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Copper, Manganese and Zinc Bioaccumulation in Some CommonPoaceae Species Along Romanian Black Sea Coast

BOGDAN STEFAN NEGREANU PIRJOL1, TICUTA NEGREANU PIRJOL1*, DAN RAZVAN POPOVICIU2

1 Ovidius University of Constanta, Faculty of Pharmacy, 1 University Alley, Campus, Corp B, 900470, Constanta, Romania2 Ovidius University of Constanþa, Faculty of Natural Sciences and Agricultural Sciences, 1 University Alley, Campus, Corp B,900470, Constanta, Romania

Copper, manganese and zinc bioaccumulation potential was screened for three common Poaceae species,Hordeum murinum ssp. murinum L., Leymus racemosus ssp. sabulosus (M. Bieb.) Tzvelev and Lolliumperenne L., abundantly growing along the Romanian Black Sea Coastal area, Constanta County. Theconcentration of the selected heavy metals was analyzed though atomic absorption spectrometry in samplesof aboveground plant organs and soils. To assess the potential for phytoaccumulation, the BiologicalAccumulation Coefficients (BAC) were calculated. Considering the minimal thresholds widely accepted inliterature for Cu, Mn and Zn hyperaccumulators, none of the selected species can fit in this category. Thehighest average copper concentration was found in Hordeum murinum ssp. murinum L. (5.45 mg/kg). Formanganese, the highest value was found in Lollium perenne L. (104.08 mg/kg), while for zinc, the maximumwas reached in Leymus racemosus ssp. sabulosus (M. Bieb.) Tzvelev, tissue (62.95 mg/kg). BAC valuesshowed that all species are manganese excluders, with a remarkable maximum value of 0.55 in Lolliumperenne. L. All species had average BAC above 1 for Cu and Zn, with a maximum in Leymus racemosus ssp.sabulosus (M. Bieb.) Tzvelev (4.85 and 2.98, respectively). However, there was a significant variation amongindividuals, usually, a high metal content in plant tissue being associated with low metal concentration insoil. The exception was Lollium perenne L., with BAC constantly above 1 (average BAC of 2.27 for Cu and1.69 for Zn). These results emphasize a significant potential for phytostabilization of copper- and zinc-richsoils and open the way for heavy metals phytoextraction capacity studies of the Poaceae species alongRomanian littoral.

Keywords: bioaccumulation, Poaceae sp., copper, zinc, manganese

A growing worldwide industry leads to an increasingdemand for various heavy metals and, subsequently, a risein soil heavy metal pollution. This has a significant impacton biodiversity, agricultural output and also on animal andhuman health.

These are the reasons why, during the past decades,much attention was given to phytoaccumulation andpotential heavy metal hyperaccumulators.

Above certain limits, metal ions have significant are toxicto plants, especially to root tissues. There are differentadaptations that ensure a higher tolerance to this type ofpollution in certain species.

Some plants are able to limit metal uptake in roots(excluders). Others accumulate metal in root tissue, butlimit translocation to other organs (photosyntheticparenchyma in leaves and stems are particularly sensitiveto metal toxicity. The last category of plants usestranslocation from roots to aboveground organs to distributeor store toxic ions (accumulators) [1, 2].

About 0.2% of known plants have extreme accumulationabilities, being considered hyperaccumulators (metalconcentration in upper organs is 100-1,000 times higherthan in average vegetation). Phytoaccumulation presentstwo potential practical applications.

Phytoremediation is the limitation or removal of soilpollution. Phytomining is the extraction of economicallyvaluable metal amounts from specific plant crops. For bothto happen, selected plants must be adapted to localenvironmental conditions, have a high rate of metalaccumulation and/or a high individual biomass or plantdensity [1, 2].

The purpose of this research was to assess thebioaccumulation potential of three extremely common* email: ticuta_np@yahoo.com

Poaceae species growing along the Romanian Black SeaCoastal area.

Hordeum murinum ssp. murinum L. (barley grass, wallbarley) is an annual winter grass, with an usually shortstem (usually up to around 30 cm), growing in tufts. It hasgreen, glabrous leaves and a barley-like spike, green or,sometimes, purplish. It is a native of Western Eurasia andNorth Africa, nowadays spread worldwide as a weed. It isimportant as a forage plant in dry climates and also usedin Chinese cuisine [3].

Leymus racemosus ssp. sabulosus (M. Bieb.) Tzvelev(mammoth wild rye) is a tall perennial grass (up to 100cm tall), with extensive rhizome, coarse, pubescent stemand long (20-40 cm), glaucous green leaves and large 15-30 cm compound spikes. It is a native of Central Asia,nowadays growing in many temperate areas around theworld, as a wild grass (this subspecies is usually found insandy areas, like marine beaches) or grown for dunestabilization or revegetation of mine tailings [4].

Lollium perenne L. (perennial ryegrass) is a perennialgrass, with short rhizomes and 30-100 cm aerial, erectstems, narrow, stiff, dark green leaves and narrow spikes.It is a native of Eurasia and North Africa, currently growingworldwide. It is valuable as forage, for turf and erosioncontrol [5].

Experimental partVegetal products samples were collected from the South

Littoral of Romanian Black Sea, Constanta County,specifically, from the Trei Papuci and Modern Beach area,in the period June 2014 – June 2015. Plant raw material(shoots and leaves) was collected from three individuals

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for each species. Adjacent soil samples were also collectedfrom the mentioned locations.

Plant organs, cut in small pieces and soil samples, wereoven dried for 3 days at 80oC. 0.25 g of each sample weredigested in 5 mL concentrated HNO3 and boiled at 150oCfor one hour. After cooling, volumes of 2 mL H2O2 (30%)were subsequently added, followed by two hours boilingat 150oC. Samples were diluted to 50 mL each, addingNH4Cl (2% solution) and CaCl2 (0.5% solution) respectively,to reduce interferences [6-8].

Metals concentration from digested samples wasdetermined through Atomic Absorption Spectrometrymethod, using HR-CS AAS Spectrometer ContrAA 700,Analytik Jena AG, Germany, acetylene flame technique, atspecific wavelengths for copper (324 nm), manganese(279 nm) and zinc (213 nm) [21]. AAS calibration curvesfor the selected Cu, Mn and Zn metals, are emphasize infigure 1 resulting values were expressed as mg/kg.

For each vegetal product sample, the metal biologicalaccumulation coefficient (BAC) was calculated. This is aratio of metal concentration in shoots to the concentrationfound in soil and is an important index showing metalaccumulation and translocation to upper organs [9, 10,22]:

BAC = [Metal]Shoot/[Metal]Soil

Results and discussionsWhen assessing metal phytoaccumulation, there are

three main factors that need to be considered.Figure 2 emphasize the heavy metals concentration in

plant tissues. The minimal detection limits for the device

used are 0.46 mg/kg (Cu), 0.16 mg/kg (Mn) and 1.44 mg/kg (Zn).

Figure 3 emphasize the average of BiologicalAccumulation Coefficients (BAC) calculated for eachvegetal species and selected heavy metals.

First of all, there are the average metal concentrationsfound throughout the plant world. Following studies on theconcentration of various elements in different plantspecies, researchers have conceived a hypotheticalstandard reference plant, which, among others, wouldcontain 10 mg/kg Cu, 200 mg/kg Mn and 50 mg/kg Zn[11]. Compared to this standard, all analyzed plants hadvalues below average for Cu (5.45 mg/kg maximum inHordeum murinum) and Mn (104.08 mg/kg maximum inLollium perenne). For Zn, Leymus racemosus had anaverage concentration of 62.94 mg/kg, however, with highvariation among individuals (27.04-118.8 mg/kg). Another,

Fig. 1. AAS calibration curves forselected heavy metals

Fig. 2. Average heavy metal concentration in plant abovegroundorgans (mg/kg)

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higher threshold, regards specifically hyperaccumulatorplants. For Cu, the minimal threshold is considered at 1,000mg/kg (with an alternative proposal: 300 mg/kg), whilefor Mn and Zn, 10,000 (or, 3,000) is considered the minimalvalue [1, 11]. None of the selected plants can be considereda hyperaccumulator according to this definition.

However, researchers point out that not allhyperaccumulators can surpass such thresholds under anycondition. There are vegetal species (for instance, the well-known zinc hyperaccumulator, Sedum alfredii) that showlower tissular concentrations on normal soils and reachthe conventional threshold only when growing on zinc-richsoil [1]. Other plants are limited in expressing theiraccumulative traits by the presence/absence of variousnutrients or various stages of their vegetative cycle (thecommon Trifolium pratense, for instance) [12].

Thus, there is a third important factor, the BAC. Neitherthe BAC is constant on all types of soils and a plant showinga high BAC on normal soil could have no bioaccumulationvalue on metal-rich soil, due to potential phytotoxic effectsor other causes.

A BAC above 1, however, can indicate a plant havingphytoaccumulation potential. It could be valuable forphytostabilization, or even for phytoextraction, or at leastas a bioindicator, but further experiments are required forconfirmation [11, 13, 23].

For the three studied species, all BAC values for Mn werebelow 1. Manganese accumulation is rare and variousdeterminations conducted on local native vegetation orcommon ornamentals have shown that most plantsexclude this metal from their roots [7, 14, 15]. Under thesecircumstances, the average 0.55 BAC for Mn in Lolliumperenne can be considered unusually high, especially onsoils with high Mn content (over 200 mg/kg on average).

All species had an average BAC above 1 for copper(Leymus racemosus had the highest average BAC – 4.85).However, in both Hordeum murinum and Leymusracemosus, these values were not constant, with high BACin some individuals and values below 1 in others. In bothcases, the maximum values were reached on the soilswith the lowest metal content.

Lollium perenne emphasize a constantly BAC above 1in all individuals (1.66-2.69, average 2.27), however, allgrowing on soil with low copper content (average 1.77mg/kg).

A similar situation was found for zinc. BAC values forHordeum murinum and Leymus racemosus were variable,with high values on soils with the lowest metal content.Only Lollium perenne had constantly high BAC (1.34-2.02,average 1.69) in all studied individuals.

Other Poaceae were also found to be metalaccumulators. Nazir et al., found BAC > 1 for Zn in Brachiaria

reptans and Eleusine indica, and root accumulation ofcopper (without translocation to upper organs) in Brachiariareptans, Dactyloctenium aegypticum and Eleusine indica[9].

Other species were found less valuable. BAC < 0.4 wasfound for all three metals in Stipa barbata and Melicapersica grown on metal-contaminated soil [16, 17].

The common reed (Phragmites australis) was alsosubject to various bioaccumulation assays (including) forthe selected heavy metals, with different results. Whilesome results point for a low BAC (for instance, < 0.35 forZn, at average concentration in adjacent sediments) [18,19], other researchers reported a significant copper andzinc accumulation and translocation [20, 24].

ConclusionsAll studied species were found to be manganese

excluders. None of the studied species can be consideredas a heavy metals hyperaccumulator.

At the same time, all three species accumulated Cuand Zn in their aboveground organs. The most promisingappears to be Lollium perenne, with a constant BAC above1 for Cu and Zn. Further research is required to confirm itsphytoextraction potential.

The obtained results emphasize a significant potentialfor phytostabilization of copper- and zinc-rich soils and openthe way for heavy metals phytoextraction capacity studiesof Poaceae species along Romanian Black Sea Coast.

References1. RASCIO, N., NAVARI-IZZO, F. Plant Sci., 180, 2011, p. 199.2. TANG, Y.T., DENG, T.H.B., WU, Q.H., WANG, S.Z., QIU, R.L., WEI,Z.B., GUO, X.F., WU, Q.T., LEI, M., CHEN, T.B., ECHEVARRIA, G.,STERCKEMAN, T., SIMONNOT, M.O., MOREL J.L. Pedosphere, 22, nr.4, 2012, p. 470.3. JACOBSEN, N., VON BOTHMER, R. Nord. J. Bot., 15, 1995, p. 449.4. UNITED STATES DEPARTMENT OF AGRICULTURE – NATURALRESOURCES CONSERVATION SERVICE. http://plants.usda.gov/plantguide/pdf/pg_lera5.pdf.5.UNITED STATES DEPARTMENT OF AGRICULTURE NATURALRESOURCES CONSERVATION SERVICE. http://plants.usda.gov/plantguide/pdf/pg_lopep.pdf.6.SHANKER, A.K., DJANAGUIRAMAN, M., SUDHAGAR, R.,CHANDRASHEKAR, C.N., PATHMANABHAN, G. Plant Sci., 166, 2004, p.1035.7.BURADA, A., TOPA, C.M., GEORGESCU, L.P., TEODOROF, L.,NASTASE, C., SECELEANU-ODOR, D., ITICESCU, C. Rev. Chim.(Bucharest), 66, no. 1, 2015, p. 48.8. POPOVICIU, D.R., NEGREANU PIRJOL, T., BERCU, R. Rev. Chim.(Bucharest), 67, no. 4, 2016, p. 670.9. NAZIR, A., MALIK, R.N., AJAIB, M., KHAN, N., SIDDIQUI, M.F. Pak. J.Bot., 43, nr. 4, 2011, p. 1925.10. OBASI, N.A., AKUBUGWO, E.I., KALU, K.M., UGBOGU, O.C. Int. J.Biochem. Res., 1, nr. 4, 2013, p. 16.11. VAN DER ENT, A., BAKER, A.J.M., REEVES, R.D., POLLARD, A.J.,SCHAT, H. Plant Soil, 362, no. 1-2, 2013, p. 319.12. MASU, S., JURJ, L.N. Rev. Chim. (Bucharest), 63, no. 12, 2012, p.1306.13. RADULESCU, C., STIHI, C., BARBES, L., CHILIAN, A., CHELARESCU,D.E. Rev. Chim. (Bucharest), 64, no. 7, 2013, p. 754.14. MUSCALU, L.A., POPOVICIU, D.R., NEGREANU PIRJOL, T., BERCU,R., FAGARAS, M., 15th International Multidisciplinary ScientificGeoConference SGEM-2015, Nano, Bio and Green - Technologies fora Sustainable Future, 18- 24 June 2015, Albena, Bulgaria, ConferenceProceedings, 2, p. 237.15. ROSCA, A.R., POPOVICIU, D.R., NEGREANU-PIRJOL, T., FAGARAS,M., 15th International Multidisciplinary Scientific GeoConference SGEM-

Fig. 3. Average Biological accumulation coefficients (BAC) forselected vegetal species and heavy metals

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2015, Nano, Bio and Green - Technologies for a Sustainable Future, 18- 24 June 2015, Albena, Bulgaria, Conference Proceedings, 2, p. 243.16. MALAYERI, B.E., CHEHREGANI, A., YOUSEFI, N., LORESTANI, B.Pak. J. Biol. Sci., 11, 2008, p. 490.17. LORESTANI, B., CHERAGHI, M., YOUSEFI, N. World Acad. Sci. Eng.Technol., 5, 2011, p. 341.18.STRBAC, S., SAJNOVIÆ, A., KASANIN GRUBIN, M., VASIÆ, N.,DOJÈNOVIÆ, B., SIMONOVIÆ, P., JOVANÈIÆEVIÆ, B. Appl. Ecol. Env.Res. 12, 2014, p. 105.19. KUCAJ, E., ABAZI, U. European Journal of Physical and AgriculturalSciences, 3, 2015, p. 54.

20. EBRAHIMI, M., JAFARI, M., SAVAGHEBI, G.R., AZARNIVAND, H.,TAVILI, A., MADRID, F. International Journal of Agricultural Science,Research and Technology, 1, 2011, p. 73.21. BUCUR ARPENTI, M., NEGREANU-PIRJOL, T., EHLINGER, T.,PARASCHIV, G.M., TOFAN, L., Rev. Chim. (Bucharest), 65, no. 9, 2014,p. 1108.22. OPREA, O., ANDRONESCU, E., FICAI, D., FICAI, A., OKTAR, F. N.,YETMEZ, M., Current Organic Chemistry, 18 (2), 2014, p.19223. LACATUSU, I., NICULAE, G., BADEA, N., STAN, R., POPA, O., OPREA,O., MEGHEA, A., Chemical Engineering Journal, 246, 2014, p. 31124. RADULESCU, M., ARSENIE, L.V., OPREA, O., VASILE, B.S., Rev.Chim. (Bucharest), 67, no. 12, 2016, p. 2596

Manuscript received: 26.01.2017