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Effect of a sugarcane straw leachate and its chemical constituents on plant growth in soil conditions

Diego A. Sampietro, María I. Isla and Marta A. Vattuone

Cátedra de Fitoquímica. Instituto de Estudios Vegetales “Dr. A. R. Sampietro”. Facultad de Bioquímica, Química y Farmacia. Universidad Nacional de Tucumán. España 2903. CP 4000. San Miguel de Tucumán. Argentina. E-mail:


In a previous study, we isolated three phenolic acids from sugarcane (Saccharum officinarum) (Poaceae) straw leachate. The aim of this work was to evaluate the leachate and its isolated constituents on plant growth in unsterilised soil, autoclaved soil and autoclaved sand. Amaranthus quitensis L., Bidens subalternans L. (Asteraceae), Brassica campestris L. (Brassicaceae), Sida rhombifolia L. (Malvaceae), Lactuca sativa L. (Asteraceae), Raphanus sativus L. (Brassicaceae), Sorghum bicolor (L.) Moench (Poaceae) and Triticum aestivum L. (Poaceae) were selected as test plants. In unsterile soil, straw leachates stimulated the growth of some test plants at the lowest assayed concentration and inhibited root elongation of all of them at higher concentrations. The leachates inhibited root elongation in unsterile soil more than in autoclaved soil. In autoclaved sand, the inhibitory effect was stronger than in autoclaved soil. In unsterile soil, phenolic acids stimulated radicle growth of some test plants at low concentrations but were inhibitory at higher concentrations. Vanillic acid was more inhibitory than ferulic and syringic acids. Phenolic acids inhibited root growth of test plants in autoclaved soil more than in unsterilised soil. Otherwise, the test plants were more inhibited in sterile sand than in autoclaved soil. Our results suggest that soil microflora and adsorption influenced the biological activity of the straw leachate and its phenolic acids. The leachate content of vanillic and ferulic acids is high enough to inhibit seedling growth of some test plants. However, the phytotoxic effect of these compounds was weak and can not completely explain the strong inhibitory activity of sugarcane straw leachate.

Media summary

The influence of soil factors on the inhibitory effect of a sugarcane straw leachate and its constituents was evaluated on plant growth.

Key Words

Sugarcane straw leachate, phenolic acids, soil conditions, root elongation.


The involvement of phenolic acids on plant-plant interactions in natural and cultivated ecosystems is often suggested (Rice 1995). In agroecosystems, these compounds can be leach from plant straws into soil (Souto et al. 2001). Once phenolic acids enter in the soil their phytotoxic activity is a function of the complex interactions among the physiological and ecological properties of both donor and receiver plants under complicated environmental conditions (Katsuichiro 2004). We isolated three phenolic acids in a bioassay guided fractionation from sugarcane straw leachate (Sampietro et al. 2004). These compounds inhibited the growth of annual weeds, usually found in the borders of sugarcane fields, in Petri dish bioassays (Sampietro et al. 2005). However, it was suggested that in absence of soil, growth bioassays, may be of little or no significance for understanding a plant-plant interaction (Inderjit and Weston 2000). We hypothesized that the straw leachate and its isolated phenolic acids can inhibit the growth of receiver plants (annual weeds) in soil conditions. The influence of soil biotic and abiotic factors was also evaluated.


Plant and soil materials

Samples of sugarcane (Saccharum officinarum cv. ´Tuc [CP] 77-42´) straw were collected from a field near Los Ralos (26° 52' S, 64° 58' W, Tucumán, Argentina) and dried at 60°C for 48 h. Seeds of pigweed (A. quitensis L.), arrowleaf Sida (S. rhombifolia L.), wild mustard (B. campestris L.) and beggarticks (B. subalternans L.) were collected from the borders of the sugarcane field. Seeds of wheat (T. aestivum L. var. ´Pegaso´), sorghum (S. bicolor [L.] Moench var. ´Supergauchazo´), radish (R. sativus L. var. ´Sparkler´) and lettuce (L. sativa L. var. ´Grand Rapids´) were provided by the Estación Experimental Agroindustrial “Obispo Colombres” (Tucumán, Argentina). Seeds of the selected test plants were sterilized with 1% sodium hypochlorite for 30 min, rinsed with sterile distilled water, dried with sterile filter paper and germinated in water. The germinated seeds with a root length of 1 mm were used in the growth experiments.

Soil (silty-loam) from 0-10 cm depth was collected from Los Ralos, air dried at room temperature, sieved (2 mm sieve) and stored in paper bags before use.

Sugarcane leachate preparation

Different amounts of dry sugarcane straw (6, 19, 45 and 64 g) were soaked in 1000 ml of water for 4 h. The leachates obtained were sterilized by passage through sterile filter membranes (Millipore, 0.22 µm) and assayed.

Growth experiments

The biological activity of the leachates was evaluated on selected test plants according to Tongma et al. (1998) with some modifications. Small glass bottles (4 cm internal diameter, 8 cm height, 12.56 cm2 transversal area) were filled with 26.6 g of dry-sieved soil or 26.6 g of dry-washed sand. The sand-filled bottles and half of the soil-filled bottles were autoclaved three times at 120 ºC and 103 kPa for 30 min. Fifteen ml of water (as control) or a sugarcane leachate were added to each bottle. Five uniformly germinated seeds of each test plant were sown 2 mm from the soil surface. The bottles were covered with sterile translucient plastic caps and placed in a growth chamber (25°C with a 16-h photoperiod at 400 µmol /m2 s photosynthetically active radiation) for 4d, after which shoot and root length of the test plants were measured. Vanillic, syringic and ferulic acids were also assayed in autoclaved soil, autoclaved sand and unsterile soil using the same conditions outlined previously.

Leachate content of the isolated phenolic acids

A known volume of the straw leachate was extracted with diethyl ether. The diethyl ether fraction was injected in a micropreparative HPLC column. The elution was performed at a flow rate of 2 ml/min with solvent A (2% HOAc in H2O) and B ( 2% HOAc in MeOH) in the ratio of 35 % B for 5 min, increasing to 90 % B in 15 min; after 5 min at 90% B, the column was cleaned-up decreasing to 35% B in 5 min. Re-equilibration was done at 35% B for 20 min. Compounds were detected at 266 nm. Peaks with retention times corresponding to vanillic, ferulic and syringic acids were collected. The peaks were injected in a GC-MS after permethylation with CH2N2 (Fieser and Fieser. 1967). GC-MS conditions were as follow: 2 µl of the sample were injected to the capillary column and the temperature gradient was: from 90 to 270° C, 90°C (2 min), 90 to 150°C (3 min), 150°C (1 min), 150 to 270°C (9 min) and 270°C (1 min). UV-Visible and MS spectra confirm the identity of the isolated peaks. The content of the phenolic acids in the straw leachate was determined by HPLC using external standards.

Statistical analysis

All experiments were duplicated using a completely randomized design with four replications. After testing normality and homogeinity of variances, all data were analyzed by one-way Anova and LSD test to compare multiple means. Data of duplicated experiments were combined and are presented as the pooled mean values.

Results and Discussion

Sugarcane straw leachates affected the growth of the test plants in unsterile soil, autoclaved soil and autoclaved sand. Shoot elongation was largely unaffected. The density of the collected straw samples varied between 72 and 764 g dry straw/m2 indicating that sugarcane straw was not uniformly distributed in the field. Consequently, we prepared straw leachates into the mentioned amounts of straw per soil area unit.

In unsterile soil, the effect of the straw leachate on plant growth was dependent of the test plant. The lowest straw leachate concentration significantly stimulated root growth of wheat, sorghum, radish, pigweed and wild mustard (not shown). The same concentration did not affect root elongation of arrowleaf sida, and beggarticks, while lettuce was inhibited. Concentrations higher than 6 g of dry straw/l inhibited root growth. Inhibition increased proportionally to the amount of leachate present.

The inhibitory activity of the isolated phenolic acids was evaluated through the concentration of these compounds needed to inhibit 30% root elongation (EC30) in unsterile soil. The biological activity of the three compounds was weak (Table 1). In general, vanillic acid was more inhibitory than ferulic and syringic acids. Otherwise, low concentrations of these compounds stimulated the growth of some of the test plants.

Table 1. EC30 values of a Saccharum officinarum straw leachate and its isolated compounds determined on root elongation of test plants.

Test plant

Straw leachate




Amaranthus quitensis L.





Bidens subalternans L.





Brassica campestris L.





Lactuca sativa L.





Raphanus sativus L.





Sida rhombifolia L.





Sorghum bicolor (L.) Moench





Triticum aestivum L.





LD: The maximun inhibition of root elongation was less than 30%

The effect of sugarcane straw leachate and its isolated compounds were also tested in both autoclaved soil and sand. The leachate inhibit root growth in autoclaved sand more than in autoclaved soil, indicating that the solid soil phase adsorbed the leachate phytotoxins (Tongma et al. 1998). On the other hand, leachate constituents inhibit root growth in unsterile soil more than in autoclaved soil (Table 1) suggesting that soil microflora modified the leachate constituents to more toxic products.

Phenolic acids inhibited radicle growth in autoclaved sand more than in autoclaved soil suggesting that soil sorption reduced the availability of these compounds to the test plants. Unlike the straw leachate, phenolic acids reduced root growth in autoclaved soil more than in unsterile soil suggesting that microbial transformation and/or degradation products were inocuous to plant growth. The concentration of the isolated phenolic acids in a straw leachate (64 g dry straw/l) was estimated to be about 187 mg/l (ferulic acid), 131.2 mg/l (vanillic acid) and 19.82 mg/l (syringic acid).


We concluded that unsterilised soil treated with sugarcane straw leachate is able to inhibit plant growth at densities of sugarcane straw usually left in the field. Furthermore, the phenolic acids were also able to inhibit the growth of most of the test plants. However, the phytotoxic effect of these compounds is weak and can not completely explain the strong inhibitory activity of the straw leachate. Besides the identified phytochemicals, sugarcane straw should contain a number of other growth inhibitors and they need to be isolated and identified. Further studies to establish the concentration, fate and residence of the phenolic acids in sugarcane soils are in progress.


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