topic title: hydrilla verticillata, a submerged aquatic plant, as phytoremediation agent of lead (ii)nitrate (pb(no3)2)

THAI NGUYEN UNIVERSITY UNIVERSITY OF AGRICULTURE AND FORESTRY NGUYEN THI THAO TOPIC TITLE: HYDRILLA VERTICILLATA, A SUBMERGED AQUATIC PLANT, AS PHYTOREMEDIATION AGENT OF LEAD (II)-NITRATE (PB(NO3)2) BACHELOR THESIS Study Mode : Full-time Major : Environmental Science and Management Faculty : International Training and Development Center Batch : 2011-2015 Thai Nguyen, 2015 DOCUMENTATION PAGE WITH ABSTRACT Thai Nguyen University of Agriculture and Forestry Degree Program : Bachelor of Environmental Science and Management Student name : Nguyen Thi Thao Student ID : DTN 1153110148 Thesis Title : HYDRILLA VERTICILLATA, A SUBMERGED AQUATIC PLANT, AS PHYTOREMEDIATION AGENT OF LEAD (II)NITRATE (PB(NO3)2) Supervisor (s): Dr Arinafril Dr Ho Ngoc Son Abstract: The experiment of this study were started from March 2015 and finished in June 2015 at Integrated Research Laboratory of Sriwijaya University Palembang in Indonesia, with applied treatments (B1: Pb(NO3)2 ppm(control), B2: Pb(NO3)2 ppm, B3: Pb(NO3)2 10 ppm and B4: Pb(NO3)2 15 ppm) and replications for each treatment through times of observing in 20 days Given results were totally evidenced strong capacity of hydrilla verticillata in absorbing and accumulating heavy metals, especially lead (Pb) which is known as a dangerous toxicity for living organisms This ability of hydrilla verticillata pulled it up to become an effective instrument for phytoremediation technique in solving environmental issues In the research phytoremediation was presented by the use of a submerged aquatic plant, hydrilla verticillata as a tool of removing Lead (II)-nitrate (Pb(NO3)2) from water The revealed results after experiment of treatments, replications in times of observing shown that hydrilla verticillata is an useful plant for treating Pb contaminant in water by taking up and storing the metal in its body After applying Pb(NO3)2 with different treatments, Pb was directly affected hydrilla verticillata morphology based on higher level of contaminant with the changes in morphological states in order of normal; leaves became yellow; leaves became faintly yellow; partial decolourization, complete decolourization and plant died through 20 days of experiment Hydrilla verticillata also demonstrated a large percentage of average Pb absorbed ability is 92.8 % and high ratio of Pb content concentrating in the plant compared to Pb i content in water is 1.88 The study also gave predictions of Hydrilla verticillata potential in improving water quality, with Pb concentration in water was forecasted to be removed all from water in treatment B1, B2 and B3 Hence, this ability of Hydrilla verticillata would be widely known and used for water assessing, monitoring and cleaning operations in the future Keywords: Phytoremediation, Hydrilla verticillata, Lead (II)-nitrate (Pb(NO3)2), absorbed capacity, water heavy metal pollution Number of Pages: 51 Date of Submission: 30/09/2015 Supervisor (Sign) ii ACKNOWLEDGEMENT First and foremost I offer my sincerest gratitude to my supervisor, Dr.-phil Arinafril of Sriwijaya University, Indralaya, Indonesia, who has supported and instructed me throughout my research time and also my thesis completion with all of his patience, devotions, enthusiasms and empathy If I have not had his helps I would have never finished this study Also, I would like to show my great appreciation to Dr Ho Ngoc Son, for supervising, guiding, counseling and advising me in writing and completing the study Besides my two supervisors, I want to send my sincere thanks to the Integrated Research Laboratory of Sriwijaya University Palembang and the Research laboratory in the Department of Chemistry, Faculty of Science, Sriwijaya University Indralaya in Indonesia with their approvals, supports and helps in using place and equipment in the research time this study is completed successfully From the bottom of my heart, I am grateful to Fadila Mutmainnah, S.Si, for her guidance, support and encouragement during the time of accomplishing this research in Indonesia My special thanks go to Ha, also, Chi, Linh, Marisa, Eka and all my classmates, friends who helped me in my months of internship time Finally, I would like to express my heartfelt gratitude to my parents and teachers for having, nurturing and teaching me from the very first steps of my life until now, and their dedications will be my indications for future life Thai Nguyen, 30th September 2015 Student Nguyen Thi Thao iii TABLE OF CONTENTS List of figures List of tables PART I INTRODUCTION 1.1 Research rationale 1.2 Research’s objectives 1.3 Research questions and hypotheses 1.4 Limitations 1.5 Definitions PART II LITERATURE REVIEW 2.1 Water environment 2.1.1 The importance of water 2.1.2 Water resources 2.2 Heavy metal-polluted water 10 2.2.1 Source of heavy metal contaminants in water 10 2.2.2 Consequences of heavy metal contaminants in water 11 2.3 Lead Compound- Lead (II) Nitrate 11 2.4 Phytoremediation 12 2.5 Hydrilla verticillata 13 PART III METHODS 16 3.1 Sampling site and time 16 3.2 Materials and equipment 18 3.3 Methods 19 3.3.1 Applying treatments 19 3.3.2 Laboratory method of analyzing Pb content in Hydrilla verticillata 20 3.3.3 Laboratory method of analyzing Pb content in water 21 3.4 Data analysis methods 21 3.4.1 Variant analysis 21 3.4.2 Lead (Pb) Reduction rate 21 3.4.3 Time Series analysis: Linear regression 22 3.4.4 Bio-Accumulation Factor 22 PART IV RESULTS 23 4.1 Pb concentration in Hydrilla verticillata 23 4.2 Pb concentration in water 25 4.3 Result of Data analysis 27 4.3.1 Variant analysis 27 4.3.2 Time series analysis: Linear regression 32 4.3.3 Lead (Pb) reduction rate 37 4.3.4 Bio-Accumulation Factor 37 4.5 Hydrilla verticillata morphological changes 38 PART V DISCUSSION AND CONCLUSION 39 5.1 Discussion 39 5.2 Conclusions 43 REFERENCES 45 List of figures Figure 3.1: Jakabaring lake in Palembang, Indonesia 3’0158.5’S and 104’7965.4’N 16 Figure 3.2: Integrated Research Laboratory of Sriwijaya University, Palembang, Indonesia which is located in 2’9910.5’S and 104’7295.5’N .17 Figure 4.1: Pb concentration in Hydrilla verticillata 24 Figure 4.2: Pb concentration in water 27 Figure 4.3: Forecasting of Pb concentration in water in treatment B3 33 Figure 4.4: Forecasting of Pb concentration in water in treatment B4 33 Figure 4.5: Forecasting of Pb concentration in Hydrilla verticillata in treatment B1 34 Figure 4.6: Forecasting of Pb concentration in Hydrilla verticillata in treatment B2 35 Figure 4.7: Forecasting of Pb concentration in Hydrilla verticillata in treatment B3 35 Figure 4.8: Forecasting of Pb concentration in Hydrilla verticillata in treatment B4 36 List of tables Table 3.1: Experimental design .19 Table 4.1: Pb concentration in Hydrilla verticillata (ppm) 23 Table 4.2: Pb concentration in water (ppm) 25 Table 3: Significantly different effects of treatments on water’s Pb concentration in the 5th day .28 Table 4: Significantly different effects of treatments on water’s Pb concentration in the 10th day 28 Table 5: Significantly different effects of treatments on water’s Pb concentration in the 15th day 29 Table 6: Significantly different effects of treatments on water’s Pb concentration in the 20th day 29 Table 7: Significantly different effects of treatments on Pb concentration in Hydrilla verticillata in the 0th day .30 Table 8: Significantly different effects of treatments on Pb concentration in Hydrilla verticillata in the 5th day .30 Table 9: Significantly different effects of treatments on Pb concentration in Hydrilla verticillata in the 10th day .31 Table 10: Significantly different effects of treatments on Pb concentration in Hydrilla verticillata in the 15th day .31 Table 11: Significantly different effects of treatments on Pb concentration in Hydrilla verticillata in the 20th day .32 Table 12: Lead (Pb) reduction rate 37 Table 13: Bio-Accmulation Factor (BAF) ratio of Lead (Pb) from water to Hydrilla verticillata 37 Table 14: Hydrilla verticillata’s morphological changes observed in different treatments through time 38 PART I INTRODUCTION 1.1 Research rationale Playing vital role as a source of life, water have been being an indispensable resource to humankind and also other organisms However, water are now getting in a strong alarmed level of pollution, which can directly impact to the whole planet, Balkis (2012) stated “Water pollution has been suggested that it is the leading worldwide cause of deaths and diseases, and that it accounts for the deaths of more than 14,000 people daily In addition to the acute problems of water pollution in developing countries, industrialized countries continue to struggle with pollution problems as well” Especially, current time, water is seriously polluted by heavy metals in the nearby industrial or populated urban areas, these heavy metals can widely distribute in water and directly or indirectly affect human and living aquatic organisms due to human activities ( Mohiuddin, et al., 2010) Lead (II) nitrate (Pb(NO3)2 ) is a toxic compound, and ingestion may cause acute lead poisoning, and also for all soluble lead compounds (USCG, 1999) Besides, when dissolve in water, Lead (Pb) from Lead (II) nitrate (Pb(NO3)2, one of the harmful chemical elements, can be huge influences for human health such as behavioral problems, high blood pressure, kidney damage, memory and learning difficulties, reduced IQ, … (Ernhart, 2006; Spivey, 2007) In term of aquatic systems, lead (Pb) has last and long effects on plants and animals by spreading and existing for long time of about 100 to 1,000 years in water (UNEP, 2010) There are many projects and researches had done in treating metal water pollution, but almost all of those projects required high techniques and also high cost for the steps and treatment processes Furthermore, with the variety in kind of metal PART V DISCUSSION AND CONCLUSIONS 5.1 Discussion All of the research results were obtained after analyzing and observing samples of water and Hydrilla verticillata with treatments of different Pb(NO3)2 concentrations: B1: ppm(control), B2: ppm, B3: 10 ppm and B4: 15 ppm), and replications for each treatment, which were shown in Part IV Hydrilla verticillata showed its absorbability as a tool of phytoremediation (Chaney, 1983) by using body and leaf (Denny & Wilkins, 1987) to absorb lead from lead (II)-nitrate (Pb(NO3)2)-polluted water, this was improved in Table 4.14 with Hydrilla verticillata’s morphological changes in treatments through times of observation with mophological states: normal; leaves became yellow; leaves became faintly yellow; partial decolourization and complete decolourization, these changes also rise evidences for the effecting of different Pb concentrations in treatments to Hydrilla verticillata’s morphology By increasing Pb concentration the lysis of Hydrilla’s cells raised through time leading to the changes in its appearances of leave and body color and state (Liden & Walker, 2015) From the 0th day until the last day - 20th day (table 4.2), the decreases of Pb concentration in water clearly indicated Hydrilla verticillata ability in absorbing heavy metal (Pb) which is detected as a mechanic of this plant (Phukan, 2015) In term of Pb reduction amount by taking initial Pb concentration applied in water minus the final Pb concentration in water of the 20th day, the absorbability of Hydrilla verticillata was given with an increasing order in of the highest Pb reduction amount of 11.86 ppm in treatment B4, followed by 9.94 ppm in treatment B3 and ppm in treatment B2 However, by using the formula to calculate Pb reduction rate (Dalun, 39 2008), the acquired results showed the decreases when Pb applied increase from treatment B1 to B4 Finished the experiment, with Pb concentrations in water of treatment B1 and B2 is ppm -> Pb reduction rate is 100% (Pb content was removed all from water), B3 still had Pb content in water but it is a small amount of 0.06 ppm > Pb reduction rate is 99.4%, only B4 had the highest Pb amount left of 3.14 ppm-> Pb reduction rate is around 79.1% When metals in polluted water are taken in aquatic flora tissues, their concentration in there can become higher than the water environment surrounding even they can come up to about 105 times higher (Albers & Camardese, 1993) As an aquatic flora Hydrilla verticillata is not an exception which had the results shown in tables 4.1 The Pb contents were accumulated into Hydrilla verticillata through its body and leaf as Cheng (2002) has concluded that plants can take up heavy metals by their stems and leaves Bases on the presented results of mean Pb concentration in Hydrilla verticillata (table 4.1) after applying treatments and bases on the formula of calculating Bio-Accumulation Factor (BAF) (Arnot & Gobas, 2006), the ratio of Pb concentration in Hydrilla verticillata per Pb concentration in water in B1, B2, B3 and B4 were determined The greatest BCF values went for B3 of 1.88 followed by B2 of 1.83 and B4 of 1.65 at the day 20th These ratios also showed the competence to accumulate Pb of Hydrilla verticillata and its tolerance capacity (Malik, 2010; Yadav, 2009), when the Pb contaminant increased to higher level by the treatments from B1 to B4 in 20 days of observing, Hydrilla verticillata displayed its best absorbability in B3 at the 20th day However in that same day with the higher applying, B4 gave the lower uptake ability comparing to B3, meant that higher concentration of toxic compound can reduce plant uptake capacity (Phukan, 2015) When contacting and accumulating 40 too high level of heavy metals the plant absorbability was no longer working because of the contaminant fierce attacks to its physiological and biochemical activities, until it could not tolerate the effects from that metal anymore due to the breakdown of cell wall, membrane and vacuolar level or even the death of plant (Barcelóa & Poschenriedera, 2008; Chibuike & Obior, 2014) Consequently, Hydrilla verticillata in B3 of 10 ppm applied in the day 20th had the best absorbability since its tolerant capacity was at the maximum level The results which were shown after analyzing by variant analysis- one-way ANOVA with Duncan’s Multiple Range test gave the Pb treatment’s significant differences effects in water and Hydrilla verticillata (tables 4.3 to 4.11) (Bewick, 2004) In the results of affecting Hydrilla verticillata, treatments illustrated the significant difference effects corresponding to the changes in Hydrilla verticillata’s morphology, when heavy metals are up taken in plant they will influence all internal activities of the plant and morphological change is one of the impacts (Bini, et al, 2012) The 0th day, Hydrilla morphology in treatments stayed normal, in the 5th day with significantly different effect of B1 and B4, the plant started changing in leave appearances And longer time until the 15th and 20th day, the significantly different effects of Pb concentration to Hydrilla verticillata in treatments gave strong differences in effecting to the plant, with normal state of B1, leaves became faintly yellow state of B2, leaves became faintly yellow state of B3 and plant died state of B4 The Pb concentration of treatments also had the significant different of B4 throughout experiment time due to its high level in water According EPA (2014), the Maximum Contaminant Level Goal (MCLG) for Lead (Pb) is zero (0ppm), for this reason any water containing Lead (Pb) amount of higher then 0ppm is harmful and 41 hazardous environment for organisms Four observation times presented B4 as the only significant difference among treatments because its mean Pb concentrations always hold the highest place in water (table 4.3 to table 4.6) and it did seriously contaminate the water All the results proved Hydrilla verticillata ability in taking up and storing heavy metal especially Pb, by this reason Hydrilla verticillata is able to live in the contaminated water and clean the contaminants in water as well (Singh, 2012) Hence, in order to see these heavy metal absorbing potentials of this aquatic plant in a longer time of experiment, the study did use Linear Regression application to predict the increasing and decreasing trend of Pb concentration in water and Hydrilla verticillata All the results showed that in more long time Hydrilla verticillata still keep impressing its ability to remove Lead (Pb) from water environment by showing the upward trend of the regression line for Pb concentration in Hydrilla verticillata and downward trend of the regression line for Pb concentration in water With the goal of removing the concerned toxic compound, Lead compound (Pb2+), phytoremediation using Hydrilla verticillata would be the best decision (Singh, 2012; Moosavi & Seghatoleslami, 2013) In addition, Hydrilla verticillata has high biomass, thus its capacity to absorb heavy metals could pose high expectations for cleaning up or monitoring in large scales in future of improving water quality activities (Dixit & Dhote, 2009; Tangahu, 2011) The study were completed in demonstrating the ability in absorbing Pb of a submerged aquatic plant Hydrilla verticillata, for treatments with replications for each treatment which were observed in 20 days in times showed the average Pb absorbability is 92.8 % From these results, other studies which have common 42 questions of research can use as the reference materials However, the procedures of how Hydrilla verticillata absorbs and accumulates Pb element inside its body which have not implemented in this study could be the interesting research topic for other learners and researchers 5.2 Conclusions This study revealed the use of Phytoremediation technique to remove Pb 2+ in Pb(NO3)2 solution, a water contaminant with the submerged aquatic plant Hydrilla verticillata as a tool of Pb absorption By showing all the results of treatments (B1: Pb(NO3)2 ppm(control), B2: Pb(NO3)2 ppm, B3: Pb(NO3)2 10 ppm and B4: Pb(NO3)2 15 ppm) and replications for each treatment through times of observing in 20 days With specific high accumulated ratio of Pb content in the plant body up to 1.88 times comparing to applied Pb in water in treatment of 10 ppm, this higher interal concentration of Pb affected the plant absorbed ability and its growth, causing the decreases of Hydrilla verticillata absorbed performance when Pb applied increased At low applied Pb concentration of ppm, Hydrilla verticillata was completely absorbed 100 % of applied Pb, but the plant was influenced by higher level of Pb, in average 92.8 % Pb content in water was absorbed by Hydrilla verticillata Furthermore, these potentials were forecasted to be increased in future Hydrilla verticillata had clearly proved itself role in answering the research questions of this study about its ability in treating heavy mental- polluted water, namely Pb(NO3)2 which were specifically 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stress tolerance of plants South African Journal of Botany: 76 (2010): 167–179 Doi:10.1016/j.sajb.2009.10.007 51 THESIS ASSESSMENT REPORT I – INFORMATION OF SUPERVISOR Title and full name: Dr.-phil Arinafril Email address: arinafril@daad-alumni.de University’s name: Sriwijaya University University’s address: Indralaya Campus, Ogan Ilir 30662, Indonesia Phone and Fax : +62 711 580663 II – THESIS ASSESSMENT Thesis title: Hydrilla verticillata, a Submerged Aquatic Plant, as Phytoremediation Agent of Lead(II)-Nitrate (PB(NO3)2) Candidate’s name: Nguyen Thi Thao Assessment of thesis: Criteria Feedback Introduction OR Ability Good The student can bring this current issue to Articulate and Explain in many countries as the topic of her the Thesis Topic experiment But, some ideas are needed to be Grade /10 broadened and sharpened Literature Review OR The student could correlate her thoughts and Ability to determine, her knowledge with the supporting statements situate, and synthesise from any sources, such as textbooks and appropriate theoretical articles from journals related to this topic concepts within an But, the understandings to find appropriate overall conceptual supporting statements are needed to be framework enhanced 19 /20 52 Criteria Methodology OR Ability to determine, situate, and synthesise appropriate methodological concepts within an overall conceptual framework Results / Analysis OR Present scholarly Feedback Grade Methodology chosen was already good and common to be used for this study Steps to conduct this study were systematically ordered But, the knowledge of student in 19.5 /20 identification of aquatic plant is still limited, and also the knowledge of laboratory skill is needed to be improved The study findings were good as expected and could be scientifically accepted 19 /20 findings and argue claims Discussion & Conclusion OR Engage in reflective and conclusive scholarly argument Student can elaborate her thoughts and her knowledge in accordance with the results, and can enhance by corresponding them to current 19.5 /20 issues But, the in-depth of discussion must be extended by citing more sources Structure & Formatting: Student has shown her ability to structure and Ability to use structure organise her arguments which were shown and organisation to from her understandings in solving the cases develop a scholarly she proposed /10 argument AND Ability to Meet rigorous scholarly presentation standards 95/100 Overall mark Date: August 3, 2015 Supervisor (Sign) 53 [...]... Hydrilla verticillata, a submerged aquatic plant, as phytoremediation agent of Lead (II)-Nitrate (Pb(NO3)2) was carefully conducted and completed 5 1.2 Research’s objectives 1 To understand how the submerged aquatic plant - Hydrilla verticillata works as a tool of phytoremediation and how it got effect from Lead (II)-nitrate (Pb(NO3)2) polluted water 2 To assess the ability of Hydrilla verticillata in... Hydrilla verticillata In order to analyze the content of Lead (Pb) in Hydrilla verticillata, AAS- Atomic absorption spectrometry was used as an analytical technique which can identify and measure the concentrations of metal elements The procedures of analyzing Pb content in Hydrilla verticillata: 1 Took 50 grams of Hydrilla verticillata from each of 12 plastic containers and dried them in the 80o C incubator... by Phytoremediation with Hydrilla verticillata? 2 How is the absorbed capacity of Lead (Pb) of Hydrilla verticillata? 3 Whether Lead in water could completely be absorbed and accumulated by Hydrilla verticillata? Hypotheses: - Lead (II)-nitrate (Pb(NO3)2) can pollute water and affect Hydrilla verticillata growth - By using Phytoremediation with Hydrilla verticillata, Lead (Pb) can be removed from water... they are the scientific and theoretical basis for this research in assessing, 14 examining and investigating Hydrilla verticillata abilities to eliminate Lead (Pb) from water, absorb and store Lead (Pb) by its body and leaves 15 PART III METHODS 3.1 Sampling site and time This study about Phytoremediation of Lead (II)-Nitrate (Pb(NO3)2) from water with a submerged aquatic plant, Hydrilla verticillata... Hydrilla verticillata was selected with selecting standard are fresh, green and healthy plants, and washed by tab water from the laboratory Then take 12 samples (12 plastic containers: 4 treatments and 3 replications) of Hydrilla verticillata (which are already selected and washed), 300 gram for each sample, then put 12 of 300 gram samples into 12 plastic containers (diameter: 90cm, height: 30cm), which already... started from March 2015 and finished in June 2015 On 09th March 2015, harvesting Hydrilla verticillata for the experiment, the total of 3 plastic containers of Hydrilla verticillata was taken from submerged waters around Ski Air Lake Jakabaring Palembang, Indonesia Then these 3 plastic containers were moved to integrated research for the Integrated Research Laboratory of Sriwijaya University Palembang,... concentrations of elements)  Plastic containers  pH meter  Hot plate, incubator  Analytical balance, mortar, rod stirrer, pipette  Glass funnels, glass conical flasks, bunchner flask, measuring cup, vacuum filtered set of tools, glass sample bottles 18 3.3 Methods 3.3.1 Applying treatments After moving to Integrated Research Laboratory of Sriwijaya University Palembang, Indonesia, 3 plastic containers of Hydrilla. .. ANOVA one-way analysis in statistics software 8.0 of statistical significance α = 0.05 was assumed Whenever analysis of variance showed statistically significant effect (p < α = 0.05), Duncan test was performed to determine the significant differences more accurately 3.4.2 Lead (Pb) Reduction rate The Mean reduction rate of Pb concentration in water was calculated based on the initial and final mean Pb... professional literatures, which are relevant to this study, play important roles in understanding the reason why water envronments need to be treated, which ways gave good resuls and which ways will have high potential of using them The literature reviews have raised background information about Hydrilla verticillata as an useful and effective phytoremediation agent of Lead (II) - nitrate (Pb(NO3)2) And also,... mean amount of Pb left in B3 is just really little 0.06 ppm From the 0th day until the 20th day Pb content in water B2 is really low, and also increased in 10th day and 15th day but until the last day of experiment- the 20th Pb concentration in this treatment is removed all from water 4.3 Result of Data analysis 4.3.1 Variant analysis After analyzing Data by Variant analysis and continued with Duncans