta de Andalucía, 1999), as is the case, heavy metal contamination of the soil has not been observed.However, a slight As contamination has been detected.One has to take into account that the values in Fig. 3 correspond to the total extraction of heavy metals, and due to the soil pH (8.6) and the carbonate content (4.5%) its bioavailability will be low as showed CF.This factor were much lower than those published by Mohamed et al. (2003), with the exception of Cu.Pb stands out with a CF of 322 in the cited study.Khan et al. (2008) determined the CF in the edible parts of many vegetables cultivated on land irrigated with purified wastewater since the 1960s.For Cu and Pb in pepper fruit they obtained CF similar to those in this study, but for Cd and Zn the CF were higher.These different CF could be due to several factors, such as the different variety of pepper, the type of soil and agronomic techniques used which will affect the availability of heavy metals and As for the plant.
Heavy metals analysis in leaf and fruit showed that PW fertigation and the use of soluble mineral fertilizers did not increase the concentration of heavy metals and As compared to the GW and FGW.In the case of Pb, the GW treatment showed a significantly greater concentration of this metal in the fruit compared to the fertilized treatments (FPW and FGW) due to possible precipitation of lead phosphates in soil of treatment with P fertilization (Hashimoto et al., 2008).The foliar reference ranges for Mn, Cu and Zn are 50-250, 6-25 and 20-200 mg kg -1 , respectively (Mills & Benton Jones, 1996), therefore, they are within suitable levels.Arora et al. (2008) determined the Mn, Cu and Zn content in different plants irrigated with wastewater or with GW, and determined that the concentration of heavy metals in plants irrigated with wastewater was greater than that found when using GW.However, all the plants showed concentrations of Cu and Zn lower than the maximums permitted by the FAO/WHO, 40 and 60 mg Kg -1 , respectively (Codex Alimentarius, 1984).
It must be pointed out that the results shown, correspond to concentration in dry matter and pepper fruits are consumed as fresh matter, therefore all the contaminants were more diluted and their risk was reduced.The average moisture of the peppers was 95%, so these fruits meet Commission Regulation (EC) 466/2001 (OJ, 2001), which fixes the maximum content of determined contaminants in food products (Cd < 0.05 mg kg -1 f.w., Pb < 0.1 mg kg -1 f.w.).Fertigation with wastewater on soil, pepper plant production and pollutant contents Organic contamination (PAH) in water, soils and fruits GW and PW met the Spanish norm concerning water quality for human consumption (RD 140/2003;BOE, 2003) with respect to these compounds, and therefore the risk from their use in fertigation will be very low.
The concentrations of PAH found in PW were much lower than those found in the purified wastewater from the metropolitan area of Paris (France) (Blanchard et al., 2004) or in Venice (Italy) (Busetti et al., 2006).A fact that confirms that the amount of PAH in wastewater is greater in large cities due to the greater intensity of traffic on the roads and fumes from heating systems and industry (Manoli & Samara, 1999).
As with the case of heavy metals in the soil, the results found in the bibliography show accumulation of PAH in soils irrigated with wastewater (Tao et al., 2004;Xiao et al., 2008).There are marked differences between this study and those cited because in the latter studies the soils were influenced by atmospheric contamination from the urban area as well as industry in the area which added PAH to the soil from the air.Other differentiating factors were the period of usage of wastewater as irrigation water and the wastewater treatment.In previous studies wastewater has been used over several decades and at least in the articles cited, there is no indication if purification treatments were carried out.In this study the use of wastewater has been limited to a year, and the water has undergone an exhaustive process of purification with primary, secondary and tertiary treatments.Chung et al. (2008) carried out a one-year trial using PW with a total PAH concentration of 1.90 μg L -1 .They found a certain accumulation of PAH in the soil with final concentrations between 3.48 and 6.60 ng g -1 depending on the crop, values within the order of magnitude of this study (11.01 ng g -1 for GW and 10.12 ng g -1 in PW).The greater values of the GW and PW soil could be due to the initial soil showed 28.09 ng g -1 of total PAH.Al Nasir & Batarseh (2008) found greater accumulations in soil irrigated with purified wastewater during a cultivation cycle in several plant species.In the case of pepper, the increase in ΣPAH was 60.7 ng g -1 .
The reduction of ΣPAH at the end of the cultivation cycle in both treatments was greater than 60%.This result means that the PW did not add PAH to the soil, and that during the cultivation period the soil micro-organisms were able to degrade PAH.By group, the biggest reduction corresponds to the PAH with three aromatic rings, the percentage being 66% for the GW treatment and 70% for the PW treatment.
These discrepancies in the results for both heavy metals and PAH show the variability in the composition of urban wastewater even after having undergone a purification process.Therefore, periodic exhaustive controls of both the irrigation water and the soil have to be carried out.
The results for the concentration of PAHs found in earlier studies are very varied for plants irrigated with wastewater, from 0.0028 ng g -1 to 984 ng g -1 (Tao et al., 2004;Chung et al., 2008) depending on the crop, agronomic techniques and industrial influence.Al Nasir & Batarsech (2008) found a concentration of ΣPAH in pepper fruit of 31 ng g -1 , value some three times greater than the found in this study.In general, in all the studies reviewed it has been seen that PAH with greater accumulation in edible parts of the plants correspond to those with low molecular weight, and those with high molecular weight were found in very low concentrations, or concentrations below detectable limits.This tendency was also observed in the results obtained with the PW treatment, except for Naph which was not detected.
According IARC (2005), no PAH classifiable as carcinogenic nor probably carcinogenic to humans were detected in fruit.Only B[a]A presented carcinogenic significance and is classified as possibly carcinogenic to humans.Therefore, the dangers of PW for consumers are low, although due to the high variability of this type of effluent depending on where it originates from, periodic controls are necessary to evaluate possible invasion from contaminants, and in this way maintain the food safety aspect of the crop and the agro-ecosystem.
In conclusion, PW from Almería (Spain) can be used to irrigate horticultural crops with localized irrigation techniques, as long as its heavy metal, As and PAH concentrations are controlled.The effects on growth, production and pepper quality were comparable with those of the GW which is usually used, producing an important saving in N, P and K fertilization.After a cultivation cycle no accumulation of heavy metals, As nor PAH was observed in the soil; the principal source of PAH being the manure used.The concentrations of Mn, Cu, Zn, Cd, Pb, As and PAH in the fruit were not significantly different when using PW or GW and the fruits are suitable for human consumption.

Figure 1 .
Figure 1.Generation of biomass (dry matter) divided into pruning, leaves, stems and biomass (total) at the end of the cultivation cycle of the treatments ground water (GW), purified urban wastewater (PW), fertilized ground water (FGW) and fertilized purified urban wastewater (FPW) (n = 4).Different letters indicate significantly different treatments, a being the lowest (p ≤ 0.05).Bars indicate standard error.GW PW FGW FPW

Figure 2 .b
Figure 2. Production of pepper in terms of fresh weight classified according to fruit calibre, medium (50-70 mm), large (70-90 mm) and very large (> 90 mm) and total production of the treatments ground water (GW), purified urban wastewater (PW), fertilized ground water (FGW) and fertilized purified urban wastewater (FPW) (n = 16).Different letters indicate significantly different treatments for each calibre and total production, a being the lowest (p ≤ 0.05).Bars indicate standard error.Medium Large Very Large Total

Figure 3 .
Figure3.Total concentration of heavy metals and As present in the soil fertigated with ground water (GW), purified urban wastewater (PW), fertilized ground water (FGW) and fertilized purified urban wastewater (FPW) after the cultivation cycle and the Junta de Andalucía (1999) reference levels for contaminated agricultural soils (n = 4).Mn has not reference level.Bars indicate standard error.

Table 3 .
pH, acidity expressed as citric acid (g L -1 ) and ºBrix of the pepper juice and hardness (kg cm -2 ) of the pepper fruit for ground water (GW), purified urban wastewater (PW),

Table 4 .
Concentrations of Mn, Cu, Zn, As, Cd and Pb in leaves and pepper fruit (mg kg -1 dry matter) for ground water (GW), purified urban wastewater (PW), fertilized ground water (FGW) and fertilizer purified urban wastewater (FPW) irrigation treatments at the end of the cultivation cycle.(n= 4) * n.d.: not detected.Different letters indicate significantly different treatments, a being the lowest (p ≤ 0.05).

Table 5 .
Concentration factors (CF) of Mn, Cu, Zn, As, Cd and Pb for leaf and pepper fruit for ground water (GW), purified urban wastewater (PW), fertilized ground water (FGW) and fertilizer purified urban wastewater (FPW) irrigation treatments Fertigation with wastewater on soil, pepper plant production and pollutant contents