In Situ Study On The Environmental Behaviors Of Typical PAHs In Mangrove Plants

Mangroves are the intertidal plant formations growing along the coastlines oftropical and subtropical regions.Due to an increase in population,and expansion ofindustrial agricultural areas and port,a large number of pollutants are dischargingdirectly or indirectly into offshore marine areas.Human activities have seriously led tothe damage of many mangroves.The unique features of mangrove ecosystems such ashigh primary productivity,abundant detritus,rich organic carbon and anoxic conditionsmake themselves a reservoir of polycyclic aromatic hydrocarbons (PAHs)fromanthropogenic inputs.However,the published research on PAHs in the mangrovewetland has mainly focused on the analysis of concentrations of individual PAH andtotal PAHs compounds in the sediment;the determination of the distribution andproportion of PAHs in different tissues of many mangrove species;the evaluation of thepossible sources of PAHs in the wetland;and the microcosm studies concerning theremoval of PAHs from surface-or bottom-contaminated sediments.As a result,research on the direct uptake of PAHs by mangrove plants as the core of the wetland israrely involved,nor are the related mechanisms being explained.In this study,a systematic method was established for in situ study on theenvironmental behaviors of typical PAHs in the mangrove wetland utilizing the solidsurface fluorimetry (SSF)and fluorescence microscopy (FM)as well as thecombination of two photon laser confocal scanning microscopy (TPLCSM).And theenvironmental behaviors of typical PAHs in mangrove plants were studied in situ withthe established methods.Main results were summarized as followings:(1)A SSF approach combining with a laboratory-made solid substrate holder anda fluorescence spectrophotometer was established for direct determination ofanthracene (An)adsorbed on the leaves of four mangrove species (Avicennia marinaAm,Bruguiera gymnorrhiza Bg,Kandelia candel Kc,Rhizophors stylosa Rs).Theexperimental results were as follows:(a)With the same exposure time (90min)of An, the leaves of the four mangrove species showed different capacities for adsorbing An,and within a certain range,the relative SSF intensities of the An adsorbed on theupper sides of leaves of the four mangrove species increased linearly with incrementsin the exposure quantity of An.The recoveries for the four species were between70.2% and 110.8%,which met the requirement of practical applications.(b)In thesame exposure concentration of An,the relative SSF intensities of leaves of the fourmangrove species increased first and then decreased as the time progressed.And themaximum relative SSF intensities of the four species were in the orderAm＞Kc＞Rs＞Bg.(c)In the same experimental conditions,the relative SSF intensitiesof the adsorbed An on the upper side of the same mangrove leaves were muchstronger than those adsorbed on the lower side,which has many stomata.Therefore,in addition to the uptake of PAH via waxy cuticle,the stomatal pathway was alsoimportant for the PAH entering into the mangrove leaves.Further studies showed thatthe relative SSF intensities in different regions of the same leaf differed from eachother,and it provided us with very important information that the adsorbed An on themangrove leaf surface might not“equally”penetrate the epidermal wax layer into theinternal leaf tissues,but might have“priority”locations.However,the reasons werenot clear.(2)Based on the established SSF method,a synchronous solid surface fluorimetry(S-SSF)method combining with an optical fiber and a fluorescencespectrophotometer was established firstly for direct determination of the An andPyrene (Py)which were adsorbed on the leaves of two mangrove species insingle-component or in mixtures.The results indicated that the quantity of An and Pyadsorbed on the mangrove leaves in single-component or in mixtures linearlycorrelated with their S-SSF intensities.And the recoveries of this S-SSF method forthe leaves of two mangrove species were between 82.9% and 116.5%,which met therequirement of practical applications;Secondly,the distribution of An adsorbed onthe living Kc leaves were studied in situ utilizing the established S-SSF approach,and the GC-MS was also used to verify the accuracy of the S-SSF method.Theexperimental results demonstrated that in a certain period of exposure time of An,thetrends of the quantity of adsorbed An on the upper side of Kc leaves determined bythe two methods were similar.Thus,the S-SSF could be used as a novel in situmethod for further studies.Finally,the photolysis of An and Py on the leaves of twomangrove species in single-component or in mixtures were directly studied by theS-SSF,and the results were as follows:(a)The processes of photolysis of An and Pyin single-component adsorbed on the leaves of two mangrove species followedfirst-order reaction kinetics.And in the same experimental conditions,the higher thequantity of adsorbed PAH on the mangrove leaves,the slower the reaction rate of thephotolysis of the PAH,moreover,the raction rate of the adsorbed Py on the leaves ofthe same mangrove species was much slower than that of the same amount ofadsorbed An.(b)The processes of photolysis of An and Py in mixtures adsorbed onmangrove leaves also followed first-order reaction kinetics.And to the same amountof PAH in mixtures,the raction rate of PAH adsorbed on Kc was slower than thatadsorbed on Ac.The experimental results also revealed that compared with the Anand Py in single-component mode adsorbed on the same kind of mangrove leaves,thereaction rate of An in mixtures slowed down,while the reaction rate of Py in mixturesspeeded up.(c)The processes of photolysis of An and Py in single-component or inmixtures,and in water followed first-order reaction kinetics.And to the same amountof PAH in single-compont mode,the reaction rate of PAH in water was faster thanthat adsorbed on mangrove leaves.Compared with An and Py in single-componentmode in water,the reaction rate of An in mixtures slowed down,while the reactionrate of Py in mixtures speeded up.(3)Under the artificial simulating conditions,the An selected as the model PAHwas applied as a homogeneous layer to the upper side of the living Kc leaves.Duringthe exposure time of 96 hours,the SSF was firstly utilized for direct determination ofthe adsorbed An on the Kc leaves,then the FM was used to visualize how An was taken up into Kc leaves and where it was located in the inner leaf tissues.The resultsdemonstrated that the overall amount of An residing on the Kc leaf surface decreasedgradually as time progressed.And after a certain period of exposure time,equilibration of the An might be achieved between the leaf surface and theatmosphere.And as the time passed,the An existing on the upper side of Kc leaveswas observed moving from the upper epicuticular wax into the first and secondhypodermis and upper palisade tissue,and the An existing on the lower side of Kcleaves moving from the stomata into the lower hypodermis and the lower palisadetissue,then ultimately into the spongy mesophyll.The results also showed that afterexposed for 24 hours,there were several light blue dots in the upper and lowerpalisade tissues and spongy mesophyll,which might be the autofluorescence of Anmetabolites.Further studies also revealed that the residual quantity of An at the centerwas much less than that remaining at the edges of the same leaf surface after exposureof 24 hours,and this suggested that as time passed,the quantity of An retaining at thecenter of the leaf surface decreased more quickly,which might provide important newinformation about how An enters into Kc leaves.(4)Under the artificial simulating conditions,the uptake,transportation anddistribution of PAHs were in situ visualized in mangrove plants using TPLCSM,some results were as follows:(a)After 20 days of culture in contaminated solutions,the Naphthalene (Na),An,Phenanthrene (Phen)and Py had been observed enteringinto the roots of Kc seedlings and being transmitted to the stem and leaf due totranspiration.The pathways of the four PAHs from the culture solutions into theplants were similar,and the results also indicated that the transmission rates of thefour PAHs in Kc seedings were in the order Na＞An and Phen＞Py.The growth of theroot of Kc seedlings was inhibited by the PAHs and the inbibitional effect was in theorder Na＞An and Phen＞Py.In addition,the inhibition of An and Phen to the rootgrowth was not significant difference,and the inhibition of Na was the most obvious.(b)After 20 days of culture in contaminated solutions,the three PAHs (Na,Phen,Py) had been observed entering into the roots ofAegiceras corniculata (Ac)seedlings andbeing transmitted to the stem due to transpiration.The pathways of the three PAHsfrom the culture solutions into the plants were similar,and the results also indicatedthat the transmission rates of the three PAHs in Ac seedlings were in the orderNa＞Phen＞Py.The growth of the root of Ac seedlings were inhibited by the PAHs andthe inbibitional effect was in the order Na＞Phen＞Py,and the inhibition of Na was themost obvious.In this study,the SSF,FM and TPLCSM were firstly combined for in situ studyon the environmental behaviors of typical PAHs in the mangrove plants.It providednovel means,methods,and research idea for the related studies of PAHs in mangroveecosystems.Through studying the processes of uptake,transportation and degradationof typical PAHs in different kinds of mangrove leaves,stem and roots,we revealedthe processes and mechanisms on phytoremediation for typical PAHs and pollutioncontrol of mangrove plants.Moreover,it layed a methodological foundation forfurther study of the growth,ecophysiology and biochemical effects of different kindsof mangrove plants under the pollution stress of PAHs.