Biosorption of Lead and Zinc by microalgae Scenedesmus sp.
Shanshan Wang, Mingzi Wang, Jian Huang, Yanqiu Fu, Min Li, Bilian Chen
（Engineering Research CenterCentre of Industrial Microbiology, Ministry of Education；, School of Life Science, Fujian Normal University, Fuzhou 350108, China）
Abstract Concerning the heavy metals uptake capacity, Scenedesmus sp. was proven to be successful as an efficient adsorbing material based on its heavy metal uptake capacity after being subjected to heavy metals metal (Pb and Zn) stress, the. The presence of them produced an effect onheavy metals affected numerous metabolic or developmental processes. The experiment in the microalgae. The effects of different heavy metal concentrations on several experimental parameters of the cells, such as the growth rate, chlorophyll a (Chl a) content, and raw fatty acid content of the cells, were determined under different concentration of each heavy metal respectively. . The results showed that Scenedesmus sp. had morehas higher tolerance to Pb and can be survivalsurvive in up to 20 mg/L. The growth of the metal. Scenedesmus sp. growth was inhibited by heavy metals when their and its biomass and Chl a concentration decreased at certain ranges of heavy metal concentrations. When the Zn concentration reached to a certain range as well as to gain a lower biomass and chlorophyllexceeded 5 mg/L, the Chl a concentration. When the concentration of Zn was above 5mg/L, the accumulation of chlorophyll a had a sharp drop to 1mg/L sharply decreased to 1 mg/L. The cells got aexhibited better growth with a concentration of and Chl a contents increased when the Pb and Zn under 1 mg/L, and chlorophyll a content increased simultaneously. Supplied with 0.5 mg/L of Pb in the concentrations were below 1mg/L. When the medium contained 0.5 mg/L Pb, the maximum biomass achieved was 0.642 g/L, while the raw. Raw fatty acid production increased with the increasing Pb concentration of Pb increasing. The main composition of the fatty acid wereacids were determined by gas chromatography–mass spectroscopy and mainly consisted of methyl palmitate, methyl oleate, methyl linoleate, and methyl linolenate, which analyzed by GC-MS.. Scenedesmus sp. reflected aexhibited higher metal sorption capacity thanfor Zn after a period of 8 days,d, reaching maximum amounts was up to values of 35.961 mg/g and 23.812 mg/g dry weight when exposed to 20 mg/L and 8 mg/L conditions. Biological multiply rate of Zn was higher compared with Pb, and that, respectively. The biological multiplication rate reachedof Zn was higher than that of Pb, reaching 4.07 underat 0.5 mg/L concentrationZn.
Key words : Pb, Zn, Scenedesmus sp., growth, metabolism, biosorption
1. Introduction
Along with the Industrial development of industry, heavy metal is a member ofhas been accompanied by the widespread use of heavy metals, which are elements that exhibit metallic properties, these effluents as a result of . Unfortunately, heavy metal effluents resulting from the extensive use of industrialization wereindustrialisation that are left untreated and subsequently discharged into the environment, which had been polluted inordinately. Because of the toxic and unbiodegradation have caused widespread pollution. Considering the toxicity, nonbiodegradability, biological accumulation, and continuelong retention time forof heavy metalmetals [1], it had been followedtheir levels are closely monitored by national and international authorities on accountbecause of a serious oftheir deleterious effects had been brought about to on human health and can have on the surrounding environment, consequently taking action indeed that aimed at minimizing their impact. In addition, actions that aim to minimise the effects of these elements have been taken.
It exposed the high cost and a limited effectiveness, when processing capacity of aqueous solution was large and concentration was present at very low level to a great extent, if it just sticked with traditional Traditional physicochemical methods applied to removefor removing heavy metals, including precipitation, solvent extraction, ion exchange, and membrane separation and so forth[2]. Removing heavy metals as well, are expensive and have limited effectiveness, particularly when used to process large volumes of aqueous solutions or when dealing with trace heavy metal concentrations [2]. By contrast, heavy metal removal using biosorption was characterized as a kind of high efficiency andis highly efficient and involves inexpensive treatment technologies. Microalgae qualified as a classical , which are considered traditional biosorbents, can survive in most sortstypes of environment systemenvironmental systems and adaptedcan adapt to different environmentenvironmental conditions [3] was appropriate to afford a superiority]. The use of microalgae as precise bioindicators is superior to conventional processing methods, that’s one of a precise bioindicators is that because of their distribution, size, longevity, presence atin pollution sites, ability to accumulate acceptable amounts of metals to a satisfactory degree, and ease of identification [4]. There have been many examples on the characteristics of toxicity effectA large number of cases demonstrating the toxic effects and physiological changes resultedresulting from heavy metalsmetal exposure have been reported [5-–9]. Their effect was that heavyHeavy metals interactedinteract with the sulfhydryl of enzyme within the organism and formed precipitation which resulted ingroup of enzymes within organisms and form precipitates, thereby inhibiting theenzyme activity of enzymes sequentially[[10]. On the other hand,Moreover, heavy metal-induced redox reactions disrupt photosynthesis and the activity of antioxidant system activities [11] were disturbed by the process that some redox reaction was induced and aggravatedand promote the generation of reactive oxygen species.
A key challenge toin using microalgae as a remediation option wasmaterial is that many a large number of microalgae wereare sensitive to the heavy metal toxicity of heavy metal in spite of extremely low content, adsorptioneven at trace concentrations. Adsorption (electrostatic attraction)[) [12] and complexation [5] were proceededoccur through the cell wall, because of the high metal -binding capacities due to the presence of polysaccharides, proteins, or lipidlipids on the cell wall surface containing. These molecules contain functional groups such as amino, hydroxyl, carboxyl, and sulphate groups, which can act as metal-binding sites for metals[2].
Scenedesmus consisted, which generally consists of commonly 2, 4, or 8 single cell wascells, is a common species of chlorophyta found in shallow lakes, which was. These algae are particularly useful in uptaking heavy metal owing to the characteristicsuptake because of their rapid propagation, stronghigh adaptability, and synthetic ability ofto synthesise fatty acids [13], furthermore]. Furthermore, Scenedesmus asare autotrophs had the photosynthesis and its and thus have photosynthetic abilities. Their large specific surface area and affinity for heavy metal cations [14] enjoyed anare added advantage overadvantageous in the detection, prediction, control, and eliminateelimination of heavy metalmetals from aqueous solutions. Heavy metals The effects of heavy metal toxicity to on aquatic organisms dependeddepend on many physical and, chemical and biological factors, therefore, the. Therefore, other environmental factors onthat affect Scenedesmus growth and, metabolism, adsorption capability and the, optimum condition of adsorption, conditions and adsorption mechanism remains tomust be explored further. The majormain purpose of this research was focused on testingis to investigate the ability of Scenedesmus to remove Pb and Zn from the point of application research. Additionally, water. Changes in the growth and metabolism of Scenedesmus were also estimated when exposed to various initial heavy metal concentrations were also determined.
2. Material and methods
2.1Microalga1Microalgal and culture conditions
GreenThe green microalga Scenedesmus sp. was obtained from the Institute of Hydrobiology, Chinese academyAcademy of Science FACHB-Collection. The experimentDetermination of microalga to uptakemicroalgal Zn and Pb uptake was proceededperformed in 500 mlmL Erlenmeyer flasksflasks containing 200 mlmL of BG-11 medium, and the. Aqueous heavy metal aqueous solutions with the intendedspecific concentrations waswere added to the flasks under controlled conditions. Each experiments consisted of experiment was conducted in triplicate with the purpose of obtaining more to obtain reliable results, a. A blank control contained containing the culture medium subtract metal(but without the metals) was also establishedprepared for each concentration. Inocula forThe inoculae of all experimentalexperiments were obtained from the exponential phase which based on optical density (this status was reachedusually achieved after 2–4 daysd of growth)and inoculation amount was 20%(in order to). To achieve the same initial optical density, more specifically, optical density lied (more specifically, between 0.11 and 0.12) then daily for a throughout the experimental period of (8 days .d), the daily inoculation amount was set to 20%.
The strain was cultivated and maintained at 25 °C under a constant fluorescent light exposure intensity of 3000 lux in a reciprocal shaker at 100 oscillations ·min-1.
2.2 Measurement of growthGrowth rate and biomass measurements
Cell growth was determined by measuring the absorbance at 650 nm in ausing an ultraviolet and –visible light spectrophotometer every 2 days,d. A growth curve reflecteding the growth status of Scenedesmus sp. exposed after exposure to different initial concentrationmetal concentrations was createdprepared according to the absorbance. At the end of the experimental period 15ml, 15 mL samples were takencollected from each flaskflask after exposure of the microalgamicroalgal cells to metal solutions for a period of 8 days, thed. The stability of the biomass subsequently collected by centrifugation (6000rpm,10min6000 rpm, 10 min, 4℃)°C) was evaluated at different initial metal concentration, wetconcentrations. Wet algal cells were rinsed with deionized water three times with deionised water and converted to dry weight (DW). The rest ofremaining samples were lyophilizedlyophilised for the follow-upsubsequent operational testexperiments.
2.3 Biosorption experiments
A stockStock solutions of Zn (10.0 g/lL) and Pb (20.0 g/lL) were prepared by dissolving the corresponding metal salt [analytical-reagent grade ZnSO4·7H2O and Pb (NO3) 2(analytical reagent) ] in deionized waters.deionised water. All glassware material employedused was previously rinsed with hydrochloric acid and cleaned off with deionized water washed several times prior to use so as with deionised water to guarantee the absence of any interferent with the chemical analyses afterwards.. All mediummedia and experimental materials were previously autoclaved at 121℃ °C and 1.01×105 Pa for 20 min.
ProperAn appropriate digestion solution consistedconsisting of a HNO3 and HClO4 mixture ( V/Vv/v=3:1) were was added dropwise added into exact weights of each algae sample (0.05g powders05 g powder) placed intoin acid -washed digestion tubes and dissolved for 30approximately30 min approximately, bathing. The bath temperature should bewas kept at approximately 80℃ or so, it was not resolved°C. The mixture was considered completely until resolved when the liquid showed up colorless andbecame clear and that the colourless. The contents were then evaporated to near dryness. ConstantA constant volume was processedmaintained on the spot with by addition of double -distilled deionizeddeionised water up to 50ml50 mL followed by cooling. The solutions were filtered through 0.22 ummm Millipore filters. The standardStandard stock solution forsolutions of Pb and Zn werefollowed the national criteria solution (NCS) with the concentrationconcentrations of 1.000 g/L (State Test CenterCentre of Ferrous Materials), the). Aqueous metal aqueous solutions came from itswere prepared through further dilution todilutions of the standard working solutionsolutions. The absorbances of Pb and Zn were measured by aan AA-240 atomic absorption spectrometer equipped with flame atomizationa flame atomiser at λ= 217.0 nm andλ=and λ=213.9nm9nm, respectively. The detection limitlimits of Pb and Zn were 1-30mg–30mg/L and 0.05-1mg–1mg/L for Pb and Zn,, respectively. The dilution ratio was determined toratios of the aqueous solution ofsolutions containing different metal concentrations were determined for metal detection of metal. In addition, the concentration of metals wasmetal concentrations were calculated and expressed in milligram per gram of dry massDW in line with the standard working curves.
2.4 Determination of chlorophyll a and raw fatty acid concentrations
For the concentration ofTo determine chlorophyll a determination(Chl a) concentrations, Scenedesmus sp. cultures were collected by centrifugation (4000rpm, 10min4000rpm, 10min) of 5 mlmL culture samples, the. The algal pellets were added rewashed with 5 mlmL of distilled water again to removedremove residual nutrients in the medium which might be that may affect the measurement accuracy of the pigment concentration. The samples contained 5ml Samples containing 5mL of 90% acetone waswere superior to other extracts for pigment extraction to maximize chlorophyllmaximise Chl a extraction and were homogenized 1 min with thus selected. These samples were homogenised in a vortex mixer. Then for 1 min. Afterwards, the samples were extracted overnight at 4℃°C and was going to be oscillating oscillated at regular intervals during the course ofentire processing procedure [15]. The absorbance of the extract was determined at the wavelength of 646 and 663 nm withusing a spectrophotometer, respectively. Chlorophyll a presented. The Chl a concentration in the extract werewas calculated derived fromusing the equations:equation Cchla (mg/L) = (12.21×A663－2.81×A646) ×5 ml / 5 mlmL/5 mL.
The rawRaw fatty wasacids were extracted by the method of using an improved chloroform/methanol extraction. 5 ml method. A total of 5mL of 5% potassium hydroxide (with 30% methanol solution) was put intoadded to 0.1 g of algae powder. The sample werewas sufficiently oscillated enough to made sure that they could makeensure full contact with each other effectivelybetween particles and put itthen set aside for 20 min, the. The studies were performed at 70℃ to be representative of environmentally °C for 5min to simulate environmental conditions for 5 min. That the subsequent accumulation. Accumulated samples were centrifuged after cooling to ambient temperature, there was a need to repeat the operation steps. The extraction procedures were repeated until the chlorophyllcomplete removal of Chl a and other pigment were removed completely.pigments. The resulting pellets were added to 3 mlmL of 4 mol/lL hydrochloric acid and kept static. Mixtures were left to stand for 30 min to separate intopromote the separation of two phases, then heated on thea boiling water bath for 5 min and freezed at -20℃then rapidly. Following frozen at –20°C. After 20 min of reaction of the mixture combined withbetween these mixtures and a chloroform and methanol mixture (V/Vv/v=2:1)), the samples were collected by centrifugationcentrifuged (5000 rpm, 10 min) [16],]. The chloroform layer in whichcontaining the raw fatty dissolvedacids plus mingled withan isometric 5% NaCl solution andwas centrifuged under the same conditions aspreviously described detailedly above, chloroform layer were vacuumized andconditions, vacuum-dried, and then weighed.
2.5 Fatty acidsacid composition analysis