Ni PHYTOACCUMULATION IN MENTHA AQUATICA L. ANDMENTHA SYLVESTRIS L.

RAMI ZURAYK∗, BISHR SUKKARIYAH, RIAD BAALBAKI andDAAD ABI GHANEM

Faculty of Agricultural and Food Sciences, American University of Beirut, Riad El Solh, Beirut,Lebanon

(∗ author for correspondence, e-mail: rzurayk@aub.edu.lb, fax: 961 1 744460)

(Received 22 January 2001; accepted 22 October 2001)

Abstract. Ni phytoaccumulation in Mentha aquatica L. and Mentha sylvestris L., two hydrophytesthat had shown high Ni accumulation potential in previous screenings, was evaluated for possible in-clusion in aquatic phytoremediation systems. The plants were grown for 14 days in full-strength (FS)and half-strength (HS) Hoagland’s solutions spiked with 1, 2, 4, and 8 mg L−1 Ni. Roots and shootswere harvested and analysed for Ni. Ni accumulation and bioconcentration factor (BCF) generallyincreased with increasing external Ni levels, except for shoot Ni in HS solution at 8 mg L−1, whichwas lower than at 4 mg L−1. We suggest that a mechanism may exist to control Ni entry to the shootat high external Ni levels in order to reduce injury. Ni phytoaccumulation and BCF were consistentlylarger in the HS solution, probably resulting from interaction between Ni and other nutrients, andindicating that manipulation of the nutrient concentration is a possible avenue for improving theefficiency of aquatic phytoremediation. Due to their large growth rates and biomass, both speciesaccumulated Ni in the roots to levels equal or superior to those of known phytoremediator plants(8327 for M. aquatica and 6762 mg kg−1 for M. sylvestris), indicating their adequacy for use inphytoremediation systems.

Keywords: accumulation, aquatic phytoremediation, Mentha aquatica L., Mentha sylvestris L.,nickel

1. Introduction

Aquatic phytoremediation involves the use of plants for the removal of contam-inants, such as heavy metals, from aqueous solutions. This may take place inconstructed wetlands (Terry and Bañuelos, 2000) or in purpose-built filtration unitsin which metals are removed by plant roots, a process also known as rhizofiltration(Dushenkov et al., 1995). In both approaches, plant biomass is harvested, driedand ashed for disposal. Metal accumulators, but not root-shoot translocators, arepreferable in aquatic phytoremediation, as efficient translocators produce pollutedabove ground biomass, which increases the cost of processing, as well as the riskof exposing wildlife to the contaminated plants.

A cursory review of the current literature (Raskin and Ensley, 2000; Terry andBañuelos, 2000) reveals that the desirable attributes of a plant for use in aquaticphytoremediation include: (i) tolerance to high environmental concentrations of

Water, Air, and Soil Pollution 139: 355–364, 2002.© 2002 Kluwer Academic Publishers. Printed in the Netherlands.

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the heavy metal, (ii) ability to accumulate very high levels of the metal, (iii) abilityto bioconcentrate the metal in low-concentration environments, (iv) fast growthand high biomass production, (v) limited root to shoot translocation, especiallyfor plants used in rhizofiltration, and (vi) ability to phytoaccumulate metal acrossa broad range of nutrient levels. Moreover, for constructed wetlands, site-specificrequirements include (i) the ability to sustain the desirable characters across a broadrange of climatic conditions, and (ii) endemism or indigenous origin, in order toavoid the introduction of foreign wetland species, which can often become noxiousaquatic weeds (e.g. water hyacinth).

Most of the research addressing the phytoremediation of heavy metal contam-ination has focused on the use of hyperaccumulator plants for metal removal, suchas Alyssum bertolonii, for instance, which is able to accumulate large quantitiesof Ni in its leaf tissue (Bennett et al., 1998). Plants such as Thlaspi caerulescensare possibly more appropriate for soil remediation, as they translocate significantquantities of metal to their shoot, although some limitations apply to their use(Brown et al., 1995). Besides the risk of heavy metal transfer to the food chain,hyperaccumulator plants are often characterized by a relatively slow growth rateand low biomass accumulation (Peterson, 1971). For instance, Thlaspi rotundi-folium L. accumulated only 5–50 mg/plant dry tissue after 5 months of growth(Cunningham and Ow, 1996). While results from initial research were promising,their slow growth rate and small biomass accumulation may limit their uses.

There has recently been a shift in research focus towards the screening of highbiomass accumulating plants, mainly crop and crop related species, for heavy metalaccumulation. It is suggested that high biomass accumulating species, even if theyare moderate accumulators, such as Indian mustard (Brassica juncea L.) may bemore effective than hyperaccumulators with lower biomass. Indeed, when B. junceaand Zn hyperaccumulator Thlaspi caerulescens L. were grown for a 6 weeks periodin a Zn contaminated soil (>11,000 mg kg−1 soil), B. juncea removed 4 timesmore Zn due to a 10-fold difference in biomass accumulation (Ebbs et al., 1997).Dushenkov et al. (1995) recommend the use of terrestrial plant roots to removethe metal from solution because they develop a much longer fibrous root system,which creates an extremely high surface area. Thus, sunflower (Helianthus annuusL.) as well as various grasses were found to efficiently remove toxic metals suchas Cu, Cd, Cr, Ni, Pb, and Zn from aqueous solution. One limitation, however, isthe low tolerance of these plants to elevated tissue levels of heavy metals, whichresults in their rapid death upon exposure to heavy metals.

Wetland plants, such as water hyacinth (Eichhornia crassipes L.) (Zhu et al.,1999), coontail (Ceratophyllum demersum L.), wild rice (Hygroryza aristata) (Raiet al., 1995), as well as salvinia (Salvinia natans) (Sen and Bhattacharyya, 1994)have shown to be effective in the removal of a broad range of heavy metals suchas Ni, Cr and Cd from contaminated water with relatively little reduction in bio-mass. Their advantage over terrestrial plants is their ability to thrive in aquaticenvironments with no special management requirements such as oxygenation. In a

Ni PHYTOACCUMULATION IN HYDROPHYTES 357

recent report, Qian et al. (1999) have demonstrated the ability of 12 wetland plantspecies to accumulate Mn, Cd, Cu, Ni, Cr, Pb, Hg, B, As, and Se to various extentsfrom nutrient solution supplied with 1 mg L−1 of the element. Highest shoot tissueconcentrations (mg kg−1) were attained for Cr (2980) and Pb (1882) by Polygonumhydropiperoides Michx, for B (1132) by Hippuris vulgaris L., and for Cd (148) andNi (80) by Wedelia trilobata Hitchc.

Our research group has recently evaluated the ability of 12 hydrophytes col-lected from the wild in Lebanon to phytoaccumulate Ni, Cd and Cr from nutrientsolutions spiked with 1 mg L−1 of each heavy metal (Zurayk et al., 2001). Thephytoaccumulation of Ni in the whole plant tissue ranged between 4.5 mg kg−1

and 235 mg kg−1. Among the best Ni accumulators were Mentha sylvestris L.(235 mg kg−1) and Mentha aquatica L. (186 mg kg−1). Both plants are facultativehydrophytes, high accumulators, produced high biomass and were also poor root-to-shoot translocators. In this article, we report on a second stage in the screeningin which we: (1) determine the concentration-dependent growth response of M.sylvestris and M. aquatica, (2) assess the effect of increasing Ni levels in solutionon metal phytoaccumulation, bioconcentration and removal for both plant species,and (3) evaluate the effect of nutrient concentration on the Ni phytoremediationpotential.

2. Materials and Methods

Twelve transplants of similar sizes and weight (±1 g) of each plant species weregrown in continuously aerated container batch system consisting of 10 L-polyethy-lene containers covered with aluminum foil to prevent algal growth. Two nutrientlevels were used: full-strength (FS) and half-strength (HS) Hoagland’s nutrientsolution, both at pH 6. The FS culture solution contained N: 56 mg L−1; P: 15.5 mgL−1; K: 58.75 mg L−1; Ca: 40 mg L−1; Mg: 6 mg L−1; S: 8 mg L−1; B: 0.27 mgL−1; Cl: 1.77 mg L−1; Mn: 0.11 mg L−1; Cu: 0.032 mg L−1; Zn: 0.131 mg L−1;Mo: 0.05 mg L−1 and Fe: 1.12 mg L−1. Nickel (as NiCl2·6H2O) was supplied tothe medium at four different concentrations: 1, 2, 4, and 8 mg L−1. Plants grown insolution without added Ni served as controls. Factorial treatments were combinedin a completely randomized design with three replicates per treatment. The culturesolutions (4 L per container) were replaced every 2 days.

The set-up was placed in a greenhouse in Beirut during the month of June(min T = 23 ◦C, max T = 39 ◦C). The plants were monitored for a period of 14days during which signs of deficiency or toxicity were recorded. They were thenharvested, their shoots and roots separated and rinsed thoroughly with distilledwater, blotted dry, and weighed for fresh weight. Plants were then oven dried at70 ◦C for 48 hr for dry weight measurement. Dried plant samples were ground topass a 20-mesh sieve and representative samples were taken for chemical analysis.Elemental analysis was carried out by acid digestion of dry samples (Zarcinas et

358 R. ZURAYK ET AL.

Figure 1. Ni accumulation in the roots of M. aquatica and M. sylvestris grown at 1, 2, 4, and 8 mgL−1 of Ni in full and half strength Hoagland’s solutions.

al., 1987). Elemental content was determined in the acid digest using an ICP-MS(Hewlett Packard 4500 series). Unless otherwise indicated, all tissue trace elementconcentrations are reported on dry weight basis throughout the manuscript. Thebioconcentration factor was calculated as follows:

BCF = Ni concentration in plant tissues (mg kg −1) at harvest/Initial concen-tration of the element in the external nutrient solution (mg L−1).

Total accumulation rate (mg kg−1 d−1) was calculated as follows (concentrationin mg kg−1 dry wt., biomass in g dry wt.):

(Shoot concentration × Shoot biomass + Root concentration × Root biomass)/[(Shoot biomass + Root biomass) × 14].

Statistical analysis of variance was performed using the MSTATC statistical pack-age version for Personal Computers. Where appropriate, treatment means wereseparated using Duncan’s multiple range test (p ≤ 0.05).

Ni PHYTOACCUMULATION IN HYDROPHYTES 359

Figure 2. Ni accumulation in the shoots of M. aquatica and M. sylvestris grown at 1, 2, 4, and 8 mgL−1 of Ni in full and half strength Hoagland’s solutions.

3. Results and Discussion

In both the FS and the HS solutions, biomass accumulation by both species wasunaffected by the presence of Ni at all levels used in this experiment. The twospecies grew to similar sizes, and reached a mean root dry weight of 0.3 g and amean shoot dry weight of 0.7 g. Leaf chlorosis was recorded after 8 days in thehighest Ni treatment (8 mg L−1). There were no toxicity symptoms on the roots.These results indicate the potential tolerance of both Mentha species to Ni, andthat any variation in phytoaccumulation will be due to experimental treatments(solution Ni concentration and nutrient levels) rather than to growth inhibition.

Phytoaccumulation in the roots of the 2 species increased in response to in-creasing external Ni levels (Figure 1). In both species, shoot phytoaccumulationpeaked at 4 mg L−1 and then declined again (Figure 2). Translocation of Ni tothe shoots was limited, and shoot Ni values were often 10–20 folds lower than thecorresponding root values. Phytoaccumulation in both shoots and roots was con-sistently reduced by increasing nutrient levels. This effect was more pronounced athigh external Ni. At the highest Ni concentration in the HS solution, M. aquaticaaccumulated more Ni than M. sylvestris on a whole plant basis (Figure 3) (2925 mg

360 R. ZURAYK ET AL.

Figure 3. Ni accumulation in plants of M. aquatica and M. sylvestris grown at 1, 2, 4, and 8 mg L−1

of Ni in full and half strength Hoagland’s solutions.

kg−1 and 2050 mg kg−1), due to a larger root accumulation (8327 mg kg−1 in M.aquatica and 6762 mg kg−1 in M. sylvestris).

Shoot BCF (Figure 5) increased with increasing Ni levels, except at 8 ppm Ni inthe HS solution. Total plant BCF (Figure 6) generally increased with increasing Nilevels, except for M. aquatica in the FS solution. It was larger in the HS solution,reaching maximal values of 365 in M. aquatica and 256 in M. sylvestris. In aquaticphytoremediation, root BCF is more relevant than whole plant or shoot BCF. RootBCF in the HS solution was very high in both species, reaching 1040 in M. aquaticaand 845 in M. sylvestris at the 8 ppm Ni treatment (Figure 4).

The recovery factor is the total amount of Ni removed from the solution bythe plants during the experimental period. Recovery was maximal at 8 mg L−1

reaching 22 mg Ni per plant or 1.59 mg kg−1 d−1 for M. aquatica, and 17.9 mg Niper plant or 1.28 mg kg−1 d−1 for M. sylvestris. Total recovery in the HS solutionwas double that in the FS solution except in the 1 mg L−1 treatment (Table I).

Wetland plants vary in their response to elevated levels of Ni. In this experiment,M. aquatica and M. sylvestris appear to be tolerant to high levels of Ni, and themaximum concentration used (8 mg L−1) was below the tolerance threshold. Ni tol-erance appears to be associated with the ability to accumulate large concentrationsof Ni in the roots, and to control translocation to the shoot. Although preferential

Ni PHYTOACCUMULATION IN HYDROPHYTES 361

Figure 4. Bioconcentration factor of Ni in the roots of M. aquatica and M. sylvestris grown at 1, 2,4, and 8 mg L−1 of Ni in full and half strength Hoagland’s solutions.

TABLE I

Recovery of Ni in whole plants of M. aquatica and M. sylvestris grown at1, 2, 4, and 8 mg L−1 of Ni in full and half strength Hoagland’s solutions

Concentration Recovery (mg kg−1)a

(mg L−1) M. aquatica M. sylvestris

Full-strength 1 2.2 b 1.3 c

2 2.3 b 1.9 bc

4 4.3 b 5.2 b

8 8.7 a 11.9 a

Half-strength 1 1.7 c 0.9 c

2 4.1 c 3.8 c

4 13.7 b 9.4 b

8 22.3 a 17.9 a

a Means within a column followed by similar letters are not significantlydifferent according to Duncan’s multiple range test (p ≤ 0.05).

362 R. ZURAYK ET AL.

Figure 5. Bioconcentration factor of Ni in the shoots of M. aquatica and M. sylvestris grown at 1, 2,4, and 8 mg L−1 of Ni in full and half strength Hoagland’s solutions.

Figure 6. Bioconcentration factor of Ni in plants of M. aquatica and M. sylvestris grown at 1, 2, 4,and 8 mg L−1 of Ni in full and half strength Hoagland’s solutions.

Ni PHYTOACCUMULATION IN HYDROPHYTES 363

root accumulation of Ni in wetland plants has been shown earlier (Sen and Bhat-tacharyya, 1994), the associated growth response differs among species. Zhu et al.(1999) for instance, showed a significant decrease in biomass accumulation of E.crassipes in the presence of high concentrations of Ni, associated with intermediatephytoaccumulation in both root (1200 mg kg−1) and shoot (120 mg kg−1) at 10 mgL−1 external Ni. These findings were similar to those of Turnquist et al. (1990),and Zaranyika and Ndapwadza (1995), indicating the sensitivity of E. crassipes toNi. In another wetland plant, duckweed (Lemna minor L.), growth was not reducedby 10 mg L−1 Ni, and whole plant accumulation reached a maximum of 1790 mgkg−1. In the same report, increasing Ni concentrations from 2 mg L−1 to 10 mgL−1 resulted in larger phytoaccumulation and in a considerable increase in BCFfor Ni. It may be deduced that the ability to tolerate high Ni levels appears to beassociated with an increase in phytoaccumulation and in BCF with increasing Niconcentrations. This response is typical of accumulators (Baker, 1981), in whichelemental content of the tissue increases linearly with increasing concentration ofthe element in the substrate until a plateau is reached at a relatively high metalcontent in the solution. Non-accumulators usually restrict entry of the metal atlow concentrations of the element in the substrate until the restriction mechanismbreaks down and there is an unlimited absorption at quite a small concentrationrange in the medium, following which the plant usually dies. In this case, however,both Mentha species restrict shoot entry, which indicates that they are not metallo-phytes, but that they have evolved a mechanism which allows them to restrict shootentry of Ni, thereby protecting vital metabolic processes, while accumulating themetal in the root.

High nutrient concentrations appear to reduce the ability of both species toaccumulate Ni. The interaction between Ni and other trace elements has beendemonstrated. Cataldo et al. (1978) showed that Ni uptake by soybean seedlingsand its transfer from shoot to root was inhibited by the presence of Cu2+, Zn2+ andFe2+, due to competition for the same transport mechanism.

4. Conclusion

The data indicates that both M. aquatica and M. sylvestris appear to be very goodcandidates for aquatic phytoremediation of Ni. They are tolerant to elevated levelsof the metal, efficient bioconcentrators and accumulators at a wide range of externalNi concentrations and of nutrients. They are also fast growing and produce largebiomass. They are widespread species and are well adapted to climatic variations.Their performance was generally better than that of water hyacinth and duckweed,both of which are commonly used wetland plant species. Our data also shows theirpotential for inclusion in a rhizofiltration-type system, as their BCF for Ni (113–365) compares favorably with that reported for B. juncea grown at 10 mg L−1 Niin a rhizofiltration system (BCF = 208) (Dushenkov et al., 1995).

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