Phytoremediation Of Petroleum Hydrocarbon Using Three .
BIODIVERSITASVolume 19, Number 3, May 2018Pages: 786-792ISSN: 1412-033XE-ISSN: 2085-4722DOI: 10.13057/biodiv/d190305Phytoremediation of petroleum hydrocarbon using three mangrovespecies applied through tidal bioreactorNURUL HIDAYATI, HAMIM HAMIM , YOHANA C. SULISTYANINGSIHDepartment of Biology, Faculty of Mathematics and Natural Sciences, Institut Pertanian Bogor. Jl. Agathis, Kampus IPB Darmaga, Bogor 16680, WestJava, Indonesia. Tel.: 62-251-8625481, email: hamim@ipb.ac.id; hamimhar@gmail.comManuscript received: 14 March 2018. Revision accepted: 9 April 2018.Abstract. Hidayati N, Hamim H, Sulistyaningsih YC. 2018. Phytoremediation of petroleum hydrocarbon using three mangrove speciesapplied through tidal bioreactor. Biodiversitas 19: 786-792. Mangrove is important component in coastal ecosystems that often facesenvironmental pollution including industrial waste, particularly oil waste (petroleum hydrocarbon). This study was aimed to analyze thegrowth, anatomical, and physiological responses of three mangroves species to petroleum hydrocarbon contaminant applied using tidalbioreactor (ebb flow system) as a model of sea-shore environmental condition. The experiment was carried out using completelyrandomized design with two factors i.e.: three species of mangrove (Rhizophora sp., Avicennia sp. and Bruguiera sp.) and four levels ofpetroleum concentration (0%, 10%, 20% and 30%) exposed using tidal bioreactor for 30 days. The parameters observed in thisexperiment were plant viability, total petroleum hydrocarbon (TPH) of media, plant growth, pH of media and roots malondialdehyde(MDA) content. Leaf anatomy was also analyzed to distinguish accumulation and translocation of oil compounds into leaf tissues. After30 days of treatments, Rhizophora sp. had the highest viability and was able to reduce the higher level of TPH in the media compared tothose of Avicennia sp. and Bruguiera sp. Petroleum hydrocarbon treatment caused the decrease of shoot and root growth of all species,as well as pH of the media, while it significantly increased malondialdehyde of the roots. Anatomical data showed that petroleumhydrocarbon was absorbed and translocated into the guard cell of the stomata. The result shows that Rhizophora sp. was the mosttolerant mangrove species to petroleum hydrocarbon compared to Avicennia sp. and Bruguiera sp., indicated by its survival, thedecrease of petroleum hydrocarbon in the sediment, and lower MDA content.Keywords: Malondialdehyde, mangrove, oil contaminant, phytoremediation, Rhizophora, total petroleum hydrocarbonINTRODUCTIONRiau Province, Indonesia has mangrove forest areamore than 120,000 ha (BPS 2016), where 14062.50 ha arein Dumai City (BPS 2013). The area of the forest had beendeclining for a long time due to human activities around theforest. The data presented by research team of YayasanLaksamana Samudera for example, showed that within 4years (1998-2002) mangrove forest in Dumai had been lostmore than 471 ha, which was similar to a decrease of 7.99percent (PAB 2014). This decrease might be caused byhuman activities including oil industry activities aroundthis area and even oil carrier crashed that may happened,which resulted in various environmental and economiclosses and the decrease of carrying capacity of the naturalresources around the city. Process of drilling and refiningof oil sometimes also produces oil sludge, which is amongthe dangerous pollutants contaminating the surface of waterin mangrove forest.Among those pollutants,, petroleum hydrocarbon is oneof the most common organic pollutant groups that ispersistent in the environment, which is normally quantifiedwith a common parameter known as Total PetroleumHydrocarbon (TPH) (Suhardi 2014). Hydrocarbonderivatives such as gasoline, kerosene, paraffin, asphalt,and others are also commonly used by the community,risking spills to the environment. All of those conditionsmay have big impact on the surface layers, surface waterand groundwater sources (Notodarmojo 2005; Cubillos etal. 2014). Therefore, some efforts to restore the coastalenvironment are urgently needed in order to maintain theecological ecosystem balance.Phytoremediation has been promoted as a potentialmethod that becomes a better alternative due to itsefficiency, affordability, and contribution to environmentalsafety (Espinosa et al. 2005). Phytoremediation is a part ofbiotechnology that utilizes physiological processes in plantbody to increase the degradation and reduction of organicas well as inorganic contaminants from the soil, water andgroundwater medium (EPA 2000). Phytoremediation mayutilize various plants including grasses (Glick 2010),shrubs (Altinözlü et al. 2012), trees (Setyaningsih et al.2007; He et al. 2013) as well as aquatic plants (Rai 2008;Sakakibara et al. 2011).In Dumai, mangrove is one of important component incoastal ecosystems that often face environmental pollutionderives from oil industry waste or even oil tanker accidents.In this area, mangrove plants have important role toimprove the environment in coastal area, and therefore theplants can be used in phytoremediation program. Previousstudy suggested that some mangrove species such asRhizophora sp. and Avicennia sp were able to be used istics and were not sensitive to the presence ofTPH in the sediment (Moreira et al. 2011). The tolerancelevel of those species sometimes is associated with theircapacity to adjust their structural and physiological changesin response to unfavorable environments. As an example,
HIDAYATI et al. – Mangrove for phytoremediation of petroleum hydrocarbonDiab and Bolus (2012) noted that the anatomy of Avicenniamarina was changed due to the presence of oil pollutantsstored in the leaf and plant roots. Other species may havedifferent mechanism in response to pollutants. This studyaimed to analyze the growth, anatomical and physiologicalresponses of three mangrove species to petroleumhydrocarbon contaminant applied using ebb flow system toinvestigate the mechanism of tolerance and the prospect forphytoremediation.MATERIALS AND METHODSSediment and oil preparationSediment that was used in this study was collectedfrom the Mangrove Conservation Area of Bandar Bakau,Dumai City, Riau, Indonesia. Sediment samples werecollected randomly from five sites. The samples werefiltered and separated from rocks, and plant materials thenmixed together in a plastic container to ensure uniformity.Oil pollutant that was used in this study was obtained fromNational Oil Company Pertamina Ltd. RU II Dumai.ProceduresMangrove seedling preparationIn this study, mangrove seedlings were prepared fromthe propagules obtained from the Conservation Area ofBandar Bakau Dumai. Three species of mangroves wereused in the experiment, i.e., Rhizophora sp., Avicennia sp.,and Bruguiera sp. Three months old seedlings wereacclimatized for 1 week in the bioreactor before beingtreated with petroleum hydrocarbons.Experimental designThis study was carried out using CompletelyRandomized Design with two factors, i.e., three mangroveplants (Rhizophora sp., Avicennia sp. and Bruguiera sp.)and four levels of petroleum concentration (0%, 10%, 20%and 30 %) with 5 replications.Tidal bioreactor for plantingTidal bioreactor was designed to resemble the tidalconditions in mangrove ecosystem environment byapplying the tidal hydroponic system (ebb flow system)with 2 times ebb and 2 times flow, each of which wasrunning for 6 hours as shown in Figure 1.Figure 1. Design of tidal hydroponics bioreactor system (ebbflow system)787Plant growth measurementPlant growth was observed by measuring plant height,the number of new roots and viability of the mangroveplant. Plant height was measured from the base of stem tothe top of the shoot. Mangrove viability was calculatedfrom the number of plants that were still alive after 30 daysof treatment. Plant morphology including the changes ofleaves color was also observed during the application withTPH treatments.TPHs DeterminationTotal petroleum hydrocarbons (TPHs) in soil weredetermined using the EPA 418.1 method (EPA 2000). Thegrinded soil samples were weighed 5 grams in vials, then 2g of sodium sulfate and 10 mL of TCE(Tetrachloroethylene) were added and stirred for 1 hour.Samples were allowed to stand for 1 minute to separateresidues and solvents. The solvent layer was poured on acolumn that had been filled with 2 g of silica gel 70-230mesh and then the absorbance of the solution was measuredusing IR spectrophotometer HC-404. The absorbance wasrecorded and the solution was diluted if the absorbance wasmore than 2.00. The TPH value was calculated as follow:TPH Cx fp TPH %TPH x 10000Where:C is the measured level (mg/L)fp is the dilution factorPH measurementThe pH measurements of the medium were carried outusing a digital pH meter at the beginning and the end of theexperiment. Calibration was carried out using a buffersolution at pH 4 and 7. The changes of pH value werecalculated from the initial pH minus the final pH of thestudy.Lipid peroxidation analysisMalondialdehyde (MDA) level is a common parameterto evaluate lipid peroxidation in membranes and biologicalsystems, as well as indicators of free-radical formation inplant tissues (Halliwell and Chirico 1993). Lipidperoxidation was calculated by measuring the level ofMDA using the method described by (Meriga et al. 2010).The 0.3 g of roots were weighed and homogenized withmortar, after which 5 ml of 0.1% TCA was added andcentrifuged at 10000 x G for 5 min. After centrifugation, 1ml of supernatant was mixed with 4 ml of 0.1%thiobarbituric acid in 20% TCA, and the mixture wasincubated in boiling water for 30 min. Absorbance ectronic, USA) at 532 nm wavelength andadjusted for non-specific absorbance at 600 nm. Thedifference in absorbance at both wavelengths is the valueof MDA concentration as the lipid peroxidation endproduct. The MDA content was estimated by using the 155(mmol/L/cm) extinction coefficient.
B I O D I V E R S I T A S 19 (3): 786-792, May 2018Data analysisAll quantitative data were analyzed using SPSS 22.0 toexamined Analysis of Variance (ANOVA), followed byfurther analysis using Duncan’s New Multiple Range Test(DNMRT) at α 5% to verify the response of mangrovespecies to petroleum treatment.RESULTS AND DISCUSSIONPlant viability and TPH of the mediaPetroleum treatment applied using tidal bioreactorsignificantly reduced plant growth and viability or survivaldepending on mangrove species and TPH concentration.The data related to the response of these plants and TPHconcentration in the media is presented in Table 1. After 30days of petroleum treatment, Rhizophora sp. had thehighest survival rate which reached 100% followed byBruguiera sp. and Avicennia sp., that was 65% and 60%,respectively (Table 1). The treatment of oil in the mediawith various concentrations did not affect the survival rateof Rhizophora sp.; while, it decreased that of the otherssignificantly even until 20% for Avicennia sp. (Table 1),suggesting that Rhizophora sp. was more resistant andadaptable to oil pollutant in the media than the other twospecies. Some chlorotic symptoms were observed on theleaves of Bruguiera sp. and Avicennia sp, which led to thedeath of some sample plants before 30 days. All theremaining plants that survive were able to grow well afterexperiment when they were transplanted to the normalmedia.The observation of the media after 30 days of thetreatment showed that residual TPH content in the mediawas varied depending on mangrove species. The lowestresidual TPH was found in Rhizophora sp. (15.65 ppm)followed by Avicennia sp. and Bruguiera sp., i.e., 26.98ppm and 32.44 ppm, respectively (Figure 2). This indicatedthat Rhizophora sp. had the ability to reduce TPH contentof the media more than the other two species. Higherreduction of TPH content in the media of Rhizophora sp.confirmed the ability of the species to reduce hydrocarboncontaminant from its environment (Figure 2).The growth of mangroveThe response of mangrove to TPH treatments was alsoobserved by measuring parameters of growth, including theincrease of plant height and the number of new rootsappeared during the treatment. The decrease of plantgrowth calculated from plant height was occurred in thethree species of mangrove in response to petroleumconcentration where Avicennia sp. had the largest reductionat 30% petroleum concentration (Figure 3). Rhizophora sp.had the lowest growth rate compared to the other twospecies, while the growth of Bruguiera sp. was average.Different from Rhizophora sp. and Avicennia sp., growthreduction of Bruguiera sp. was not significantly differentas compared to that of control plants (0% of petroleum) inresponse to petroleum treatment up to 30% concentration(Figure 3).Table 1. Effect of treatment of petroleum concentration onviability of 3 mangrove plantsOiltreatmentPlant viability (%)(%)0100 a100 a100 a100.010100 a60 b60 b73.320100 a60 b60 b73.330100 a20 d40 c53.3Average100606575Note: The number followed by similar letter does not significantlydifferent based on Duncan Analysis at 5% of Histochemical analysis of leaves and roots tissuesPreparation of leaf paradermal tissues was carried outby scraping adaxial parts of the leaves to get the abaxialsamples. The preparation of leaf and roots transversalsection was carried out by cutting leaves and roots using afrozen microtome (Yamato RV-240) with a thickness of17-20 μm. The leaf sections were immersed to 70% ethanolfor 1 min then stained in filtered solutions of Sudan IV in70% ethanol for 30 min at 40oC in water bath, and washedrapidly with 70% ethanol (Boix et al. 2011). The sectionwas placed in the object glass and closed by cover glassafter added by glycerin. Samples were examined, and thephotograph was taken using a light microscope camera(Olympus CX23 asp.sp.RhizophoraAvicennia sp.sp.AvicenniaMangrove re 2. The residue of TPH observed from the media grown by3 mangrove species after 30 days treatmentHigh increase (cm)788Petroleum concentration (%)Figure 3. The increase of plant height after 30 days of petroleumtreatment
HIDAYATI et al. – Mangrove for phytoremediation of petroleum hydrocarbonLipid peroxidation analysisThe negative effect of petroleum treatment was alsoobserved, indicated by the increase of malondialdehyde(MDA) content in the root tissues of all mangrove specieswhich suggested the occurrence of lipid peroxidation. Theincrease of MDA levels occurred in line with the increaseof petroleum concentration observed in all mangrovespecies. The increase of MDA levels in the roots indicatedthat the plants underwent stress due to the petroleumtreatment. Three species of mangroves showed differentresponses to the petroleum treatment; the content of MDAin the roots of Rhizophora sp. was lower than that ofAvicennia sp. and Bruguiera sp. under all petroleumtreatments. The MDA of Bruguiera sp. was the highestamong the three species in almost all treatments (Figure 5).Histochemical analysis of leaves and roots tissuesAnatomical analysis of mangrove leaves was carriedout to observe the absorption, distribution andaccumulation of hydrocarbon compounds into the planttissues especially in the leaf tissues, including stomata. Theaccumulation of petroleum hydrocarbon compounds in allmangrove species was found in roots and leaves. Theobservation of paradermal tissues using Sudan IV solutionshowed that petroleum accumulation was occurred inmangrove leaves. The accumulation of petroleum wasobserved in adaxial as well as abaxial of epidermal leavesof all mangrove species, which was marked by theTable 2. Effect of treatment of petroleum concentration onsediment pH reduction after 30 tration(%)0102030pHBefore5.85 a5.45 a5.36 ab5.26 abpHAfter5.67 a4.65 b4.51 bc4.30 cpHReduction0.18 c0.80 ab0.85 a0.96 a01020305.10 ab4.95 b4.93 b4.86 b4.80 b4.70 b4.67 b4.59 b0.30 c0.25 c0.26 c0.27 c05.72 a5.51 a0.21 c105.58 a4.95 b0.63 b205.56 a4.87 b0.69 b305.38 ab 4.69 b0.69 bNote: The number in the similar column followed by similar letterdoes not significantly different based on Duncan Analysis at 5%of αBruguierasp.Number of rootThe changes of pH of planting mediumIn general, petroleum treatment during 30 daysdecreased pH of the media. The decrease of pH occurred inall mangrove species treated with different petroleumconcentrations. The decrease of pH medium in treatedplants was higher than that in the control plants, except inAvicennia sp. In Rhizophora sp. and Bruguiera sp. the pHof the media treated with petroleum was more acidic thanthe control media, where the higher concentration ofpetroleum tended to reduce pH of the mangrove mediummore (Table 2).appearance of yellowish-orange to red color in the object(Figures 6 and 7). The accumulation of petroleum insidethe leaves was clearly observed in the stomata ofRhizophora sp. and Bruguiera sp., while for Avicennia sp.,the accumulation of petroleum in the stomata was notclearly visible because it was covered by heavy trichomeon the leaf surface (Figure 6.H). Petroleum treatment alsoaffected the color of trichome in Avicennia sp. in which thetrichome of control plant was brighter compared to themangroves treated with petroleum which had darker color(Figures 6.F and H).Petroleum concentration (%)Figure 4. The number of new roots that appear on mangroveplants after 30 days of petroleum treatmentMDA level (μmol)Petroleum treatment not only affected the shoot growth,but also involved in the emergence of new roots, theimportant organs required to absorb water and nutrients.The result showed that there was a decrease in the numberof new roots that emerged after 30 days of petroleumtreatment (Figure 4). Rhizophora sp. and Avicennia sp. hadrelatively equal number of new roots at control level, but inthe TPH treatment, Rhizophora sp. had more new rootsthan Avicennia sp. The number of new roots in Bruguierasp. was less than in Rhizophora sp. and Avicennia sp., bothin the control and petroleum treatments. Bruguiera sp. hadthe slowest root growth compared to the others, but thedecrease of root growth was not significantly different inresponse to petroleum treatments. Based on the data ofshoot and root growth, it can be seen that petroleumtreatment had a varied influence in mangrove growth,where the roots were more sensitive to the presence ofpetroleum hydrocarbon compounds compared to theshoots.789Petroleum concentration (%)Figure 5. Effect of petroleum concentration treatment on MDAlevel on root of mangrove plant
B I O D I V E R S I T A S 19 (3): 786-792, May 2018790ABCDEFGHIJKL0%30%0%Adaxial30%AbaxialFigure 6. Paradermal observation of mangrove leaves. Rhizophora sp. (A, B, C, and D), Avicennia sp. (E, F, G and H) and Bruguierasp. (I, J, K and L)ABCDEFGHIJKL0%30%Adaxial0%30%AbaxialFigure 7. Transversal observation of mangrove leaves. Rhizophora sp. (A, B, C, and D), Avicennia sp. (E, F, G and H) and Bruguierasp. (I, J, K and L)
HIDAYATI et al. – Mangrove for phytoremediation of petroleum hydrocarbonDiscussionThe presence of hydrocarbon compounds in the mediaapplied using tidal bioreactor caused disturbances to themangrove plants, which was observed by the decrease ofsurvived mangrove or mangrove viability after petroleumtreatment. The higher viability revealed the better tolerancelevel of mangrove in response to TPH contaminant, asindicated by Rhizophora sp. (Table 1). The decrease ofviability indicated the mortality rate of mangrove 1 monthafter exposure to petroleum treatment. The mortality ofsome mangrove species can also be observed by thepresence of chlorosis and necrosis symptom, which leads toleaf abscission and even plant death (Data not shown).Symptoms of mangrove mortality
Volume 19, Number 3, May 2018 E-ISSN: 2085-4722 Pages: 786-792 DOI: 10.13057/biodiv/d190305 Phytoremediation of petroleum hydrocarbon using three mangrove species applied through tidal bioreactor NURUL HIDAYATI, HAMIM HAMIM , YOHANA C. SULISTYANINGSIH Department of Biology, Faculty of Ma
B1.2 Support Hydrocarbon Production – Processing Facilities 38 B1.3 Support Hydrocarbon Production – Pipeline Export Power 40 B1.4 Support Hydrocarbon Production – Utilities and Offsites 40 B1.5 Support Manufacture of Downstream Hydrocarbon Products – Oil Refinery Products 41 . (ARENA) to provide a “desk-top” assessment of the .
Fort Jackson, S.C., March 23 and 24, 1995. 32 12. Field-property determinations, and petroleum- hydrocarbon and dissolved-lead concentrations detected in ground-water samples from Site 1062, Fort Jackson, S.C. , November 3, 1995 . 34 13. Petroleum-hydrocarbon concentrations detected in
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Petroleum Refining Process Descriptions Petroleum refining is a complex industry that generates a diverse slate of fuel and chemical products, from gasoline to heating oil. The refining process involves separating, cracking, restructuring, treating, and blending hydrocarbon molecules to generate petroleum products. Figure 1 shows the overall .
How to cite: Z. Mohebi, M. Nazari, Phytoremediation of wastewater using aquatic plants, A review, Journal of Applied Research in Water and Wastewater, 8 (1), 2021, 50-58. . and the high efficiency of purification. The primary role of plants in this system is to supply the oxygen required by heterotrophic microorganisms in the root zone .
polycyclic aromatic hydrocarbons (PAHs). Polycyclic aromatic hydrocarbons are undoubtedly the most widespread carcinogens in the environment (Harms et al., 2003) and are commonly associated with the disposal of combusted materials or petroleum residues (Suess, 1976). Polycyclic aromatic hydrocarbons are a group of ubiquitous hydrocarbon .
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