Peroxidase Fractions Research Articles

Peroxidases from grass clippings for the removal of phenolic compounds from wastewater

Green waste, especially of municipal origin, is currently used as a material only to a limited extent. However, the large material flows could also be used in a more economical way if they were integrated into biorefinery concepts. Besides the production of basic and fine chemicals, green waste could also be used as source of industrial relevant enzymes. Here, the purification and characterization of peroxidases from common lawn grass species Lolium perenne and Festuca arundinacea are reported. The purified peroxidase fractions as well as crude extracts were investigated for the removal of common wastewater pollutants such as phenol, m-cresol, and 2,4-dichlorophenol by oxidative polymerization. The grass-derived peroxidases showed the highest affinity towards 2,4-dichlorophenol, followed by m-cresol and phenol. A crude extract of real lawn grass was able to remove over 95 % of 0.5 mM 2,4-dichlorophenol within 20 min.

ROLE OF THE ANTIOXIDANT IN FUNCTIONING OF THE AGING MECHANISMS ON THE EXAMPLE OF BARLEY

Introduction. Despite close attention to the process of seed aging, there is still no unambiguous answer to questions about the mechanisms of aging. The solution of these issues is due to both economic needs and the need to store plant genetic resources. Aim. The aim was to study the effect of long-term storage of seeds of different barley varieties, previously differentiated by tolerance to seed aging, on the state of the antioxidant system enzymes. Methods. The studies were carried out on three varieties of barley (Hordeum vulgare L.) Rosava, Selena Star and Trudivnik which had been stored under non-specialized laboratory conditions in paper envelopes for 8 years. The seed viability was assessed by germination energy and germinating ability according to the state standards. The direct test was supplemented with a cytogenetic study: an anatelophase test. The condition of the antioxidant system was characterized by an electrophoretic method for studying nonspecific peroxidase and superoxide dismutase in germinated shoots. The number of multiple forms of enzymes, their relative electrophoretic mobility and specific gravity (share) in percent in the total spectrum were determined. Enzymatic activity was assessed by the area of the peaks of the corresponding multiple forms on the densitograms. Results. In terms of tolerance to long-term storage in direct and cytogenetic tests, the varieties were arranged as follows: Trudivnik > Rosava > Selena Star. It has been shown that the viability of seeds after storage is closely related to the activity of peroxidase and superoxide dismutase: more tolerant varieties are characterized by the stability of the total activity of peroxidase and superoxide dismutase. Conclusion. The specific activity of the medium-mobile peroxidase fraction and low- and medium-mobile SOD fractions plays a leading role in determining the viability of seeds during long-term storage.

Antioxidant activity of fractions isolated from hemolymph of garden snail Helix lucorum

Context: The snail hemolymph is a multi-component mixture comprising various substances with pharmacological activity. Aims: To evaluate the antioxidant activity of hemocyanin and fractions isolated of hemolymph from garden snail Helix lucorum. Methods: The antioxidant activity was tested in chemical systems generating superoxide radicals (O2•–) and hydroxyl radicals (•OH) and was presented as percent inhibition. The fractions’ composition was analyzed by MALDI-TOF/TOF mass spectrometry. Results: At concentration of 90 µg/mL the hemolymph fraction with compounds with molecular weight (MW) <100kDa demonstrated the strongest O2•–– scavenger effect (82%), followed by the hemocyanin (58%), and hemolymph fraction with compounds with MW<1kDa (15%). The most powerful •OH scavenger effect at concentration of 90 µg/mL showed hemocyanin (43%) followed by MW<100kDa fraction (26%). No •OH scavenger effect was observed by the MW<1kDa fraction. The highest chelation properties (55%) were demonstrated by the MW<100kDa fraction at concentration of 130 µg/mL; both the hemocyanin and MW<1kDa hemolymph fraction showed 33% inhibitory effect at the same concentration. The analysis of the composition of the tested fractions revealed high content of hydrophobic surfaces in MW<1kDa fraction and presence of antioxidant enzymes-catalase, superoxide dismutase and glutathione peroxidase in MW<100kDa fraction that contribute to the overall antioxidant effect of H. lucorum hemolymph. Conclusions: The complex antioxidant effect of hemolymph from H. lucorum is thought to be related to the presence of low molecular weight peptides in the MW<1 kDa fraction including glutathione and mainly to the antioxidant enzymes in the MW<100 kDa fraction.

Processing of pre-cooked frozen Brussels sprouts: Heat transfer modelling as related to enzyme inactivation and quality stability

The finite element numerical method was applied to simulate heat transfer processes (heating, cooling and freezing) in irregular domains during the production of precooked frozen Brussels sprouts; the effect of temperature on thermo-physical properties in the partial differential equation, and experimental heat transfer coefficients were considered. During precooking, heat transfer was coupled to the inactivation kinetics of thermo-labile and thermo-resistant fractions of peroxidase. The temperature distribution, and enzyme activity in the vegetable domain were modeled; these simulations were validated, obtaining a satisfactory agreement between the predicted values and: (i) the experimental time-temperature curves, (ii) the residual enzyme activity of peroxidase measured in the vegetable.The simulation of the time-temperature curve during the freezing process is a complex nonlinear problem that requires knowledge of the thermo physical properties changing with temperature. A change of variables based on Enthalpy and Kirchhoff formulations, was adopted for the numerical solution. The effects of precooking times (3,6,10min) on quality parameters of the frozen vegetable (Browing Index, textural properties, residual peroxidase activity, ascorbic acid content) were measured in the samples after freezing and storage during 4 months at −20°C. The pre-cooking time was optimized to get acceptable quality parameters avoiding peroxidase reactivation during frozen storage.

Chilling prior to low intensity pulsed electric field processing improved vitamin C stability of carrot purée (Daucus carotacv. Nantes)

Summary Effects of chilling (5 °C) prior to low intensity pulsed electric fields (PEF) (0.1–1.1 kV cm−1) on vitamin C and activity of peroxidase (POD) and ascorbic acid oxidase (AAO) in carrot puree were studied. The effect of PEF on carotenoids extractability was evaluated and compared to that of puree pre-conditioned at 20 °C (control). PEF enhanced carotenoids extractability for puree with and without pre-chilling, up to 25% and 66%, respectively. Vitamin C content decreased after PEF but combined pre-chilling and PEF maintained vitamin C stability and reduced AAO and POD activities. AAO and the heat-stable POD fraction were found to be more thermolabile (i.e. higher kref value) after PEF, particularly at 0.6 kV cm−1, which implies elimination of their enzymatic action towards oxidation of bioactive compounds. It was suggested that chilling prior to low intensity PEF modified enzyme properties and secured the stability of vitamin C and carotenoids.

Impact of weak permanent magnetic field on antioxidant enzyme activities in radish seedlings

The impact of weak permanent magnetic field (PMF) with magnetic flux density of 185–650 μT on activities of antioxidant enzymes in 5-day-old radish (Raphanus sativus L. var. radicula D.C.) seedlings, cv, Rosovo-krasnyi s belym konchikom was demonstrated. In the range of 185–325 μT PMF suppressed superoxide dismutase (SOD) activity in seedling grown in darkness and catalase (CAT) grown in both darkness and light. At the same values of magnetic flux density, all fractions of guaiacol peroxidase were activated and MDA accumulation was enhanced. At the higher values of magnetic flux density, SOD was activated in seedlings grown in darkness and CAT was activated in seedlings grown in both darkness and light. Other indices decreased or remained unchanged. It was concluded that PMF action depends on its intensity: at its low values, antioxidant enzymes are inhibited and at high values — activated.

Prevalence of, risk factors for, and oxidative stress associated with Toxoplasma gondii antibodies among asymptomatic blood donors in Egypt

Methods Blood donors were recruited (169 men and 61women) from blood banks, Mansoura University Hospital, Egypt. We interviewed blood donors about sociodemographic characteristics and potential risk factors for T. gondii infection using a structured questionnaire. A venous blood sample was taken to document their T. gondii antibody status using enzyme-linked immunosorbent assay (ELISA). Also, serum level of malondialdehyde (MDA) and activity of glutathione peroxidase (GSH-Px) and tocopherol fractions (a, g, δ) was assessed.

Different peroxidase activities and expression of abiotic stress-related peroxidases in apical root segments of wheat genotypes with different drought stress tolerance under osmotic stress

One-week-old seedlings of Triticum aestivum L. cv. Plainsman V, a drought tolerant; and Cappelle Desprez, a drought sensitive wheat cultivar were subjected gradually to osmotic stress using polyethylene glycol (PEG 6000) reaching 400mOsm on the 11th day. Compared to controls cv. Plainsman Vmaintained the root growth and relative water content of root tissues, while these parameters were decreased in the drought sensitive cv. Cappelle Desprez under PEG-mediated osmotic stress. Simultaneously, H(2)O(2) content in 1-cm-long apical segment of roots comprising the proliferation and elongation zone, showed a transient increase in cv. Plainsman V and a permanent raise in cv. Cappelle Desprez. Measurements of the transcript levels of selected class III peroxidase (TaPrx) coding sequences revealed significant differences between the two cultivars on the 9th day, two days after applying 100 mOsm PEG. The abundance of TaPrx04 transcript was enhanced transitionally in the root apex of cv. Plainsman V but decreased in cv. Cappelle Desprez under osmotic stress while the expression of TaPrx01, TaPrx03, TaPrx19, TaPrx68, TaPrx107 and TaPrx109-C decreased to different extents in both cultivars. After a transient decrease, activities of soluble peroxidase fractions of crude protein extracts rose in both cultivars on day 11, but the activities of cell wall-bound fractions increased only in cv. Cappelle Desprez under osmotic stress. Parallel with high H(2)O(2) content of the tissues, certain isoenzymes of covalently bound fraction in cv. Cappelle Desprez showed increased activity suggesting that they may limit the extension of root cell walls in this cultivar.

Inactivation kinetics of food enzymes during ohmic heating

Ohmic heating of milk and fruit and vegetable juices was carried out at several incubation temperatures to investigate inactivation of alkaline phosphatase, pectin methylesterase and peroxidase. Mechanisms of inactivation of these enzymes and corresponding kinetic models were verified for each food material, using the multitemperature evaluation of inactivation data. Compared to inactivation by conventional indirect heating, kinetic parameters were changed but inactivation mechanisms remained the same. The kinetic parameter changes were relatively minor for pectin methylesterase and alkaline phosphatase. A significant destabilization of the labile isozyme fraction of peroxidase occurred by the effect of ohmic heating when the greatest decrease of stability was obtained for carrot juice.

Discrimination of Pinus eldarica Medw. and its two new species by epicuticular wax, lignin content, electrophoretic isozyme and activity of peroxidase

AbstractMondell pine (Pinus eldarica Medw.) is a rare and drought resistant pine, found naturally only in semi desert environment southeast of Tbilisi, Georgia, and has been probably introduced to Iran more than 800 years ago. This species has gradually altered both in shape and growth rate in Nashtifan‐Khaf, and generated two species nova which so called as ball shaped and conical, according to characteristics of their shape and smaller stature. This study was conducted to discriminate these species nova using epicuticular wax features and certain biochemical characteristics i.e., peroxidase electrophoretic pattern (PAGE), the activity of soluble (SPO), ionically (IPO) and covalently (CPO) wall‐bound fractions of peroxidase, lignin and total protein content. Observation via scanning electron microspopy showed differences that in adaxial and abaxial surface of needles, and also seed surface, in ball shaped type with the others. But none of the biochemical characteristics had significant difference in these three types. The electrophoretic analysis showed that the peroxidase isozyme is not suitable marker for discriminating the species nova from wild type, in spite of obvious micro‐ and macromorphological differences. Results of current study, propose the use of molecular markers and cytogenetic studies in order to specify cause of species nova occurrence in the study area. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Avaliação da termoestabilidade e da regeneração da atividade da peroxidase extraída de laranja (Citrus spp.)

Soluble and ionical-bound Peroxidase (EC 1.11.1.7) was extracted from orange cultivars Citrus sinensis (L.) Osbeck and Citrus aurantium (Rutaceae). Oranges were washed with distilled water and cut into small pieces for extract preparation; they were then homogenized with sodium phosphate buffer 100mM and pH 6.0, by a liquidizer. Afterwards, the extract underwent heat treatment at 70oC, 75oC and 80oC, and the peroxidase activity measured at 1 to 10 minutes. Whereas fractions with soluble peroxidase in both cultivars were more heat stable, those with ionical-bound peroxidase were less heat-stable. Total inactivation was not achieved with the heat treatments used. A regeneration of 15% of the initial activity was observed for fractions with soluble peroxidase. No regeneration was observed in the ionical-bound peroxidase fraction. This seems to show that regeneration may be linked to different isoenzymes with different heat stability

Peroxidase activity can dictate the in vitro lignin dehydrogenative polymer structure

The objective of this study was to assess the influence of the peroxidase/coniferyl alcohol (CA) ratio on the dehydrogenation polymer (DHP) synthesis. The soluble and unsoluble fractions of horseradish peroxidase (HRP)-catalyzed CA dehydrogenation mixtures were recovered in various proportions, depending on the polymerization mode (Zutropf ZT/Zulauf ZL) and HRP/CA ratio (1.6–1100 purpurogallin U mmol −1). The ZL mode yielded 0–57%/initial CA of unsoluble condensed DHPs (thioacidolysis yields <200 μmol g −1) with a proportion of uncondensed CA end groups increasing with the HRP/CA ratio (7.2–55.5%/total uncondensed CA). Systematically lower polymer yields (0–49%/initial CA) were obtained for the ZT mode. In that mode, a negative correlation was established between the β-O-4 content (thioacidolysis yields: 222–660 μmol g −1) and the HRP/CA ratio. In both modes, decreasing the HRP/CA ratio below 18 U mmol −1 favoured an end-wise polymerization process evidenced by the occurrence of tri-, tetra- and pentamers involving at least one β-O-4 bond. At low ratio, the unsoluble ZT DHP was found to better approximate natural lignins than DHPs previously synthesized with traditional methods. Besides its possible implication in lignin biosynthesis, peroxidase activity is a crucial parameter accounting for the structural variations of in vitro DHPs.

The activity of peroxidase in various cell fractions of wheat plants infected with Septoria nodorum berk.

The activities of peroxidase isoforms and hydrogen peroxide content in leaf cuttings of wheat (Triticum aestivum L., cv. Diamant) resistant to Septoria blotch were studied during aging and following the infection with Septoria nodorum Berk. The differential activation of peroxidase isoforms was regulated by hydrogen peroxide level in the tissue. At early stages of fungus development in plant tissues, the decrease in the activities of soluble, membrane and ion-bound fractions of peroxidase elevated the level of hydrogen peroxide in infected tissues and rapidly activated peroxidase isoforms in infected tissues as compared to the aging ones even before disease symptoms appeared. The anionic peroxidases, which were first to respond to the pathogen, seem to stand for wheat resistance to fungal infections and the protection of leaf tissues from oxidative stress.

Monolignol oxidation by xylem peroxidase isoforms of Norway spruce (Picea abies) and silver birch (Betula pendula)

We partially purified peroxidase isoform fractions from xylem extracts of a gymnosperm, Norway spruce (Picea abies (L.) Karst.), and an angiosperm, silver birch (Betula pendula Roth.), to determine the participation of xylem-localized peroxidases in polymerization of different types of lignin in vivo. Several peroxidase fractions varying in isoelectric point values from acidic to basic were tested for their ability to catalyze the oxidation of the monolignols coniferyl alcohol, sinapyl alcohol and p-coumaryl alcohol in vitro. All of the xylem peroxidases extracted from Norway spruce and most of those from silver birch showed the highest rate of oxidation with coniferyl alcohol in the presence of hydrogen peroxide. The exception was an acidic peroxidase fraction (pI 3.60-3.65) from silver birch that exhibited higher oxidation activity for sinapyl alcohol than for coniferyl alcohol. For the xylem enzyme fractions extracted from silver birch, the ability to oxidize the artificial phenolic substrate syringaldazine coincided with high specific activity for sinapyl alcohol. Therefore, we conclude that the acidic, neutral and basic xylem peroxidases of Norway spruce all function in the synthesis of guaiacyl-type lignin, whereas in silver birch the acidic peroxidases preferentially oxidize sinapyl subunits. The latter provides a mechanism for synthesis of guaiacyl-syringyl lignin typical of tracheid cell walls in angiosperm trees.

Quantitative and qualitative aspects of peroxidases extracted from cladodes of Opuntia ficus indica

Peroxidases (EC 1.11.1.7) were extracted from fresh cladodes harvested from nine ecotypes of the cactus species Opuntia ficus indica Mill. growing as a collection, in Marrakech (South Morocco). Two enzyme fractions were obtained by a progressive solubility method leading to soluble peroxidases (S) and ionically wall-bound peroxidases (I). The preferred substrate for cladode peroxidases was determined to be o-dianisidine over 4-chloro-1-naphtol and guaiacol. Contrarily to roots, no guaiacol-based activity was found in cladodes. The late ecotypes 1‘Haddaouia’ and ‘Moussa’ showed a relatively high peroxidase ratio S/I (units g −1 fresh weight). When subjected to electrophoresis on polyacrylamide gels, S and I peroxidase fractions each exhibited two enzyme forms based on their electric charges in basic and acidic gel media. Acidic peroxidase forms, well separated on basic gels, showed two principal migration zones with great differences in their enzyme activities depending on the fraction types S and I. Acidic ionically wall-bound peroxidases exhibited fast, highly active isoforms with R f values 0.42–0.58. Basic forms, represented essentially in fractions S and resolved on acidic gels, were typified by slow and fast isoforms with a double banded pattern in most ecotypes. Opuntia ecotypes collected from localities surrounding Marrakech exhibited fast basic soluble isoforms, while distant ecotypes from Marrakech, including late ones, were typified by a fast acidic ionically bound isoperoxidase of R f 0.58. Possible roles of O. f. indica peroxidases in growth and their evaluation as markers in this cactus species are discussed in this study.

Kinetics and inactivation of carrot peroxidase by heat treatment

The activity and kinetics of carrot peroxidase were determined by using pyrogallol, guaiacol and o-dianisidine as hydrogen donors and peroxidase inactivation was studied by thermal and microwave heating. The kinetics of peroxidase showed characteristics which were dependent upon the identity and concentration of the hydrogen donor used. With pyrogallol and guaiacol, the true K m was found to be 0.34 and 1.4 mM for hydrogen peroxide, respectively, whereas the apparent K m with o-dianisidine was 7.7 × 10 −3 mM. The lowest K m and highest V max/ K m with o-dianisidine exhibited the greater tendency of the enzyme toward hydrogen peroxide and the specificity of the competing substrate, o-dianisidine. Thermal treatment of carrot peroxidase was done in the range of 35–75 °C for 0.5–180 min. Inactivation kinetics of peroxidase showed a biphasic first-order model, while at 75 °C, peroxidase showed monophasic first-order behaviour. Kinetic parameters, k and E a, were determined for heat labile and heat resistant fractions of peroxidase. Biphasic behaviour of enzyme inactivation was observed for the microwave treatment at 70 and 210 W, whereas at 350 and 700 W microwave powers enzyme inactivation was monophasic. Microwave heating was found to be more effective for inactivating the enzyme than thermal treatment and additionally vitamin C retention was higher in microwave treated samples compared to heat treatment.

Ecophysiological approach to study of the role of cell-bound peroxidase forms in wheat leaves of different morphology

Extracellular peroxidase enzymes, both ionically - and covalently bound, have been implicated in the biochemical changes of cell wall and furthermore for regulation of leaf and root extension growth. However, the role of cell wall bound peroxidase in changes of mature leaf morphology is poorly understood. The aim of this study was to determine whether ionically and covalently bound peroxidase activities exhibit any relationships with drought-induced leaf rolling of wheat. An activity and thermostability of three wheat genotypes with different leaf rolling degree were studied. The results clearly show that two genotypes with leaf rolling trait exhibited significantly higher activity of both extracellular peroxidase fractions in comparison to non-rolling cv. 'Omskaya 9'. In addition, the significant genotypic differences were found in thermostability of ionically and covalently bound peroxidase. A 35 °C-treatment decreased the activity of ionically bound peroxidase (IBP) in non-rolling leaves of parent cultivar and enhanced the activity of IBP in rolling leaves of 'Alba' genotype. Under 45 and 55 °C-treatment the IBP was more stable in rolling leaves of both genotypes 'Otan' and 'Alba'. Despite to low activity of covalently bound peroxidase it was more stable after heating by 45 °C compare with ionically bound fraction. The heating of covalently bound peroxidase (CBP) extracts by 35 °C and 45 °C also has enhancing effect on its activity by both leaf rolling genotypes (on 16 and 40% by 'Otan' and 'Alba', respectively). At the same time the CBP activity of non-rolling genotype decreased significantly on 40%. Even after 55 °C-treatment the CBP of both rolling genotypes remained very stable, although there were further inhibition of CBP purificated from leaves of non-rolling genotype. Both wheat genotypes with inserted leaf rolling trait exhibited a much higher blue-green fluorescence emission than non-rolling leaves of parent cultivar 'Omskaya 9'. Thus, the molecular mechanism of leaf rolling regulation seems to include an adjustment of cell wall properties and probably involves an activity of cell wall bound peroxidase. Further investigation is expected to reveal the regulatory mechanism for leaf rolling in wheat.

Peroxidase from oranges ( Citrus sinenses (L.) Osbeck

Soluble peroxidase was extracted from oranges (Citrus sinenses (L.) Osbeck). Small amounts of purified anionic and cationic isoperoxidases have been obtained by gel filtration and ion-exchange chromatography. It has been shown that peroxidase activity present in crude extracts of oranges (juice, albedo, and peel) is less stable to heat than the enzymes activity of highly purified individual orange isoperoxidases. For the purified isoperoxidases heat-inactivation is still non-linear. It is suggested that this may be due to micro heterogeneity in covalently bound oligosaccharide residues at the molecular level. The higher activity was detected in the crude extract from the peel, around 3.2 and 7.8 times more than the fractions from the albedo and juice respectively. The crude soluble peroxidase fractions were particularly heat stable. The pI (4.5, 5.2, and 9.0) of the purified isoperoxidase (A1, A3, and C1) from the peel soluble fraction was measured using a surface electrode and the molecular weight (42 kDa, 30 kDa, and 26 kDa) was estimated by gel-filtration.

Diversity of leaf peroxidases in date palm ( Phoenix dactylifera L.) as revealed in an example of marginal (seedling derived) palm groves

Peroxidases (EC 1.11.1.7) were extracted from fresh leaves of date palm ( Phoenix dactylifera L.) harvested from 90 non-fruit-producing male trees and 75 adult female plants growing all in the Marrakech (South Morocco) marginal palm grove of unknown seedling origin. Two enzyme fractions were obtained by a progressive solubility method leading to soluble peroxidases (S) and ionically wall-bound peroxidases (I). When subjected to electrophoresis on polyacrylamide gels, S and I peroxidase fractions exhibited each one, two enzyme forms based on their electric charges occurred in basic and acidic gel media. Acidic peroxidase forms, well separated on basic gels, showed two migration zones with a maximum of nine isoperoxidases. Basic forms, more represented in fractions I and resolved on acidic gels, were typified by two major isoforms. When combining patterns of acidic and basic isoperoxidases, enzyme fractions S and I of individual trees, showed, respectively, 10 and nine isoenzyme phenotypes. No sex-specific isoforms were found at the electrophoresis-based qualitative study. However, some phenotypes were found to be highly represented in male trees. Evaluation of peroxidases as markers in date palm is discussed in this study.

Removal of Phenol by Alfalfa Plants (Medicago sativa L.) Grown in Hydroponics and its Effect on Some Physiological Parameters

The plant-assisted removal of phenol, with special emphasis on the effects of this compound on some plant's physiological parameters, was investigated. Hydroponic cultures of alfalfa (Medicago sativa L., var. Romagnola) were employed as a model system. These cultures were exposed to two phenol concentrations: 100 and 500 mg/l. A first order kinetic approach was used to describe the removal of phenol from the solution. After 30 days of cultivation, the initial amount of phenol (100 mg/l) was reduced to non-detectable levels in the presence of plants. In the absence of plants, 20% of phenol remained in the solution. The half-life of phenol was reduced from 7.2 to 4.5 days in the presence of plants. After 25 days, the initial amount of 500 mg/l of phenol was reduced to non-detectable levels in the presence of plants not previously exposed to phenol and to approximately 20% with plants previously exposed to the contaminant. In the absence of plants, almost 40% remained in the solution. The presence of plants reduced the half-life of phenol from 18.3 days to 10.4 in the case of plants previously exposed and to 7.8 days in the case of plants without previous contact. Chlorophyll contents in alfalfa leaves of plants exposed to 100 mg/l of phenol were similar to those of control plants and a decrease in total chlorophyll content was observed when plants were exposed to 500 mg/l of phenol. The activity of soluble peroxidases of the roots increased in the presence of 100 mg/l of phenol but the amount of 500 mg/l had a negative effect on the peroxidase fraction. No changes were observed in the case of the ionically-bound cell wall fraction. The growth index of the plants exposed to 100 mg/l of phenol was comparable to that of non-exposed plants, while this parameter was negatively affected in the case of plants exposed to 500 mg/l of phenol. Although alfalfa plants were able to survive an exposure to 500 mg/l of phenol, their physiological parameters and their removal capacity were negatively affected.