Bio-availability of heavy metals
DTPA-extractable bioavailable fraction of heavy metals was varied across different stages of incubation for various fly ash-soil-ameliorants combinations used in this experiment (Fig. 2). The extractable amounts of Pb, Cd, Ni, and Cr significantly differed for different treatment combinations after 60 days of incubation (Fig. 3; A1-D1) and a noticeable reduction in metal concentration was detected in different combinations. A1T1 recorded the lowest value for bio-available Pb (1.07mg kg-1; ~49% lower), Cd (0.05mg kg-1, ~171% lower), Ni (0.11mg kg-1, ~233% lower), and Cr (0.06mg kg-1, ~65% lower) whereas A3T0 recorded the highest (2.24, 0.26, 0.65, 0.22 mg kg-1respectively for the same; ~29-55% higher) among the treatments. On an average, the amount of DTPA-extractable Pb, Cd, Ni, and Cr was increased with the increased proportion of fly ash in fly ash-soil combinations (A3> 1.09 A2> 1.26 A1, A3> 1.28 A2> 1.59 A1, A3> 1.16 A2> 1.46 A1, and A3> 1.15 A2> 1.31 A1 for Pb, Cd, Ni, and Cr respectively) irrespective of ameliorants applied (Fig. 3; A2-D2). However, the efficacy of ameliorants in reducing the bio-availability of heavy metals was varied significantly within themselves. In this regard, T1 (mean ~1.26, 1.51, 1.83, and 1.65 times lower for Pb, Cd, Ni, and Cr, respectively) retained the lowest amount of bio-available HMs followed by the variant T3 (mean ~1.35, 1.70, 0.80, and 1.24 times lower, respectively, for the same) among all the treatments.
Changes in DTPA-extractable heavy metal to total heavy metal ratio under different fly ash-soil-ameliorant combinations
After 60 days of incubation, the ratio of DTPA-extractable to total heavy metal concentrations for Pb, Cd, Ni, and Cr was varied from its initial across the interventions imposed (Fig. 4). On average, a declining trend was observed in the ratio for all the heavy metals with the application of different ameliorants (mean~0.009, 0.003, 0.004, and 0.001 for Pb, Cd, Ni, and Cr respectively). All the treatments comprised of A1 i.e. fly ash-soil (50%+50%) combination showed a greater declining trend as compared to A2 (fly ash-soil: 75%+25%) and A3 (fly ash: 100%), respectively, irrespective of the heavy metals considered (A1>2 A2> 5 A3). Overall, A1T1 (0.01) followed by A1T3 (0.009) recorded the highest decrease whereas A2T0 (-0.001) followed by A3T0 (-0.0006) recorded the lowest among the treatments. In most cases, these ratios for different metals were highly correlated with their respective bioavailable fraction than the total metal load under all the treatments (Table 4).
Efficacy of different fly ash-soil-ameliorant combinations in minimizing bio-availability of heavy metals
Among different treatment combinations, a greater decrease of Pb and Cd by A2T3; Ni by A1T1, and Cr by A2T1 in the tune of 0.89, 0.15, 0.46, and 0.07 mg kg-1 was achieved after 60 days incubation. However, considering the four heavy metals, the total decrease was highest for A1T1 (1.43 mg kg-1; ~226% decrease) followed by A2T1 (1.36 mg kg-1; ~126% decrease) averagely whereas A3T0 (0.26 mg kg-1, mean ~10% increase) recorded the lowest (Fig. 5). Overall, among the treatments, the trend in decreasing the bio-availability of heavy metals observed in this experiment was: A1T1> A2T1> A2T3> A1T3> A2T2> A1T2> A3T1> A3T3> A3T2> A1T4> A3T4> A2T4> A1T0> A2T0> A3T0.
PCA results showed that ~97% of the total variation in declining the bioavailability of HMs for different fly ash-soil-ameliorant combinations was explained by the first three components (Table 3). Principle component 1 (PC1) explained 73% of the total variance having a significant loading on each of the variables. The highest loading was attained by the A1T1 treatment combination in PC1 (PC1 loading on A1T1: 2.32). The second (PC2) and third principal components (PC3) explained only ~18 and 6% of the total variance respectively and loaded highest on A2T3 and A1T0.
PCA-scatter plot distinctly represented the distribution of observations corresponding to different treatment combinations and resulting interactions between the two main components (Fig. 6). The treatments positioned on the positive X-axis performed well over others in decreasing the bio-availability of HMs and the treatment with the highest positive value was considered the best. Accordingly, A1T1 followed by A2T1 performed the best whereas A3T0, the poorest in this experiment.
Evaluation of environmental risk
The efficacy of different amendments to improve fly ash-soil quality in terms of reduced metal load and its impact on soil ecology was evaluated by using the two most widely used indexes i.e. contamination factor (CF) and ecological risk factor (ERF) for four different HMs (Fig. 7).
Values for CF for different metals under the influence of different fly ash-soil-ameliorant combinations significantly varied among themselves. All the treatment combinations exhibited a ‘moderate contamination’ level for Pb (CF: 1-3) excepting the A3T0 combination which showed a ‘considerable contamination’ (CF: 3-6). Pollution load for Cd under the influence of A1T0 and A2T0 showed a ‘considerable contamination’ level whereas the others depicted a ‘moderate contamination’ considering all A1 and A2 combinations. In addition, a ‘considerable contamination’ level for Cd was attained for all the treatments constituted by 100% fly ash in conjunction with different inorganic and organic amendments excepting sole application of fly ash (A3T0) which recorded the ‘high contamination’ level (CF: >6). For Ni, only A1T1 had a ‘low contamination’ level and the others presented mostly of moderate level. A3T0, A3T2, A3T3 had a value between 3 to 6 and categorized ‘considerable contamination’ level. All the treatments showed a ‘low contamination’ level for Cr apart from A3T0 which was categorized under ‘moderate contamination’ level.
Results for ERF showed that all the treatment combinations had a value less than 40 and classified under ‘low ecological risk’ for Pb, Ni, and Cr. But, the values of ERF for Cd varied considerably. A3T0 showed a ‘high ecological risk’ (ERF: >160) whereas A1T0, A2T0, and all other treatments comprising A3-ameliorant combinations were classified under ‘considerable ecological risk’ (ERF: 80-160) for Cd. A1T1 and A1T3 recorded a value less than 40 and considered as ‘low ecological risk’ and the remaining treatments presented moderate level-values for the same.
Assessment of microbial endpoints under the influence of fly ash-soil-ameliorant combinations
To support the ecological risk factor in the earlier section, two widely used soil health indicators viz. microbial biomass carbon (MBC) and dehydrogenase activity under the influence of different treatment combinations were assessed (Table 5). Among the base material, MBC load and dehydrogenase activity was highest in A1 followed by A2 and A3 (A1>1.8A2>8.9A3 and A1>1.9A2>19.7A3 respectively), irrespective of the ameliorants applied. While different ameliorants were imposed on the base materials, a noticeable increase in MBC was recorded in A1 followed by A2 over sole fly ash as a base material (A3) by 52 and 26 ug g-1. However, in all cases, irrespective of the base material, T4 excelled over others. On the whole, A1T4 holds significantly higher MBC followed by A1T1 by ~493% and 391%, respectively. A similar trend was also followed in the case of dehydrogenase activity under different treatment combinations and A1T4 followed by A1T1 recorded the highest values (~9 and 7 µg TPF g-1 h-1 respectively) over others.