Evaluation of Acid Leaching on the Removal of Heavy Metals and Soil Fertility in Contaminated Soil

Heavymetal-contaminated soils were leached with various acid reagents, and a series of treatments was assessed to understand soil fertility after acid leaching. Aqua regia digestion and a five-step sequential extraction procedure were applied to determine heavy metal distribution. *e average total concentrations of Zn, Cd, Cu, and Pb for contaminated soil were 1334, 25, 263, and 525mg·kg based on the ICP/AES quantitative analysis. Other than Pb extracted by H2SO4, over 50% removal efficiency of other heavy metals was achieved. A five-step sequential extraction revealed that the bound-to-carbonate and bound-to-Fe-Mn oxides were the major forms of the heavy metals in the soil. *e addition of organic manure considerably promoted soil fertility and increased soil pH after acid leaching. Seed germination experiments demonstrated that after acid leaching, the soil distinctly inhibited plant growth and the addition of manure enhanced seed germination rate from 35% to 84%. Furthermore, the procedure of soil turnover after acid leaching and manure addition greatly increased seed germination rate by 61% and shortened the initial germination time. Seed germination in untreated soil was superior to that in acid-leached soil, illustrating that the phytotoxic effect of acid leaching is more serious than that of heavy metals.


Introduction
Soil pollution is a serious environmental issue worldwide [1,2].Heavy metals are considered a serious soil pollutant because of their toxicity, persistence, and nondegradable characteristics in the environment [3][4][5].In addition, the self-purification process of heavy metals present in contaminated soil is slower than that in contaminated air and water; thus, inadequate treatment of contaminated soil can pose a great human health risk.In Taiwan, more than 50,000 hectares of farmland has been contaminated by heavy metals to the fourth level classification, and more than 160 sites have been identified to be contaminated by heavy metals.Surveys state that approximately 785 hectares of farmland is contaminated by zinc to the fifth level classification [6].
e Er-Jen River located in the south western Taiwan is one of the most seriously contaminated rivers, caused by the waste from metal reclamation, smelting, and disposal of motors and electrical capacitors that released a large amount of heavy metals, polychlorinated biphenyls (PCBs), and polychlorinated dibenzo-p dioxins and dibenzofurans (PCDD/DFs) into the water [7].Due to lack of environmental consciousness during early agricultural cultivation, the Er-Jen River was used as an irrigation source, which caused a detrimental heavy metal contamination to the paddy field.erefore, remediation of the contaminated farmland near the Er-Jen River is of high priority.
Although acid leaching is an effective treatment for heavy metal removal from soil, it severely damages soil fertility and its use for remediating farmland is timeconsuming.
e rapid enrichment of soil fertility after acid leaching is increasingly becoming a topic of concern for farmers.erefore, this study used acid leaching techniques to remove heavy metals from contaminated farmland that can be helpful in understanding the mobility and bioavailability of metals in contaminated soil and to investigate the extraction efficiencies of heavy metals.In addition, the assessment of soil fertility and seed germination tests after the acid leaching procedure were investigated to understand the condition of remediation in the contaminated farmland.

Experimental
2.1.Sampling Area.Soil samples were collected from the top layer (0-30 cm) of a paddy field near the Er-Jen River in Tainan city, Taiwan province, China, which has been recognized as a previous history of irrigation with metals-rich wastewater.
e paddy field contaminated with heavy metals has been strictly prohibited cultivating.Soil samples were collected by mixing ten subsamples from each site within a 50 × 50 meter area and were recorded for the central point position by using GPS equipment.
e soil samples were air-dried at room temperature and then were ground with an agate mortar to pass through a 2 mm sieve.After sieving, the soil samples were experimentally pretreated by a vortex mixer with 100 rpm for 24 h to ensure the complete mixing of the samples.

Chemical Analysis.
Soil properties, including soil pH, organic matters, soil texture, and cation exchange capacity, were determined.Particle size distribution was obtained by the pipette method after removal of carbonate, organic matters, and MnO 2 .Carbonate was removed by 1 M NaOAc with a pH 5 at 60 °C, and organic matters and MnO 2 were digested by 30% H 2 O 2 [16].e soil pH value was measured on a mixture of 1 : 1 soil/deionized water by a glass electrode [17].Organic matter content was determined by the Walkley-Black wet oxidation method [18].Cation exchange capacity was determined by the ammonium acetate method at pH � 7 [19].Total content of heavy metals in soils was extracted by aqua regia digestion with a volume ratio of 1 : 3 HNO 3 /HCl at 120 °C for 8 h.e suspension was filtered through a 0.2 μm filter paper after the digestion process and was placed into precleaned 100 ml volumetric flasks with 0.5 M HNO 3 for analysis.e contents of heavy metals were analyzed by inductively coupled plasma atomic absorption spectrometry (ICP/AES).To preclude unexpected contaminations in the analysis procedure, all equipment used were washed with a phosphate-free soap and cleaned with deionized H 2 O. e detection limits for metals were as follows: Cd (0.05 mg•kg −1 ), Zn (0.02 mg•kg −1 ), Pb (0.06 mg•kg −1 ), and Cu (0.05 mg•kg −1 ).e percentages of recoveries for each metal were ranged from 96.7% for Cd to 103.2% for Zn. e standard stock reagents (Merck, Germany) containing 1000 mg•kg −1 of metals were used.Calibration reagents containing between 0.5 and 5000 mg•kg −1 of Cd, Cu, Pb, and Zn in 3% HNO 3 were daily prepared.

Acid Leaching Experiment.
In this study, five acid reagents were taken into account for leaching heavy metals from contaminated soil, including 0.05 M HNO 3 , HCl, H 3 PO 4 , and H 2 SO 4 .A liquid-to-soil ratio of 1:10 was adopted by adding 5 g of soil with 50 mL above acid reagents in a glass vessel, and they were continuously stirred at 100 rpm for 5 h.After leaching experiments, the suspension was centrifuged at 3500 rpm for 30 min and then was filtered through a 0.2 μm membrane filter for metals analysis.e contents of metals were analyzed by ICP/AES.To reduce the systematic errors, duplicate measurements were carried out for each sample, and the average values were determined.A five-step sequential extraction procedure used in this study was developed by Tessier [20].e five fractions of bound heavy metals included the following: exchangeable form (EXC) was extracted with 1 M NH 4 OAc at pH 7, carbonates form (CAR) was extracted with 1 M NaOAc at pH 5, Fe-Mn oxide form (MNO) was extracted with 0.04 M NH 2 OH•HCl in 25% HOAc, organic matter and metals associated with easily oxidizable solids or compounds (OM) were extracted with H 2 O 2 (30%) at pH 2 and 0.02 M HNO 3 , and residual form (RES) was extracted with a 5 : 1 mixture of HF-HClO 4 .All extracted reagents were centrifuged for 30 min at 10000 rpm and filtered with a 0.2 μm Teflon filter.e suspensions were analyzed by ICP/AES.Soil particle size is an important factor to affect removal efficiency of heavy metals.However, it is difficult to screen soil particle size when the in situ remediation is throughout used to a real heavy metal-contaminated site.erefore, the effect of soil particle size is not mentioned in this study.

Seed Germination Experiment.
For the germination tests, seeds of cabbage (Brassica chinensis L.) were used because this species is a general vegetable and can be popularly seen for a whole year.Twenty-five seeds of cabbage were sowed in a sterilized individual tray (15 × 20 × 3 cm 3 ) containing 1200 cm [3] of modified leached soils.Duplicate of the seed germination experiment was simultaneously conducted.e trays were placed in a greenhouse at 25-28 °C and 70% humidity.e experiment was examined daily for 30 days, and seeds were recognized to have germinated when the cotyledon was observed.After cotyledon appearance, the root lengths in each tray were measured and recorded every 12 h.e untreated soil sample was also conducted under the identical condition for comparison.To assess the seed germination criteria, the percentage of relative seed germination was chosen to understand the seed germination condition [21].

Results and Discussion
e physical and chemical properties of the soil samples along with their total heavy metal concentrations are listed 2 Journal of Chemistry in Table 1.e average total concentrations of Zn, Cd, Cu, and Pb were 1334, 25, 263, and 525 mg•kg −1 , respectively, and their maximum concentrations of Zn, Cd, Cu, and Pb were 4852, 46, 1470, and 1430 mg•kg −1 , respectively.e total heavy metal concentrations were not only higher than the tolerable limits recommended by the World Health Organization (WHO) and European Union (EU), but also exceeded the Chinese standard for agricultural soil [22]. is contaminated site needs to be appropriately treated to avoid heavy metals moving into deeper layers and causing phreatic zone pollution.As shown in Table 1, the organic matter content is much lower than the normal content.
is is because that the Er-Jen River is contaminated by metal reclamation, smelting, and electrical capacitors and results in a high content of heavy metal accompanying a low pH value.Due to long-term irrigation from Er-Jen River, it is believed that the organic content of soil in this site is much lower than that of the uncontaminated site.e removal e ciency of heavy metals by various acid reagents is shown in Figure 1. e leaching experiments indicated that Zn and Cu had the highest removal e ciency and more than 70% removal e ciency was achieved for all acid reagents.For the Cd species, regardless of the acid reagents used, the removal e ciency ranged from 55% to 66%, indicating a high chemical a nity between the Cd and the soil, yielding lower removal e ciency for all acid reagents.A sequential extraction method was used to determine the Cd distribution in the surface sediment and found the major binding structure between Cd and soil was attributed to a residual form [23].Among the four acid reagents, HNO 3 had the lowest Cd, Zn, and Cu removal e ciency, whereas H 2 SO 4 had the highest Cd, Zn, and Cu removal e ciency; this result was not observed for Pb.To further understand the speciation distribution of heavy metal in soil, the chemical sequential extraction was used and is shown in Figure 2. e highest Cd concentration was associated with the RES fraction, whereas the EXC had the lowest concentration.e presence of Cd in the RES fraction was consistent with the result obtained by Mendoza et al. in which their report showed that Cd was distributed predominantly in the residual form with a value of 45% in the contaminated soil [24].us, the Cd retained in the crystal lattices of minerals has a strong bond and consequently is not easily released into the environment.e overall percentage of Cd in the fractions was in the order: RES > OM > CAR > MNO > EXC.Unlike Cd, the largest portion of Zn was observed in the CAR fraction (39%) and followed in the descending order of EXC > MNO > OM > RES.
is fraction could be easily  made exchangeable under conditions such as pH change [25].Pb and Cu demonstrated an identical trend in that CAR and MNO appeared to be the major fraction.e proportion of heavy metals bounds to Fe-Mn oxides was highly variable and depended on the depth of the water and redox reactions with the soils.e redox cycle is important in controlling Fe and Mn in most aquatic and soil systems.However, it varies seasonally in summer, when the oxygen between the water and soil interface often decreases.is causes the reduction of Fe 3+ and Mn 4+ into soluble Fe 2+ and Mn 2+ , which is transported upward in the water column and oxygenated, resulting in the reoxidation of insoluble metals to settle the soil and repeat the cycle [26].ese forms of metals are potentially dangerous for plants because they can transform into being bioavailable under environmental changes [27].Organic matter (OM), nitrogen (N), potassium (K), and phosphor (P) are important substances for vegetable and plant growth in soil.To understand the changes in these substances after acid reagents leaching, OM, total N, total K, and total P contents were determined and are depicted in Figure 3. e content of OM in the soil decreases by 18.5% and 8.74% when H 3 PO 4 and H 2 SO 4 were used for pickling, respectively.No signi cant change in the OM content for HNO 3 and HCl was observed.e total N content increased  e result of changes in total K is also depicted in Figure 3(c).e soil K concentration changed because of acid liquid precipitation.HNO 3 reduced the total K content by 26.2%, followed by others in a descending order HCl > H 3 PO 4 > H 2 SO 4 .Commercial organic manure was used after the leaching treatment to fertilize the soil.Commercial organic manure contains 21% OM, 4.2% total N, 4.1% total P, and 3.5% total K.Most manure containing considerable amounts of soluble salts increases soil's electrical conductivity (EC), resulting in damage to plants.e result of the experiments showed that the EC increased after manure treatment and reached stability after 30 days.Over 30 days, the EC ranged from 0.86 to 0.41 mS•cm −1 , an acceptable range for plant growth.Figure 4 shows the changes in soil pH as a function of duration after HCl leaching and fertilization by organic manure.e initial pH for original and leached soils was 7.6 and 6.3, respectively.e acid leaching treatment reduced the soil pH consequentially caused an e ect to crops cultivation.e pH increased with the duration and reached a maximum of 7.02 at 30 days; it then gradually decreased and eventually stabilized to approximately 6.3.
e addition of organic manure signi cantly improved soil pH over a short period; however, in the long-term, the soil pH returned to a meta-acid state, which was probably associated with the soil bu ering power.Figure 5 shows the changes in the OM, total N, total P, and total K contents as a function of the duration after HCl leaching and fertilization treatment.
e identical feature could be observed after the addition of organic manure, which increased the OM, total N, total P, and total K content within 30 days.Over the rst 15 days, the OM and total N contents became more than twice of this manure addition, and a steady state was gradually achieved.However, the total K content rapidly increased with the duration with the highest content value of 2.5% within 5 days.On the basis of these ndings, the organic manure additive improves soil fertility within a short period.To assess the suitability of farmland for cultivation, a seed germination experiment was performed to evaluate suitability for planting after various treatments.Seeds were cultivated in an incubator and observed daily for germination with the seed germination rate as a function of time at various treatments presented in Figure 6.
e lowest germination rate was observed when soil was treated through acid leaching because after acid leaching the soil fertility decreased and a ected plant growth.On the other hand, initial seed germination occurred earlier than the other treatments.Although heavy metals were moderately removed after acid leaching, the nutrient loss was a major barrier to seed germination.After the addition of organic manure, the seed germination rate considerably increased by more than 80% and the initial seed germination time was four days, earlier than that of the experiments assessed after acid leaching.Notably, the seed germination rate in untreated soil was superior to that in acid-leached soil. is demonstrates that the inhibition e ect of acid leaching to the plant is more serious than that of heavy metals, and the addition of organic manure could be a suitable approach to improve soil fertility and plant cultivation.To obtain a higher seed germination rate, the procedure of soil turnover was applied simultaneously.As shown in Figure 6, the procedure of adding manure to soil turnover rapidly increased the seed germination rate and initial seed germination time, and a seed germination rate of approximately 85% was achieved.e procedure of soil turnover diluted heavy metals and acidic species and thus resulted in the redistribution of these species.
Although seed germination rate is a suitable indicator to evaluate soil fertility, heavy metal concentration in test plants should be considered to ensure food safety.erefore, cabbage roots and leaves were collected and analyzed to understand heavy metal distribution.e experimental result is presented in Table 2. e major uptake was found in the root of cabbage, and the fraction of Cd, Zn, Pb, and Cu uptake by the root was 46.8%, 51.3%, 17%, and 25.4%, respectively.is nding is in agreement with the observation of Shi et al., who reported that heavy metal concentration is much higher in the root than in the shoot and leaf [28].e root is the rst mode of entry for heavy metal pollutants from the soil into the plant.Nevertheless, it excretes organic acids, amino acids, sugars, and growth substances into the rhizosphere, a ecting metal adsorption.By changing the physiochemical properties of the rhizosphere, the heavy metals will be detained in the roots through chelation, complexation, and disposition with root exudates.Notably, in this case, the Pb uptake by the root was the lowest, whereas a much higher fraction was noted for Zn; the leaf demonstrated the same result.Pb and Zn are essential elements for plants growth, but their excessive amounts in soil lead to the inhibition and poisoning of plants.e normal Pb, Cu, and Zn contents in plants are 0.5-10 mg•kg −1 , 3-30 mg•kg −1 , and 10-150 mg•kg −1 , respectively, whereas their toxic concentrations to plants are 100-400 mg•kg −1 , 70-400 mg•kg −1 , and 60-125 mg•kg −1 , respectively [29].Here, the Pb, Zn, and Cu concentrations in bulk cabbage were 120.23 mg•kg −1 , 283.52 mg•kg −1 , and 37.14 mg•kg −1 , respectively.Except Cu, Pb and Zn concentrations were slightly higher than the aforementioned range, indicating the soils after acid leaching and organic manure addition remain a risk when used to cultivate plants.e procedure of soil turnover may be a suitable approach to overcome this obstacle.

Conclusions
A series of acid leaching tests and soil fertility assessment for contaminated soil containing Cd, Pb, Zn, and Cu were performed to investigate leaching behavior.Most heavy metals could be removed through acid leaching with a nearly 60% removal e ciency achieved.e major species for Cd, Zn, and Cu were distributed in residual, bound-tocarbonate, and bound-to-Fe-Mn oxides forms, respectively.e addition of organic manure distinctly increased soil nutrient content as well as soil pH within a short period.Seed 6 Journal of Chemistry germination in untreated soil was superior to that in acidleached soil, revealing that the phytotoxic e ect of acid leaching is more serious than that of heavy metals.

Figure 1 :Figure 2 :
Figure1: Removal e ciency of heavy metals by using various acid reagents.

Figure 3 :
Figure 3: e changes in content of (a) organic matter, (b) total N, (c) total K, and (d) total P after leaching by various acid reagents.

Figure 4 :
Figure 4: e change in soil pH as a function of the duration period after HCl leaching and fertilized by adding organic manure.

Figure 5 :
Figure 5: e changes in contents of (a) organic matter, (b) total N, (c) total P, and (d) total K as a function of the duration period after HCl leaching and fertilized by organic manure.

Figure 6 :
Figure 6: Seed germination rate as a function of time at various treatments.

Table 1 :
e brief description of contaminated soil used in this study.