The role of low molecular weight organic acids in the release of phosphorus from sewage sludge-based biochar

Abstract

Low molecular weight organic acids (LMWOAs) are common and abundant compounds in the environment, responsible for the release of the nutrients from rhizosphere into the soil solution and, therefore, facilitating the availability of these nutrients for plants. Sewage sludge biochar (SSB) is considered to be a potential source of phosphorus (P) in soil. In this context, the efficacy of twelve LMWOAs in releasing SSB-bearing P was tested in a laboratory extraction experiment. Moreover, the potential effects of (i) biochar pyrolysis temperature (varying from 220°C to 620°C) and (ii) other elements in SS, to estimate the P bonds attacked predominantly by the LMWOAs, were also assessed. The results revealed citric acid was the most effective extraction agent enhancing the mobility of the SSB-derived P. P release was strongly affected by biochar pyrolysis temperature (the highest P release was from the 320°C biochar, significant at p < 0.05). Moreover, the results showed that P was released preferably from Fe-P bearing compounds in SS. Citric acid was recorded as less effective at releasing other nutrients, such as Ca, Mg, K, indicating the exceptional performance of citric acid in P release from biochar without substantial effect on the other nutrients.

KEYWORDS:

• biochar
• organic acids
• phosphorus
• leachability

Introduction

Phosphorus belongs among the most important nutrients for plant growth but 30–40% of arable soils are low in phosphorus (P), all over the world (Kirkby and Johnston 2008). Low P availability is one of the primary limiting factors for crop production in soils, which is attributed to its low solubility, chemical fixation, and complex chelation (Li et al. 2016). To provide plants with a balanced nutrient status inorganic (Pi) fertilizers are applied, where the P compounds are mainly derived from non-renewable resources, i.e. phosphate rocks, which, based on current production rates of P fertilizers, will only last for the next 300–400 years (Glaser and Lehr 2019). However, P-rich waste materials can be used as a potential P source for plants. For instance, sewage sludge (SS) contains high amount of P (14%) (Qian et al. 2019). The predominant position of sewage sludge within the P rich wastes documents that 35% of P wasted from consumption sector is wasted through sewage sludge (Van Dijk 2016).

In this context, the potential risk of SS application to the soil from the enhanced contents of risk elements such as various organic contaminants including pharmaceuticals and personal care products. Jones et al. (2014) surveyed 28 wastewater treatment works in Great Britain for over a period of 12 months and contaminants such as trace metals, pharmaceuticals, polycyclic aromatic hydrocarbons (PAHs), ‘emerging’ and regulated organic pollutants were detected in at least some of the sludges sampled. Many investigations documented that the regular long-term application of SS can result in the regular increase of hardly degradable organic pollutants in soils (Singh and Agrawal 2008; Pulkrabová et al. 2019). Major limitation of the SS use in agriculture is a potential release of heavy metals from the SS and their accumulation to toxic levels in the topsoil (Hanč et al. 2007). In the Czech Republic, the required parameters of sewage sludge suitable for landfill application are given by the public notice of the Ministry of the Environment No. 437/2016. According to these regulations, the maximum allowable contents of the risk elements in SS are 30 mg/kg of As, 5 mg/kg of Cd, 200 mg/kg of Cr, 500 mg/kg of Cu, 100 mg/kg of Ni, 200 mg/kg of Pb, 2500 mg/kg of Zn in SS dry matter. In order to suppress the potential harmful effects of SS in the soil, the pyrolytic thermal treatment of this material resulting in biochar has been widely investigated. Biochar is a carbon-rich solid product of the thermochemical conversion of biomass residues in conditions of minimal or zero oxygen supply, and most of the P in SS can be concentrated in biochar (Qian et al. 2019).

Many previous studies have documented the role of organic acids in P solubilization in the soil. Generally, an acidic environment can significantly enhance the solubility of P minerals (Chen et al. 2006; Li et al. 2016; Wang et al. 2018). Soil organic acids (OAs) can be broadly classified into two groups, namely high molecular weight OAs (HMWOAs) and low molecular weight OAs (LMWOAs). The HMWOAs are characterized by their high molecular weight and low solubility in water whereas the LMWOAs (molecular weight ranging from 46 DA to a few 100 DA) are present in the soil solution (Adeleke et al. 2017). Generally, the major sources of organic acids in the soil are plant root exudations and decomposing microbial as well organic matter (Adeleke et al. 2017). It has been demonstrated that some dicotyledonous plant roots, and especially non-mycorrhizal plants such as Lupinus albus (but not L. angustifolia) and Brassica napus, are capable of releasing large amounts of organic acids into the rhizosphere in response to P deficiency, while other dicotyledonous (e.g. Sisymbrium officinale) and graminaceous (wheat, maize) plants do not appear to express the trait (Jones 1998).

Among microorganisms, bacteria, fungi, and lichen species produce significant amounts of soil OAs. Similarly, decomposed organic matter also contributes to high concentrations of OAs that enrich the soil environment (Adeleke et al. 2017). Chen et al. (2006) stated that phosphate-solubilizing bacteria P release LMWOAs to solubilize phosphorus. Jones (1998) also reported that rhizospheric bacteria and plant roots release large amounts of organic acid, especially in nutrient-limited conditions, for example, malate and citrate appear to be the primary components released by roots under P deficiency. In addition, the co-incubation of biochar and phosphate-solubilizing bacteria can promote the degradation of organic contaminants (for instance the herbicide atrazine), where biochar increases the total and bioavailable P content of the soil (Tao et al. 2020). Carboxyl and hydroxyl anions of organic acids chelate cations or reduce the pH to release P (Alori et al. 2017). These organic acid anions can therefore solubilize P from mineral surfaces either by ligand exchange or by ligand-promoted dissolution (Oburger et al. 2011). In the case of biochars, the effect of LMWOAs on the release of P is dominated by the molecular structure of LMWOAs, the dissociation degree of LMWOAs, and the mineral element stoichiometric ratio of biochars (Zhang et al. 2020). However, the informations concerning the role of LMWOAs in the release of P from biochars are still limited.

This study describes the P solubilizing efficiency of twelve different organic acids (acetic, quinic, benzoic, oxalic, mucic, malic, succinic, maleic, phthalic, trans-aconitic, citric, and kojic acid) from sewage sludge biochar (SSB) produced at different pyrolysis temperatures. The ability of LMWOAs to release P from SSB has an impact on the potential enrichment of the soil with microorganisms containing these acids predominantly in their exudates. A laboratory experiment was conducted to: (i) evaluate the availability of SSB-bearing P as related to both pyrolysis conditions and particular organic acids, and, (ii) recommend the most effective organic acids for further choice of the rhizospheric organisms facilitating the bioaccessibility of the biochar-based P. Therefore, the main objective of the study was to identify the most effective organic acids able to release the biochar-based P for the enhancement of the soil P status via a short-term extraction experiment. In this context, the microorganisms dominating in the exudation of the most effective P-releasing acids could be selected for the further experiments.

Materials and methods

Biochar preparation

SS was sampled from a wastewater treatment plant (WWTP) in the Czech Republic with a design capacity of 29,000 population equivalent (PE) where Fe₂(SO₄)₃ is used for P precipitation. The SS used was anaerobically stabilized (mesophilic anaerobic digestion) and dewatered using a decanter centrifuge (dry matter content 24 wt%). After collection, the sample was air-dried in thin layers at 105°C to constant mass. The dried SS was milled and passed through a 1 mm stainless sieve before thermal treatment. The biochar preparation process is described by Mercl et al. (2020). Briefly: the thermal treatment of the dried SS was performed in an inert atmosphere of N₂ (99.99%) using an electric laboratory tube furnace (GHA 12/600, Carbolite Gero Ltd., Hope Valley, United Kingdom). The torefaction temperatures were: 220°C, and 320°C, the pyrolysis temperatures were 420°C, 520°C, and 620°C. In addition, non-pyrolyzed SS, dried at 105°C, was included in the experimental design. The detailed chemical composition and physicochemical characteristics of the biochars were assessed by Mercl et al. (2020). The contents of risk elements, as well as the emerging organic contaminants in the SS, as well as their changes with increasing pyrolysis temperature were summarized by Mercl et al. (2021) in the Supplementary tables. These materials document that the risk element contents in the SS used in this study was far below the limits given by the Public notice No, 437/2016.

Experimental design and analytical procedure

Three monocarboxylic acids (acetic, quinic, benzoic), six dicarboxylic acids (oxalic, mucic, malic, succinic, maleic, phthalic), two tricarboxylic acids (trans-aconitic, citric), and one representing the enols, kojic acid, were used for P extraction. The main characteristics of the individual organic acids with commercial reagent grade are summarized in Table 1. The appropriate amount of each acid was dissolved in deionized water to make 1 mM solution, and pH of the solution was immediately determined using a Sentron SI400 pH meter. The concentration of organic acids took into account the rhizosphere value reported by Jones (1998). Each treatment was prepared in triplicate. The 0.15 g aliquots of biochar samples (220°C, 320°C, 420°C, 520°C, and 620°C biochar and non-pyrolyzed SS), were weighed into a 50 mL Falcon tube and equilibrated with 15 mL of the 1 mM acid solution. The acid solutions without biochar were included as a blank. The flasks were then incubated at 25°C for 4 h with shaking in an orbital shaker at 150 rpm. At the end of the extraction period, centrifugation was performed at 10,000 × g for 5 min followed by filtration using filter paper (Whatman 42). The concentrations of P, K, Ca, Mg, Fe, and Al were determined using an inductively coupled plasma-optical emission spectrometer (ICP-OES, Agilent 720, Agilent Technologies Inc., USA), equipped with a two-channel peristaltic pump, a Sturman-Masters spray chamber, and a V-groove pneumatic nebulizer made of inert material.

Table 1. The main chemical characteristics of organic acids used for extraction of phosphorus from biochar.

Carboxyl groups	Name (Reagent grade)	Molecular weight	Formula	pKa	pH	Chemical structure
Monocarboxylic	Acetic acid (99%)	126.07	CH3CO2H	pKa = 4.75	3.93
	Quinic acid (98%)	192.17	C7H12O6	pKa = 3.46	3.24
	Benzoic acid (≥99.5%)	122.12	C6H5CO2H	pKa = 4.204	3.68
Dicarboxylic	Oxalic acid (99.6%)	126.07	(CO2H)2	pKa1 = 1.2pKa2 = 4.2	2.87
	Mucic acid (97%)	210.14	C6H10O8	pKa = 2.83	3.31
	Malic acid (99%)	134.09	C4H6O5	pKa1 = 3.51pKa2 = 5.03	3.35
	Succinic acid (≥99.5%)	162.05	C4H6O4	pKa1 = 4.21pKa2 = 5.60	3.40
	Maleic acid (99%)	116.08	C4H4O4	pKa1 = 2.00pKa2 = 6.26	3.06
	Phthalic acid (≥99.5%)	166.13	C8H6O4	pKa1 = 2.76pKa2 = 4.92	3.16
Tricarboxylic	Trans-aconitic acid (98%)	174.11	C6H6O6	pKa1 = 2.80pKa2 = 4.46	3.15
	Citric acid (99.9%)	192.13	C6H8O7	pKa1 = 3.13pKa2 = 4.76pKa3 = 6.39	3.19
Enol	Kojic acid (≥98.5%)	142.11	C6H6O4	pKa = 9.40	5.62

Data analysis

Factorial analysis of variance (ANOVA, p < 0.05), and Tukey’s honestly significant difference (HSD) post-hoc test was performed using Statistica 12.0 software (TIBCO Software Inc.) to determine significant differences between the individual variables. A correlation analysis was used for the assessment of relationships between individual variables; Pearson’s correlation was used with p < 0.05 as the criterion for significance. Principal component analysis (PCA) was performed using XLSTAT (a statistical add-in for Microsoft Excel) software.

Results and discussion

Phosphorus release with the LMWOAs

The total element contents of the biochar have been presented elsewhere (Mercl et al. 2020), where the P content of the non-pyrolyzed dry SS was 32.6 ± 0.3 g/kg, and increased with increasing pyrolysis temperature from 36.4 ± 0.40 g/kg (220°C) to 60.8 ± 0.39 g/kg (620°C). Table 2 presents the proportions of extractable P associated with the individual LMWOAs from the biochars pyrolyzed at different temperatures. Among the twelve organic acids, citric acid extracted the highest P proportions (significant at p < 0.05), regardless of the biochar pyrolysis temperature; mucic and oxalic acid also showed quite good extraction potential. This result is well supported by the findings of Ivanova et al. (2006) where citric acid was confirmed as the most efficient organic acid for P extraction from Tunisian phosphorite. Wei et al. (2010) also found the same result in forest soils where citric acid significantly enhanced the solubilization of organic P by 34.7%, as compared with water; whereas no significant differences were observed in the mobilization of organic P by maleic acid, oxalic acid, or water. The efficiency of citric acid to release soil P was also confirmed by Chatterjee et al. (2015). In the wastewater treatment plants, Fe³⁺ is for P precipitation as FeCl₃ and/or Fe₂(SO₄)₃. Therefore, the total Fe contents in the SS reached up to 4.8% (Mercl et al. 2020), and the presence of P as FePO₄ and other iron bearing phosphates is expected. Pyrolysis degrade the organic P present in the SS on inorganic, which is again associated predominantly with Fe (due to its high content), but also with Al and Ca (Huang et al. 2017). As stated by Zhang et al (2020), deprotonated citric acid is polyvalent anions able to exchange with P in biochar with a high ratio of trivalent metals to P, such as Fe³⁺ in this case.

Table 2. Extractable phosphorus content from the biochar matrix by the individual acids (mg/kg); data expressed as mean ± standard deviation (n = 3); averages marked by the same letter did not significantly differ at p < 0.05 within individual rows (lower case letters), and columns (upper case letters).

	Biochar pyrolysis temperature
Acid	Non-pyrolyzed SS	220°C	320°C	420°C	520°C	620°C
Acetic	1268 ± 37^aF	1304 ± 26^aA	812 ± 30^eAB	547 ± 16^dB	372 ± 23^cA	189 ± 3^bB
Quinic	1138 ± 27^aE	1207 ± 29^aA	859 ± 31^eAB	620 ± 60^dB	440 ± 4^cAB	242 ± 11^bAB
Benzoic	1133 ± 7^cE	1187 ± 37^cA	752 ± 126^dA	532 ± 14^bB	380 ± 31^abA	222 ± 5^aAB
Oxalic	1674 ± 19^aD	1740 ± 53^aAB	1642 ± 146^aDE	1093 ± 15^bA	1096 ± 228^bD	675 ± 11^cA
Mucic	1920 ± 3^eG	2225 ± 2^fB	1510 ± 93^dCDE	1370 ± 25^cD	1083 ± 44^bD	683 ± 24^aA
Malic	1537 ± 28^aAB	1735 ± 24^aAB	1712 ± 437^aE	987 ± 56^bA	1216 ± 95^abD	678 ± 44^bA
Succinic	1553 ± 45^cABD	1584 ± 46^cAB	980 ± 170^bAB	805 ± 16^bC	461 ± 24^aAB	394 ± 8^aAB
Maleic	1472 ± 69^aABC	1556 ± 21^aAB	1109 ± 16^eABC	815 ± 16^dC	599 ± 20^cABC	421 ± 12^bAB
Phthalic	1364 ± 28^dCF	1538 ± 18^eAB	946 ± 107^aAB	833 ± 32^aC	589 ± 17^cABC	422 ± 18^bAB
Trans-aconitic	1580 ± 4^cBD	1741 ± 44^cAB	1232 ± 158^bBCD	1029 ± 44^bA	688 ± 45^aBC	566 ± 29^aAB
Citric	2932 ± 99^aH	3053 ± 869^abC	4174 ± 77^bF	3237 ± 114^abE	3090 ± 183^abE	1436 ± 541^cC
Kojic	1445 ± 15^bAC	1404 ± 35^bA	1049 ± 64^aABC	1000 ± 5^aA	788 ± 40^eC	389 ± 8^cAB

Conversely, Khademi et al. (2009) and Ström et al. (2005) found that oxalate was slightly more effective than citrate in mobilizing P from calcareous soils. The superiority of oxalic acid compared to other LMWOAs in releasing P from calcareous soils was also reported by Moradi et al. (2012). The release of P with the LMWOAs differed for inorganic and organic forms of P, where the extractability of organic P followed the order oxalic acid > citric acid > malic acid, whereas the release of inorganic P followed the order citric acid > oxalic acid > malic acid (Wang et al. 2018). These findings indicate the importance of the P speciation in the soil/biochar, as well as the physicochemical parameters of the soil, and the fate of P having contact with LMWOAs. In the case of biochar, the presence of residual organic compounds of P can be expected and, therefore, citric acid should be preferred to achieve effective release of biochar-bearing P. A comparison of the P release with LMWOAs is limited also due to different composition of both materials. Moreover, in soil, citric acid is an excellent substrate for soil microorganisms, resulting in the fast degradation of citric acid compared to for instance oxalic acid (Oburger et al. 2009). Thus, lower effect of citric acid compared to other LMWOAs in soils is expectable. In this context, Wang et al. (2020) reported an increase in microbial P after the addition of citric acid, malic acid, oxalic acid, or their mixture to the soil, where the changes in microbial P were related to the changes in organic C, soil pH, and available P in the soils.

Basak (2019) tested the efficiency of the LMWOAs on P release from rock phosphates, where the proportions of the extracted P with the individual acids followed the order oxalic acid > citric acid > tartaric acid > formic acid > malic acid > succinic acid > acetic acid. Moreover, the proportions of P released increased as the pH of the extracting solution decreased. Similarly, an inverse relationship between the amounts of P released from soils and the pKa values of the organic acids was reported by Harrold and Tabatabai (2006). The importance of the extractant concentrations was also highlighted by Yang et al. (2019a), where when the concentrations were ≤1 mM, oxalic acid lower proportion of P than citric acid, and when the concentrations were ≥1.5 mM, oxalic acid was more effective. Thus, the similar pattern was expected also in this study, if 1 mM concentration of the acids was used. The pH values of the extracting agents varied between 2.87 and 5.62 (Table 1), and the different efficiency of the individual acids could be at least partially related to their different degrees of acidity. Yang et al. (2019b) showed the importance of pH in the P-biochar interactions. However, although the extractable proportion of P tended to decrease with increases in the pH of the extracting agent, Pearson’s correlation coefficients did not indicate any significant (p < 0.05) relationship between the extracting solution’s pH and extractable P levels, and, therefore, the role of the solution pH remains in question. Moreover, as reviewed by Yang et al. (2021), the soil application of biochar will affect the potential P-release an additional interaction with the soil, where the P mobility in the soil will be a result of (i) the mobile P pool originating from the biochar, (ii) the enhancement of the mobility of the soil-bearing P due to the soil-biochar interaction, and (iii) reduction of the P losses in the biochar treated soils.

Other macro-and micronutrient release with the LMWOAs and their role in the P release

Tables 3–6 summarize the extractable contents of K, Ca, Mg, and Fe from the biochar samples. The maximum K content (899 ± 8 mg/kg) was extracted from the non-pyrolyzed SS by succinic acid (Table 3); pyrolysis resulted in a decrease in the extractable K proportion. Of the other LMWOAs, malic and citric acid also showed quite high extractable K in high-temperature biochar (420, 520, and 620°C), whereas for lower temperature biochars, only occasional significant (p < 0.05) differences in the K extractability were recorded among the LMWOAs. Previous research presented by Khademi et al. (2009) found high concentrations of citric acid were more effective than oxalate in mobilizing Ca, Fe, Mn, and Zn. This study found that trans-aconitic acid was the most effective OA for calcium extraction and the highest amount of extractable calcium was obtained from 220°C biochar (Table 4), but Ca was better extracted with citric acid than with oxalic acid. Similarly as in the case of P, the extractable nutrient proportions remained unaffected by the solution pH, where no significant (p < 0.05) correlations were recorded.

Table 3. Extractable potassium content from biochar matrix by the individual acids (mg/kg); data expressed as mean ± standard deviation (n = 3); averages marked by the same letter did not significantly differ at p < 0.05 within individual rows (lower case letters) or columns (upper case letters).

	Biochar pyrolysis temperature
Acid	Non-pyrolyzed SS	220°C	320°C	420°C	520°C	620°C
Acetic	726 ± 23^eA	657 ± 5^dABCD	246 ± 11^bA	360 ± 25^cA	184 ± 21^aA	204 ± 11.9^abAB
Quinic	729 ± 3^bA	683 ± 2^bA	173 ± 28^aA	372 ± 13^cA	222 ± 49^aA	226 ± 17^aABCD
Benzoic	676 ± 6^bA	624 ± 20^bB	157 ± 19^aA	328 ± 26^cA	167 ± 32^aA	204 ± 20^aAB
Oxalic	729 ± 11^bA	678 ± 7^bA	218 ± 10^aA	384 ± 10^cA	227 ± 42^aA	242 ± 8^aACDE
Mucic	764 ± 36^dAB	674 ± 14^cAD	161 ± 27^aA	370 ± 15^bA	209 ± 15^aA	219 ± 4^aABC
Malic	724 ± 133^cA	649 ± 16^cABCD	189 ± 78^aA	402 ± 38^bA	270 ± 5^abA	291 ± 34^abDE
Succinic	899 ± 8^eB	804 ± 16^dE	209 ± 65^aA	419 ± 13^cA	190 ± 21^aA	301 ± 3.8^bE
Maleic	756 ± 19^dAB	669 ± 5^cACD	196 ± 10^aA	393 ± 43^bA	252 ± 33^aA	231 ± 6^aABCD
Phthalic	684 ± 29^bA	667 ± 15^bABCD	173 ± 23^cA	383 ± 21^dA	251 ± 15^aA	257 ± 10^aACDE
Trans-aconitic	708 ± 39^bA	629 ± 23^bBC	203 ± 38^aA	323 ± 129^aA	189 ± 117^aA	221 ± 30^aABC
Citric	684 ± 90^cA	632 ± 11^cBCD	201 ± 6^aA	388 ± 65^bA	253 ± 26^abA	281 ± 57^abCDE
Kojic	670 ± 13^bA	646 ± 22^bABCD	204 ± 29^aA	316 ± 3.8^cA	192 ± 27^aA	171 ± 5.8^aB

Table 4. Extractable calcium contents from biochar matrix by the individual acids (mg/kg); data expressed as mean ± standard deviation (n = 3); averages marked by the same letter did not significantly differ at p < 0.05 within individual rows (lower case letters) or columns (upper case letters).

	Biochar pyrolysis temperature
Acid	Non-pyrolyzed SS	220°C	320°C	420°C	520°C	620°C
Acetic	4657 ± 87^cBF	8012 ± 55^dABC	3119 ± 314^bABC	4237 ± 40^cB	2606 ± 207^aAB	2924 ± 58^abA
Quinic	4511 ± 159^bAB	8161 ± 70^cAC	3208 ± 274^aABC	4079 ± 81^bAB	2695 ± 366^aAB	2810 ± 37^aA
Benzoic	4369 ± 48^bAB	7584 ± 115^cB	2750 ± 645^aAB	3879 ± 21^bA	2298 ± 405^aAB	2606 ± 23^aA
Oxalic	3679 ± 27^dE	6845 ± 40^eE	2754 ± 55^aAB	2857 ± 21^aD	1763 ± 76^cA	1254 ± 12^bD
Mucic	5428 ± 151^cC	7851 ± 258^dBC	3314 ± 290^aABC	4105 ± 202^acAB	2717 ± 1423^abAB	1175 ± 40^bD
Malic	5202 ± 98^bC	8427 ± 95^cA	4491 ± 275^aCD	5222 ± 113^bG	4394 ± 76^aCD	4181 ± 23^aBC
Succinic	6167 ± 123^cD	9373 ± 153^dD	3413 ± 939^abABCD	5723 ± 29^cC	2704 ± 285^aAB	4368 ± 33^bC
Maleic	5117 ± 415^cCF	8538 ± 59^dA	3658 ± 500^aABCD	4863 ± 275^bcF	3677 ± 380^aBC	3973 ± 27^abB
Phthalic	6063 ± 286^cD	9510 ± 58^dD	3914 ± 181^aBCD	5885 ± 58^cC	4601 ± 252^bCD	4328 ± 47^abBC
Trans-aconitic	7171 ± 92^bG	10588 ± 167^cF	4828 ± 986^aDE	7175 ± 85^bI	4567 ± 358^aCD	5405 ± 45^aE
Citric	5965 ± 83^abD	9260 ± 461^cD	6172 ± 63^abE	6736 ± 150^bH	5815 ± 232^aD	5587 ± 425^aE
Kojic	4003 ± 92^cAE	6969 ± 135^dE	2253 ± 440^aA	3282 ± 54^bE	2346 ± 316^aAB	2594 ± 58^aA

Table 5. Extractable magnesium contents from biochar matrix by the individual acids (mg/kg); data expressed as mean ± standard deviation (n = 3); averages marked by the same letter did not significantly differ at p < 0.05 within individual rows (lower case letters) or columns (upper case letters).

	Biochar pyrolysis temperature
Acid	Non-pyrolyzed SS	220°C	320°C	420°C	520°C	620°C
Acetic	993 ± 33^dBCD	1291 ± 25^eAC	363 ± 32^bcABC	340 ± 34^abABC	264 ± 51^aABDE	442 ± 9^cAD
Quinic	919 ± 27^cAB	1259 ± 4.6^dABC	337 ± 43^abABC	336 ± 3^abABC	310 ± 29^aABCE	406 ± 33^bBCD
Benzoic	878 ± 9^dAE	1164 ± 14^eBD	271 ± 50^abA	284 ± 25^bC	201 ± 13^aD	386 ± 16^cBC
Oxalic	790 ± 31^dE	1214 ± 26^eAB	368 ± 5.9^cABC	297 ± 25^aBC	217 ± 11^bAD	286 ± 6^aH
Mucic	1059 ± 8^cD	1361 ± 27^dC	369 ± 77^aABC	406 ± 5^abAD	370 ± 40^aCF	512 ± 33^bEF
Malic	907 ± 23^cAB	1160 ± 22^dBD	406 ± 43^abBC	380 ± 21^aABD	363 ± 41^aCEF	474 ± 6^bAE
Succinic	1022 ± 30^dCD	1218 ± 30^eAB	317 ± 52^abABC	358 ± 39^bcABC	246 ± 34^aABD	436 ± 8^cACD
Maleic	880 ± 56^bAE	1062 ± 47^cD	285 ± 52^aAB	328 ± 48^aABC	248 ± 24^aABD	361 ± 16^aB
Phthalic	974 ± 75^cABCD	1287 ± 41^dAC	315 ± 11^aABC	386 ± 29^abAD	317 ± 54^aABCE	466 ± 7^bAE
Trans-aconitic	1165 ± 16^cF	1362 ± 68^dC	441 ± 60^abCD	443 ± 22^abDE	374 ± 39^aCF	535 ± 25^bFG
Citric	957 ± 18^cABC	1249 ± 18^dAB	567 ± 29^bD	500 ± 13^aE	461 ± 4^aF	567 ± 10^bG
Kojic	949 ± 6^cABC	1274 ± 53^dAC	368 ± 27^aABC	371 ± 28^aABD	337 ± 22^aBCE	532 ± 3.6^bFG

Table 6. Extractable iron contents from biochar matrix by the individual acids (mg/kg); data expressed as mean ± standard deviation (n = 3); averages marked by the same letter did not significantly differ at p < 0.05 within individual rows (lower case letters) or columns (upper case letters).

	Biochar pyrolysis temperature
Acid	Non-pyrolyzed SS	220°C	320°C	420°C	520°C	620°C
Aceic	79 ± 7^bA	54 ± 4^cA	89 ± 9^bA	16.7 ± 0.8^aA	6 ± 1^aA	7 ± 2^aA
Quinic	85 ± 8^cA	58 ± 4.2^bA	365 ± 19^dBCD	16 ± 1.5^aA	13 ± 1.6^aA	7 ± 3.4^aA
Benzoic	68 ± 10^bA	44 ± 3.6^abA	230 ± 41^cABC	14.7 ± 2.5^aA	11 ± 4^aA	8 ± 0.2^aA
Oxalic	135 ± 39^aA	131 ± 30^aA	577 ± 113^cD	10.8 ± 3^abA	14 ± 4.8^abA	2.2 ± 3^bA
Mucic	266 ± 8^cB	659 ± 8^dC	1382 ± 5^eE	486 ± 12^aB	466 ± 100^aC	144 ± 12^bC
Malic	289 ± 3.4^abB	467 ± 159^bB	1883 ± 109^dF	86 ± 42^aA	898 ± 96^cB	334 ± 45^abB
Succinic	78 ± 22^aA	69 ± 5^aA	379 ± 71^bCD	12 ± 3.3^aA	7 ± 2.7^aA	4.6 ± 1.3^aA
Maleic	73 ± 8^bA	61 ± 0.8^bA	152 ± 8^cAB	20 ± 11^aA	9 ± 2.9^aA	8 ± 4.1^aA
Phthalic	79 ± 5.7^bA	49 ± 9^abA	30 ± 21^aA	0.43 ± 0^aA	n.d.	n.d.
Trans-aconitic	83 ± 3.9^aA	83 ± 25^bA	150 ± 26^cAB	3.5 ± 1.9^aA	1.7 ± 1.1^aA	n.d.
Citric	3293 ± 97^cD	4319 ± 97^aE	6045 ± 104^dG	4132 ± 154^aD	3951 ± 380^aD	1719 ± 81^bD
Kojic	917 ± 20^abC	1020 ± 54^cD	1213 ± 62^dE	1136 ± 16^cdC	845 ± 76^aB	330 ± 8^eB

n.d. data below detection limits.

Wang et al. (2016) reported that LMWOAs (oxalic, citric, and malic acids) can enhance P release from soils due to ligand exchange and the chelation of LMWOAs with Fe and Al. The role of Fe/Al (hydr)oxides in the P release from biochar was presented by Peng et al. (2019). The organic acids exhibit chelation and complexing properties, where citric acid belongs to the effective (Kpomblekou and Tabatabai 2003; Alori et al. 2017) and is a well-known chelating agent (Kubota et al. 2010). As mentioned above, in the SS-based biochar, characterized by the high content of Fe-bearing phosphates, the LMWOAs with tri-carboxylic groups have excellent chelating ability with polyvalent metal ions. Similarly, Wu et al. (2015) showed LMWOAs stimulated release of inorganic P from the soil due to their ability to complex or precipitate with cations such as Fe, Al, and Ca. As reviewed by Kratz et al. (2019), in a chemical waste water treatment process, P will be adsorbed to Fe/Al-hydroxides, or precipitate as Fe/Al-phosphate or basic calcium phosphates, and these compounds can be easily dissolved with citric acid. Taghipour and Jalali (2013) found a different pattern for the individual LMWOAs in contact with individual soil fractions, where oxalate mainly released P contained in Ca–P minerals, whereas citrate predominantly released the P contained in Fe–P and Al–P minerals. The interactions of these elements could indicate which acid releases P from particular compounds. In this experiment, Figure 1 indicated that there was a close relationship among the proportions of Al, Fe, and P extractable with citric acid, whereas a weaker relationship was observed for the P – Ca interaction. Moreover, a similar pattern was observed for Ca, Mg, and K, which were extractable with most of the tested LMWOAs (except citric acid) from the non-pyrolyzed SS and from biochar pyrolyzed at 220°C. No relationships were recorded between the LMWOAs and extractable Ca, K, Mg proportions at the higher pyrolysis temperatures. The findings presented in Figure 1 are supported by the significant (p < 0.05) Pearson’s correlations between P and Fe proportions extractable with citric acid (r = 0.97).

Figure 1. Principal component analysis (PCA) plot of biochar pyrolysis temperatures. The first two principal components (PCs) are plotted. PCA was performed using all analyte data. The percentage of variation accounted for by each PC is shown in brackets with the axis label.

Additionally, Bolan et al. (1994) found (decades ago) decreasing P adsorption by soils after the addition of organic acids in the order tricarboxylic acid > dicarboxylic acid > monocarboxylic acid. In our study, the extractable P proportions correlated significantly (p < 0.05) with the number of carboxylic groups of the acids (the r-value varied between 0.62 and 0.68), partially confirming these findings. Whereas enhanced extraction efficiency was proven for citric acid compared to the other extraction agents tested, P extractability with trans-aconitic acid did not differ from the dicarboxylic acids such as succinic, phthalic, and maleic acids (Table 2). Thus, the results showed the exceptional performance of citric acid for potential enhancement of P release from SS-derived biochar, where the release of the other nutrients remained unaffected.

Moreover, the efficiency of the individual LMWOAs can be modified in real conditions by the presence of other acids. For instance, P release from sediments was enhanced by using an oxalate/citrate mixture, compared to the application of individual extracting agents, indicating the potential synergetic effect of these LMWOAs (Gaviglio et al. 2019). Similarly, Mondala et al. (2017) reported that 2:1 and 1:1 (molar ratios) oxalate-citrate mixtures were able to release up to 61% of total bound P from sediments. LMWOAs are used by rhizosphere microorganisms as sources of carbon and energy, and citric acid favored the proliferation of the Clostridiaceae, Micrococcaceae, and Pseudomonadaceae families (Macias-Benitez et al. 2020). In addition, the enhanced release of P from various P-bearing minerals and rocks due to enhanced soil LMWOAs produced by various soil microorganisms, such as Penicillium spp., Talaromyces spp., Aspergillus niger, etc. has been described (Arcand and Schneider 2006; Scervino et al. 2010; Xiao et al. 2013; Solans et al. 2019).

The role of biochar pyrolysis temperature in the nutrient release performances

Among the biochars, a significantly higher amount of P was extracted (4174.6 ± 76.6 mg/kg) from 320°C biochar compared to the non-pyrolyzed SS. In contrast, the highest pyrolysis temperature, 620°C, resulted in a significantly (p < 0.05) lower proportion of P released with citric acid compared to all the biochars pyrolyzed at the lower temperatures and to the non-pyrolyzed SS (Table 2). Compared to citric acid, the other investigated LMWOAs showed increased (p < 0.05), or similar proportions of P vs. the non-pyrolyzed SS, with regular decreases in extractable P with increasing biochar pyrolysis temperatures (Table 2).

The extractable calcium proportion was highest in the case of the 220°C biochar, as also reported by Mercl et al. (2020). The extractability of Mg showed a similar pattern, where the highest value of Mg (1362.7 ± 68.1 mg/kg) was released by trans-aconitic acid (Table 5). A significant (p < 0.05) decrease in extractable Mg was observed for the biochars prepared at temperatures >220°C. Citric acid was the most effective OA for all types of biochars (and the non-pyrolyzed SS) in the case of iron (Table 6). Citric acid showed the highest iron extraction (60451 ± 104 mg/kg) with the 320°C biochar. Kojic acid, malic acid, and mucic acid showed a small amount of extractable iron for the high-temperature biochars (520°C and 620°C). One important finding was that as the pyrolysis temperature increased from 420°C to 620°C iron extraction decreased. Apart from the elements mentioned above, the extractability of Al was tested, where most of the measured data were under the detection limit of the analytical technique used. Detectable Al content was found only in the case of five LMWOAs, and their extractability decreased in the order citric > mucic > kojic > malic > oxalic acids. The extractable contents of Al in the case of non-pyrolyzed SS varied between 268 mg/kg (citric acid) and 7.3 mg/kg (oxalic acid). In regard to the biochars, the citric acid extractable proportions of Al slightly increased compared to the non-pyrolyzed SS, and the pyrolysis temperature did not affect Al extractability. For malic, kojic, and mucic acids the extractable Al proportions from biochars significantly (p < 0.05) increased with increasing pyrolysis temperature, and the ambiguous results obtained for the Al content extractable with oxalic acid can be related to the low levels of this element, which are close to the detection limit.

Conclusions

The results showed a different response for P extractability depending on the application of different LMWOAs, but the effect of the chemical properties and the composition of the individual acids, and the presence of other elements also impacted the response. Moreover, the results showed specific interactions between LMWOAs and SS-based biochar, which could modify the P mobility in biochar treated soils. Among the P solubilizing microorganisms, those producing preferably citric acid should be considered as potential bioaugmentation measures for the effective enhancement of P release from SS-derived biochar, and, subsequently, to improve the availability of P originating from biochar. Therefore, in further experiments, the role of the LMWOAs in the P release should be tested in the soil-biochar system.

Acknowledgements

The authors thank NAZV QK1710379 project and European Regional Development Fund – Project No. CZ.02.1.01/0.0/0.0/16_019/0000845 for financial support. Correction and improvement of language was provided by Proof-Reading-Service.com Ltd., Devonshire Business Centre, Works Road, Letchworth Garden City SG6 1GJ, United Kingdom.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

The data that support the findings of this study are openly available in Mendeley Data: https://doi.org/10.17632/5vf53yy24p.1.

Additional information

Funding

This work was supported by European Regional Development Fund [grant number CZ.02.1.01/0.0/0.0/16_019/0000845]; National agency for agricultural research [grant number QK1710379].