efecto compost vegetales de hojas verdes

Compost Science & Utilization, (2011), Vol. 19, No. 2,114-122

Effects of Green Compost on Soil BiochemicalCharacteristics and Nutritive Quality of Leafy Vegetables

S. Tavarini, R. Cardelli, A. Saviozzi, E. Degl'Innocenti and L. Guidi*Dipartimento di Biologia delle Piante Agrarie, Universit di Pisa, Pisa, Italy

*E-mail contact: [email protected]

Green compost can improve the quality of fruits and vegetables through an increase in sugars and organicacids, which is extremely important because today's consumer is concerned about healthy and nutritionalproperties of food. The aim of this research was to evaluate the effects of green compost used as soil amend-ment on phenols and vitamin C content and on antioxidant capacity of leafy vegetables. Plants of Lactucasativa var. acephala cultivar Red Salad Bowl and Spinacia olercea cultivar Lorelay were grown in three soiltreatments: a) 100% soil, b) 75% soil plus 25% compost c) 50% soil plus 50% compost. Results show thatadding green compost to soil improved its fertility and the growth of plants. The use of compost for lettuce

:, did not show any differences in antioxidant capacity and vitamin C content at either compost concentration(25 and 50%) while a significant reduction in phenols was evident. The Chi content decreased in spinach asa consequence of compost addition while no changes were observed in lettuce. However, the healthy prop-erties of spinach decreased as a ftmction of the compost concentration in the soil.

Introduction

When mature composts are used in agriculture,improvements in the growth of plants are seen, due tohigher levels of nutrients and organic matter (Maynard1995; Ozores-Hampton 1998; Cox et al. 2001; Ros et al.2005). Compost has been shown to be beneficial infruit, vegetable and ornamental crop production (Fitz-patrick et al 1998) because of its ability to improve sug-ars and organic acids levels. Its influence on phyto-chemicals content in vegetables is not yet well known.This is extremely important because today's consumeris concerned about the healthy and nutritional proper-ties of food. So the importance of phytochemicals in thehuman diet is growing, because they have a high an-tioxidant capacity and are helpful in preventingand/or curing a wide range of diseases (Ames et al.1993; Hertog et al 1993; Kaur and Kapoor 2001). In fact,the role of phytochemicals in oxidative stress is due tothe presence of Reactive Oxygen Substances (ROS) in-side the aerobial organisms (Rico et al 2007).

During decomposition, microorganisms assimi-late complex organic substances and release inorgan-ic nutrients. An adequate composting process killspathogens and stabilizes compost organic carbon be-fore material is land applied (Erickson et al. 2009).The phytonutritive capacity of compost has oftenbeen demonstrated to be analogous to that of ma-nure; the same level of productivity, both quantita-tively and qualitatively, can be maintained by re-

placing manure with compost (Hepperly et al. 2009).One method to improve soil quality involves the

addition of several kinds of composts made from solidorganic waste, sewage sludge, green and industrialwastes, or animal manure (Masciandaro et al 2000; Coxet al 2001; Madrid et al 2007). Compost is considered tobe environmentally safe, agronomically advantageous,and a relatively cheap soil organic amendment (Hep-perly et al 2009). Compost green wastes (from recycledgarden waste which includes grass clippings, hedgecuttings and wood materials), used as organic amend-ments in soil, are considered advantageous comparedwith other aforementioned organic wastes as they havea low a risk of toxicity due to the presence of heavy met-als, pollutants, etc. (Nicholson et al 2003). Moreover,mass production of green compost helps local authori-ties achieve recycling targets and provides a worthy al-ternative to traditional amendments.

The objectives of this research were to evaluatethe effects of green compost, used as soil amendment,on soil biochemical characteristics and on the nutri-tional aspects of two vegetables widely used as freshsalad: (i) spinach (Spinacia olercea) and (2) lettuce (Lac-tuca sativa var. acephala cultivar Red Salad Bowl).

Materials and MethodsPlant Material, Soil and Green Compost Characteristics

Seeds of spinach (Spinacia olercea L., cultivarLorelay) and lettuce (Lactuca sativa var. acephala Dill.,

114 Compost Science & Utilization Spring 2011

Effects of Green Compost on Soil Biochemical Characteristics and Nutritive Quality of Leafy Vegetables

cultivar Red Salad Bowl) were sown in a sterilizedsoil. Lettuce seedlings emerged after 15-20 days andspinach emerged after 30-40 days.

The soil used was collected from the top 15 cm ofthe soil surface at the Interdepartmental Centre "E.Avanzi" (University of Pisa, Pisa, Italy). Samples offield-moist soil were air dried, crushed to pass a 2-mmsieve to remove large residue fragments and analyzedfor physical and chemical properties (Table 1). Granu-lometric composition, carbonate content, cation ex-change capacity (CEC) and pH were determined ac-cording to the MAAF (1994). Organic carbon and total Swere determined by dry combustion (induction furnace900CS, ELTRA) after removing carbonate-C (Nelsonand Sommers 1982); total N determination was madeby the Kjeldahl procedure after acid digestion (Bremn-er and Mulvaney 1982). Available P was determined ac-cording to the Olsen method (sample:extractant; 1:20).

TABLE 1.Main characteristics of soil and green compost

Sand (g kg')sut (g kg')Clay (g kg')pHTotal organic carbon (g kg')CaCO,(gkg')Total N(g kg')RaHo C/NCation exchange capacity (cmol kg )Available P (mg kg')Total P (g kg')Total S (g kg')

Soil

65.922.611.58.29.656.11.029.47.017.0

-

0.11

Compost

-

-

-

8.1310.0

-

11.8026.3

-

-

1.312.50

The green waste compost (GWC) was taken fromthe CERMEC facility (Massa Carrara, Italy), which isdesigned to accept green waste from neighbouringproducers. The composting process was designed as aforced-air, in-vessel composting process, which pro-vided an initial decomposition process over a pre-de-termined period. The composted material was re-moved from the tunnels and placed in "windrows" ina maturation area, for a 3 months period before beingscreened, ready for delivery to end-users.

The main characteristics of the GWC were deter-mined according to the methods used for the soil. ,

Treatments

Both species were grown in three soil treatments:a) 100% soil, b) 75% soil plus 25% compost and c) 50%soil plus 50%. They were planted in pots (20x50cm)

and grown in a greenhouse (T=253C; RH 75 5%with a photoperiod of 12/12 (day/night) at an irradi-ance of about 400 \in\o\ m s ).

Soil Analysis

Dissolved orgarc carbon (DOC) was determinedby stirring samples of soil with distilled water (soil: HO;1:50) for 24 h at room temperature, centrifuging the sus-pension at 10,000 rpm for 10 min. After filtrationthrough a 0.4 mm glass fibre, carbon content was deter-mined by using dichromate oxidafion titration (Ciavat-ta et ai 1991). On an aqueous extract obtained by shak-ing samples of soils and distilled water in a 1:5 ratio for1 h, the following parameters were determined: electri-cal conductivity (E.C.), using a cell calibrated with NaClstandards; NO ' content by ion chromatography(Dionex apparatus, mod. DX-lOO); and phenolic com-pounds, expressed as coumaric acid, by a modificafionof the method of Folin (Kuwatsuka and Shindo 1973).

Soil dehydrogenase activity was determined us-ing triphenyl tetrazolium chloride (TTC) as a substrate(Casida et al. 1964). Microorganisms reduced TTC totriphenyl formazan (TPF) after 24 h of incubation at37C. TPF product was determined spectrophotomet-rically at 488 nm.

The catalase activity was determined as describedby Beck (1971) by using H O as a substrate. Oxygenwas measured with a gas volumetric method duringincubation of 3 minutes.

Arylsulfatase activity was determined using p-ni-trophenyl sulfate as a substrate. Soil samples were in-cubated at 37C for 1 hour and the formed p-nitrophe-nol was extracted by using dilute alkali (CaCl 0.5Mand NaOH 0.5M) and determined at 40t) nm(Tabatabai and Bremner 1970).

Amylase activity was determined using a starch-NaCl solution as a substrate. Soil samples were incu-bated at 50C and pH 6.5 for 24 hours and the formedglucose was measured with Reflectoquant test(Cardelli et. al. 2001).

Alkaline phosphatase activity was determinedbased on colorimetric estimation of the p-nitrophenolreleased by phosphatase activity when soil was incu-bated with buffered (pH 11) sodium p-nitrophenylphosphate solution and toluene at 37 C for 1 hour(Tabatabai and Bremner 1969).

Protease activity was analysed by a colorimetricmethod, using sodium caseinate as substrate. Soilsamples were incubated at 50C for 2 hours and theformed tyrosine was determined at 700 nm (Ladd andButler 1972).

Two indexes of soil quality, the biological index offertility (BIF) and the enzyme activity number (EAN),

Compost Science & Utilization Spring 2011 115

s. Tavarini, R. Cardelli, A. Saviozzi, E. Degl'Innocenti and L. Guidi

were calculated as follows: the BIF is (dehydrogenaseH- k catalase)/2 where k is a proportionality catalasecoefficient (Stefanie et al. 1984); the EAN is 0.2 (dehy-drogenase-i-catalase/10 -i- alkaline phosphatase/40-i-protease/2-t-amylase/20) (Beck 1984).

Plant Analysis

Three discs (9 mm D) were taken from each leafsample, extracted with 80% (V/V) acetone solution,then centrifuged at lOOOOg for 5 min. An aliquot of thesupernatant were used for spectrophotometric deter-mination of Chi a, Chi t at a wavelength of 663 and 648nm respectively and then the total Chi content wasmeasured. The concentrations were determined basedon the formulas reported by Lichtenthaler (1987). Thefinal value was expressed as ]ig Chi /g FW.

The extraction of phenolic compounds was per-formed as described by Degl'Innocenti et al. (2005).The total phenolic content in the methanol extractswas determined according to Folin Ciocalteu byDeglTnnocenti et al. (2007). Absorbance was measuredat 765 nm using a UV-Vis spectrophotometer. The fi-nal was expressed as mg gallic acid/100 g FW.

ASA and DHA were determined spectrophoto-metrically as described by Kampfenkel et al. (1995).Leaf material (about 0.4 g) was transferred to 6%(W/V) trichloroacetic acid (TCA) and the mixture wasstirred continuously for 15 min; the mixture was cen-trifuged at 15000g for 5 min at 4C and the supernatantwas immediately used for analysis of ascorbate. The fi-nal value was expressed as mg ascorbic acid/lOOg FW.

The free radical scavenging activities (DPPH as-say), as well as the ferric reducing ability (FRAP as-say), were used to evaluate the antioxidant capacityof the extracts.

Radical Scavenging Activity DPPH (a,a-difenil--picrilidrazile) - A 0.5 mL aliquot of the methanol ex-tract prepared above was mixed with 0.25 mL of anethanolic 0.5 mM DPPH solution and 0.5 mL of 100mM acetate buffer (pH 5.5). The tubes were mixed for15 s and, after standing for 30 min, the absorbance ofthe mixture was measured at 517 nm (Kang andSaltveit 2002). The final value was expressed as

OD/100 g FW.Ferric-Reducing Antioxidant Power (FRAP) Assay -

The assay was based upon the methodology of Benzieand Strain (1996). The FRAP reagent was preparedfresh. It contained 1 mM 2,4,6-tripyridyl-2-triazine(TPTZ) and 2 mM ferric chloride in 0.25 M sodium ac-etate at pH 3.6. A 100 i^L aliquot of the methanol ex-tract prepared above was added to 900 pL of FRAPreagent, and the they were mixed. After the mixturestood at 20C for 4 min, the absorbance at 593 nm wasdetermined against a water blank. Calibration wasagainst a standard curve (50-1000 \iM ferrous ion) pro-duced by the addition of freshly prepared ammoniumferrous sulfate. The final value of antioxidants capaci-ty was expressed as mol Fe^^/100 g FW.

Statistical Analysis

The experiment was conducted twice inspringer-summer 2008. Data represent the mean ofdifferent replicates. Results of soil characteristics andbiochemical activity were subjected to one-wayanalysis of variance (ANOVA) with treatment asvariability factor. A two-way ANOVA test was ap-plied to plant characteristics and nutritional qualityin which treatment and species are the source of vari-ability. Means are compared by using a least signifi-cant difference (LSD ) test. For correlation betweenchemical characteristic of soil and nutritional qualityof vegetables a regression analysis was carried outand the correlation coefficient and its significancewere determined.

Results

Physical, Chemical and Biochemical Soil Properties

The soil used for the experiment (Table 1) had asub-alkaline pH, a low cation exchange capacity and asandy loam texture. Organic carbon content was ratherlow (9.6 g Kg "^ ) but C/N ratio was very good. Nitrates,phenols, dissolved organic carbon and salinity in-creased with increasing doses of compost (Table 2).There were significant differences in dehydrogenases

TABLE 2.Influence of green compost added at different percentage on some chemical and physical characteristics of soil.

In each column, means followed by the same letter are not significantly different at p


40'

30-

20-

10-

(a)

i I '(b)

b

c

a

25% 50% 25% 50%

FIGURE 1. Dehydrogenase (a) and catalase (b) activities in soil amended with different percentage of green compost. Means with the sameletters are not significantly different at p


Quality of Leafy Vegetables

There were no significant differences in the freshweight of leaf spinach among the three different treat-ments (Table 3); however, a significant increase infresh weight of leaf lettuce was observed when plantsare grown in soil amended with the green compost.

TABLE 3.Leaf fresh weight (FW) of spinach and lettuce grown in soilamended with different percentage of green compost. For

the two species spinach and lettuce, means followed by thesame letters are not significantly different for p = 0.05.

500-

Treatments Spinach Lettuce

0 Compost25% Compost50% Compost

6.9+0.12 a5.00.57 a5.90.79 a

2.20.36c4.50.46b19.21.02a

2500'(a)

FIGURE 3. Chlorophyll a (a), b (b) and total (chlorophyll fl+W (c) inspinach (black) and lettuce (white) grown in soil amended with dif-ferent percentage of green compost. For the two species spinachand lettuce, means followed by the same letters are not significant-ly different for p = 0.05.

25% 50%FIGURE 4. Total phenol content in spinach (black) and lettuce(white) grown in soil amended with different percentage of greencompost. For the two species spinach and lettuce, means followedby the same letters are not significantly different for p = 0.05.

Total chlorophyll content in spinach was higherthan in lettuce (Figure 3), but it decreased with the useof green compost. The lowest value of Chi a content wasrecorded in plants grown in soil amended with 50% ofcompost, whereas for this treatment Chi b values weresimilar to plants grown in soil without compost.

Figure 4 showed the phenols content in leaves oftwo vegetables grown with or without compost in thesoil. Phenols were higher in spinach leaves of plantsgrown on soil control and on 25% compost treatmentscompared to lettuce. In lettuce the compost did not in-fluence positively the phenol content.

Vitamin C content was much higher in spinach,with no strong differences among treatments, exceptfor the treatment with 25% compost in which a sig-nificant decrease in vitamin C was observed (Figure5). In lettuce there were no statistical differencesamong treatments.

25% 50%

FIGURE 5. Vitamin C content in spinach (black) and lettuce (white)grown in soil amended with different percentage of green compost.For the two species spinach and lettuce, means followed by thesame letters are not significantly different for p = 0.05.

The antioxidant capacity assays were reported inFigure 6. Both FRAP and DPPH showed higher val-ues in spinach as compared to lettuce extracts. Thecapacity to reduce TPTZ-Fe"'* complex to TPTZ-Fe^ "^of spinach extracts decreased significantly when



0.8'

25% 50%

FIGURE 6. Antioxidant capacity determined by FRAP (a) andDPPH (b) assay in spinach (black) and lettuce (white) grown in soilamended with different percentage of green compost. For the twospecies spinach and lettuce, means followed by the same letters arenot sigrficantly different for p = 0.05.

plants were grown with compost and a similar be-haviour was observed also for lettuce. The results ob-tained with the DPPH assay showed a different be-haviour. In spinach the antioxidant capacityincreased significantly in leaves of plants grown withsoil amended with 25% of green compost whereasthe highest dose of compost induced a significant de-crease as compared to plants grown without com-post (Figure 6b). In lettuce, no variations was ob-served for DPPH values in relation to the presence ofcompost in the soil.

Correlation Between Chemical and Biochemical SoilCharacteristics and Quality of Vegetables

In Table 4 the correlation coefficients betweensome soil characteristics and quality of leaf vegetablesare reported. Different results were found between thetwo vegetables. Nitrates, phenols and DOC, that in-creased in soil amended with compost (see Table 2),induced a significant decrease in total Chi of spinachleaves, as well as phenol content, while no correlationswere observed with vitamin C. BIF index was nega-tively correlated with all parameters with the excep-tion of vitamin C, whereas EAN index was signifi-cantly correlated with total Chi and phenols. A verydifferent picture was shown in lettuce; in fact, onlyleaf fresh weight was positively correlated with all soiltested parameters (Table 4).

TABLE 4Correlation coefficients between some chemicalcharacteristics of soil and quality parameters ofspinach and lettuce. In the table the correlationcoefficients and their significance are reported.

*": p


ment of the native soil DOC. In our study, the appli-cation of compost caused a significant increase inDOC, depending on the doses.

With increasing doses of compost applied to soil,phenolic compounds in soil and salinity increase, butthey never reached levels that disturb soil, plant health orgrowth (Mizrahi and Pasternak 1985; Okolo et ai 2007).

There is considerable interest in soil enzyme activi-ties as indicators of changes following soil treatment ef-fects such as the incorporation of organic materials. Thepractice may influence enzymafic acfivifies in soil be-cause the added organic fractions may contain intra-and extra-cellular enzymes (Garcia et ai 1993) and canalso stimulate the growth of soil microbiota in responseto the presence of easily available C (Bhattacharyya etai 2001; Saviozzi et al. 2002). For example. Cooper andWarman (1997) and Saviozzi et ai (2002) reported a sig-nificant increase of dehydrogenase by amendments forsoils low in organic matter content. In this research, allenzyme activifies in amended soil, both oxidoreductaseand hydrolase and the soil quality indexes (BIF andEAN), were much higher than the respective controls(no compost), showing an increase almost proportionalto the applied dose. This confirms that compost addi-tion to soil causes the stimulafion of microbial activityas a result of the rapid mineralization of the organicmatter in sou and availability of the more easily degrad-able substrates. Indeed, the highest level of enzyme ac-tivities occurred at the greatest dose of compost. Thevery large increase of amylase activity may be ex-plained with the high hydrosoluble sugars content sup-ply from this well matured compost, which representsan easily available source of energy for microorgan-isms. Spinach and lettuce may have taken advantage ofthe high soil enzyme acfivities, and thus were able to as-sure a fuU availability of nutrients.

The only excepfion of the general increasing trendshowed by the enzyme activity following compost ad-difion is the arylsulphatase. Arylsulphatase is consid-ered to be responsible for sulphur cycling in soil be-cause it is involved in the processes of organic Smineralization. In our research, although the sou sam-ples treated with compost showed higher arylsul-phatase activity than the control soil, suggesting thatthe amendment was able to stimulate the enzyme insoil, a dose-response was not effective. This indicatesthat to enhance the arylsulphatase activity of the usedsoil the low applicafion rate is sufficient. As suggestedby Maynard et al. (1985) and Vong et al. (2004), the aryl-sulphatase acfivity may be under feedback control in in-organic S, i.e. the greater amount of inorganic sulphates,likely released by the samples treated with the highestdose of compost, may have had a negative effect on theactivity of the enzyme.

Conclusion

The addition of compost to soil improved itschemical and physical characteristics. However, thisdoes not induce an improvement of leaf vegetablequality as evidenced by studied nutritional aspectsand by statistical analysis of correlations. Moreover,the type of soil is important in determining the re-sponse of crop to compost (Ouedraogo et al. 2001).Also the crop species used are important. Ribeiro et al.(2007) have found that the use of compost obtainedfrom forestry wastes and solid phase of pig slurry didnot affect the germination and growth of lettuce, con-trasting with tomato seedlings where the highestgrowth occurred at the maximum concentration ofcompost used (100%).

The addition of compost as a soil supplementdecreased the total Chi content in spinach leavesthat could be attributed to the increases of the valuesof soil parameters, as confirmed by the negative cor-relations (see Table 4), while no changes were ob-served in lettuce. Other authors have been found asthe addition of compost to soil did not induce varia-tions in chlorophyll content in Cuphea hyssopifolia, inwhich also a strong decrease in the growth was ob-served (Wilson et al. 2001). In our study we havefound no changes in leaf fresh weight in spinach. Anincrease in lettuce has already been reported by oth-er authors (Ouedraogo et al. 2001; Lee et al. 2004; Aliet al 2007).

The use of compost for lettuce did not show anydifferences in antioxidant capacity and vitamin C con-tent at either compost concentration (25 and 50%)while a significant reduction in phenols was evident.This is confirmed by the lack of significant correlationsbetween soil parameters and quality of the vegetable.

The antioxidant capacity of spinach leaves de-creased when 50% compost was added to soil. Asimilar behaviour was observed for phenols contentwhile vitamin C, the most important phytochemicalin this leafy vegetable, showed the lowest valuewhen 25% compost was used. Such results showthat the addition of compost to soil improves fertil-ity with a beneficial effect on growth of lettuce.However the healthy properties of spinach were re-duced as a function of compost concentration in thesubstrate. .

This study represents a first approach to an im-portant thematic that involves the influence of agro-nomical practices (such as the use of amendment insoil) on the nutritional quality of leaf vegetables. Inaddition, the usage of green compost represents animportant economical and technical aspect for a sus-tainable agriculture.



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