A discussion and evaluation of statistical procedures used by JIMB authors when comparing means.

Out of the 166 articles published in Journal of Industrial Microbiology and Biotechnology (JIMB) in 2019-2020 (not including special issues or review articles), 51 of them used a statistical test to compare two or more means. The most popular test was the (Standard) t-test, which often was used to compare several pairs of means. Other statistical procedures used included Fisher's least significant difference (LSD), Tukey's honest significant difference (HSD), and Welch's t-test; and to a lesser extent Bonferroni, Duncan's Multiple Range, Student-Newman-Keuls, and Kruskal-Wallis tests. This manuscript examines the performance of some of these tests with simulated experimental data, typical of those reported by JIMB authors. The results show that many of the most common procedures used by JIMB authors result in statistical conclusions that are prone to have large false positive (Type I) errors. These error-prone procedures included the multiple t-test, multiple Welch's t-test, and Fisher's LSD. These multiple comparisons procedures were compared with alternatives (Fisher-Hayter, Tukey's HSD, Bonferroni, and Dunnett's t-test) that were able to better control Type I errors. NON-TECHNICAL SUMMARY: The aim of this work was to review and recommend statistical procedures for Journal of Industrial Microbiology and Biotechnology authors who often compare the effect of several treatments on microorganisms and their functions.

The Effects of Oil Content on the Structural and Textural Properties of Cottonseed Butter/Spread Products.

Plant-based butters from nuts and seeds have steadily increased in consumer popularity due to their unique flavors and healthy nutritional properties. Oil content is a critical parameter to measure the proper consistency and stability of plant butter and spread products. Previous work has shown that glandless cottonseed can be used to formulate cottonseed butter products to increase the values of cottonseed. As part of the efforts made in the valorization of cottonseed, this work evaluated the effects of oil content on the microstructural and textural properties of cottonseed butter/spread products. While the oil content in the raw cottonseed kernels was 35% of the kernel biomass, additional cottonseed oil was added to make cottonseed butter products with six oil content levels (i.e., 36, 43, 47, 50, 53, and 57%). The values of three textural parameters, firmness, spreadability, and adhesiveness, decreased rapidly in an exponential mode with the increasing oil content. The particle size population in these butter samples was characterized by similar trimodal distribution, with the majority in the middle mode region with particle sizes around 4.5-10 µm. Higher oil content decreased the butter particle size slightly but increased oil separation during storage. The oxidation stability with a rapid oxygen measurement was gradually reduced from 250 min with 36% oil to 65 min with 57% oil. The results of this work provide information for the further optimization of formulation parameters of cottonseed butter products.

Microbiome Analysis of Sugarcane Juices and Biofilms from Louisiana Raw Sugar Factories.

During postharvest processing of sugarcane for raw sugar, microbial activity results in sucrose loss and undesirable exopolysaccharide (EPS) production. Historically, culture-based approaches have focused on the bacterium Leuconostoc mesenteroides as the main contributor to both processes. However, recent studies have shown that diverse microbes are present in sugarcane factories and may also contribute to sugarcane juice deterioration. In the present study, high-throughput amplicon-based sequence profiling was applied to gain a more comprehensive view of the microbial community in Louisiana raw sugar factories. Microbial profiling of the bacterial and fungal microbiomes by 16S V4 and ITS1 sequences, respectively, identified 417 bacterial amplicon sequence variants (ASVs) and 793 fungal ASVs. While Leuconostoc was indeed the most abundant bacterial genus overall (40.9% of 16S sequences), multiple samples were dominated by other taxa such as Weissella and Lactobacillus, underscoring the microbial diversity present in sugarcane factories. Furthermore, flask cultures inoculated with the same samples demonstrated differences in the rate of sucrose consumption, as well as the production of exopolysaccharides and other organic acids, which may result from the observed differences in microbial composition. IMPORTANCE Amplicon-based sequencing was utilized to address long-ignored gaps in microbiological knowledge about the diversity of microbes present in processing streams at Louisiana sugarcane raw sugar factories. These results support an emerging model where diverse organisms contribute to sugarcane juice degradation, help to contextualize microbial contamination problems faced by raw sugar factories, and will guide future studies on biocontrol measures to mitigate sucrose losses and operational challenges due to exopolysaccharide production.

Recovery of Aconitic Acid from Sweet Sorghum Plant Extract Using a Solvent Mixture, and Its Potential Use as a Nematicide.

Trans-aconitic acid (TAA) is naturally present in sweet sorghum juice and syrup, and it has been promoted as a potential biocontrol agent for nematodes. Therefore, we developed a process for the extraction of aconitic acid from sweet sorghum syrup. The process economics were evaluated, and the extract was tested for its capability to suppress the motility of the nematodes Caenorhabditis elegans and Meloidogyne incognita. Aconitic acid could be efficiently extracted from sweet sorghum syrup using acetone:butanol:ethanol mixtures, and it could be recovered from this solvent with a sodium carbonate solution, with an overall extraction and recovery efficiency of 86%. The estimated production cost was USD 16.64/kg of extract and this was highly dependent on the solvent cost, as the solvent was not recycled but was resold for recovery at a fraction of the cost. The extract was effective in reducing the motility of the parasitic M. incognita and causing over 78% mortality of the nematode when 2 mg/mL of TAA extract was added. However, this positive result could not conclusively be linked solely to TAA. An unidentified component (or components) in the acetone:butanol:ethanol sweet sorghum extract appears to be an effective nematode inhibitor, and it may merit further investigation. The impact of aconitic acid on C. elegans appeared to be entirely controlled by pH.

Oxidative Stability of Cottonseed Butter Products under Accelerated Storage Conditions.

Cottonseed is a natural product of cotton (Gossypium spp.) crops. This work evaluated the oxidative stability of cottonseed butters through accelerated autoxidation by storage at 60 °C for 25 days. Three oxidative stability parameter values (peroxide value, p-anisidine value, and total oxidation value) were monitored over the storage time. These chemical measurements revealed that the storage stability of the butter products was dominated by primary oxidation of lipid (oil) components, while the secondary oxidation levels were relatively unchanged over the storage time. An analysis of the tocopherols (natural oxidants in cottonseed) suggested not only the protection function of the molecules against oxidation of the cottonseed butter during storage, but also the dynamic mechanism against the primary oxidation of lipid components. Attenuated total reflectance-Fourier-transform infrared spectroscopy (ATR-FTIR) data confirmed no changes in the major C functional groups of cottonseed butters over the storage time. On the other hand, characteristic minor peaks of conjugated dienes and trienes related to lipid oxidation were impacted by the accelerated storage. As each day of accelerated oxidation at 60 °C is equivalent to 16 days of storage at 20 °C, observations in this work should have reflected the oxidative stability behaviors of the cottonseed butters after about 13 months of shelf storage under ambient storage conditions. Thus, these data that were collected under the accelerated oxidation testing would be useful not only to create a better understanding of the autooxidation mechanism of lipid molecules in cottonseed butters, but also in developing or recommending appropriate storage conditions for cottonseed end products to prevent them from quality degradation.

Molecular level characterization of the effect of roasting on the extractable components of glandless cottonseed by Fourier transform ion cyclotron resonance mass spectrometry.

Roasting is a technological process in some food applications of agricultural products. To investigate the composition changes of the extractable functional/bioactive components of cottonseed, in this work, glandless cottonseed kernels were roasted at 110, 120, 140 and 150 °C for 15 min, respectively. The UV/vis data of the 80 % ethanol extracts found that roasting increased the level of phenolic compounds. Electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry of the extracts identified about 44 % to 55 % of total formulas as potential phenolic compounds. Roasting (up to 140 °C) mainly increased carbohydrate-, lignin-, and tannin-like compounds while lipid-like compounds decreased. The compositional changes at 150 °C were less than those at 140 °C, attributed to devolatilization at the higher temperature. The information of chemical profiling of cottonseed and the roasting impact would be greatly useful in enhanced utilization of cottonseed as nutrient and functional foods or food supplements.

Aconitic Acid Recovery from Renewable Feedstock and Review of Chemical and Biological Applications.

Aconitic acid (propene-1,2,3-tricarboxylic acid) is the most prevalent 6-carbon organic acid that accumulates in sugarcane and sweet sorghum. As a top value-added chemical, aconitic acid may function as a chemical precursor or intermediate for high-value downstream industrial and biological applications. These downstream applications include use as a bio-based plasticizer, cross-linker, and the formation of valuable and multi-functional polyesters that have also been used in tissue engineering. Aconitic acid also plays various biological roles within cells as an intermediate in the tricarboxylic acid cycle and in conferring unique survival advantages to some plants as an antifeedant, antifungal, and means of storing fixed pools of carbon. Aconitic acid has also been reported as a fermentation inhibitor, anti-inflammatory, and a potential nematicide. Since aconitic acid can be sustainably sourced from renewable, inexpensive sources such as sugarcane, molasses, and sweet sorghum syrup, there is enormous potential to provide multiple streams of additional income to the sugar industry through downstream industrial and biological applications that we discuss in this review.

Butanol production from sweet sorghum bagasse with high solids content: Part I-comparison of liquid hot water pretreatment with dilute sulfuric acid.

In these studies, we pretreated sweet sorghum bagasse (SSB) using liquid hot water (LHW) or dilute H2 SO4 (2 g L-1 ) at 190°C for zero min (as soon as temperature reached 190°C, cooling was started) to reduce generation of sugar degradation fermentation inhibiting products such as furfural and hydroxymethyl furfural (HMF). The solids loading were 250-300 g L-1 . This was followed by enzymatic hydrolysis. After hydrolysis, 89.0 g L-1 sugars, 7.60 g L-1 acetic acid, 0.33 g L-1 furfural, and 0.07 g L-1 HMF were released. This pretreatment and hydrolysis resulted in the release of 57.9% sugars. This was followed by second hydrolysis of the fibrous biomass which resulted in the release of 43.64 g L-1 additional sugars, 2.40 g L-1 acetic acid, zero g L-1 furfural, and zero g L-1 HMF. In both the hydrolyzates, 86.3% sugars present in SSB were released. Fermentation of the hydrolyzate I resulted in poor acetone-butanol-ethanol (ABE) fermentation. However, fermentation of the hydrolyzate II was successful and produced 13.43 g L-1 ABE of which butanol was the main product. Use of 2 g L-1 H2 SO4 as a pretreatment medium followed by enzymatic hydrolysis resulted in the release of 100.6-93.8% (w/w) sugars from 250 to 300 g L-1 SSB, respectively. LHW or dilute H2 SO4 were used to economize production of cellulosic sugars from SSB. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:960-966, 2018.

High solid fed-batch butanol fermentation with simultaneous product recovery: Part II-process integration.

In these studies, liquid hot water (LHW) pretreated and enzymatically hydrolyzed Sweet Sorghum Bagasse (SSB) hydrolyzates were fermented in a fed-batch reactor. As reported in the preceding paper, the culture was not able to ferment the hydrolyzate I in a batch process due to presence of high level of toxic chemicals, in particular acetic acid released from SSB during the hydrolytic process. To be able to ferment the hydrolyzate I obtained from 250 g L-1 SSB hydrolysis, a fed-batch reactor with in situ butanol recovery was devised. The process was started with the hydrolyzate II and when good cell growth and vigorous fermentation were observed, the hydrolyzate I was slowly fed to the reactor. In this manner the culture was able to ferment all the sugars present in both the hydrolyzates to acetone butanol ethanol (ABE). In a control batch reactor in which ABE was produced from glucose, ABE productivity and yield of 0.42 g L-1 h-1 and 0.36 were obtained, respectively. In the fed-batch reactor fed with SSB hydrolyzates, these productivity and yield values were 0.44 g L-1 h-1 and 0.45, respectively. ABE yield in the integrated system was high due to utilization of acetic acid to convert to ABE. In summary we were able to utilize both the hydrolyzates obtained from LHW pretreated and enzymatically hydrolyzed SSB (250 g L-1 ) and convert them to ABE. Complete fermentation was possible due to simultaneous recovery of ABE by vacuum. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:967-972, 2018.

Effects of Particle Size on the Morphology and Water- and Thermo-Resistance of Washed Cottonseed Meal-Based Wood Adhesives.

Water washing of cottonseed meal is more cost-efficient and environmentally friendly than protein isolation by means of alkaline extraction and acidic precipitation. Thus, water-washed cottonseed meal (WCSM) is more promising as biobased wood adhesives. In this work, we examined the effects of the particle size on the morphology and adhesive performance of WCSM. Pilot-scale produced and dried WCSM was treated by three grinding methods: (1) ground by a hammer mill and passed through a 0.5-mm screen, (2) further ground by a cyclone mill and passed through a 0.5-mm screen, or (3) further ground by a ball mill and passed through a 0.18-mm screen. Micro-morphological examination revealed two types of particles. The filament-like particles were mainly fibrous materials from residual linters. Chunk-like particles were more like aggregates or accumulations of small particles, with proteins as the major component. Further grinding of the 0.5-mm Hammer product with the Cyclone and Ball mill led to more fine (smaller) particles in the WCSM products. The impact of further grinding on the dry and soaked adhesive strengths was minimal. However, the decrease of the hot and wet strengths of WCSM products by the additional grinding was significant (p ≤ 0.05). Data presented in this work is useful in developing the industrial standards of WCSM products used in wood bonding.

Dedicated Industrial Oilseed Crops as Metabolic Engineering Platforms for Sustainable Industrial Feedstock Production.

Feedstocks for industrial applications ranging from polymers to lubricants are largely derived from petroleum, a non-renewable resource. Vegetable oils with fatty acid structures and storage forms tailored for specific industrial uses offer renewable and potentially sustainable sources of petrochemical-type functionalities. A wide array of industrial vegetable oils can be generated through biotechnology, but will likely require non-commodity oilseed platforms dedicated to specialty oil production for commercial acceptance. Here we show the feasibility of three Brassicaceae oilseeds crambe, camelina, and carinata, none of which are widely cultivated for food use, as hosts for complex metabolic engineering of wax esters for lubricant applications. Lines producing wax esters >20% of total seed oil were generated for each crop and further improved for high temperature oxidative stability by down-regulation of fatty acid polyunsaturation. Field cultivation of optimized wax ester-producing crambe demonstrated commercial utility of these engineered crops and a path for sustainable production of other industrial oils in dedicated specialty oilseeds.

Developmental regulation of diacylglycerol acyltransferase family gene expression in tung tree tissues.

Diacylglycerol acyltransferases (DGAT) catalyze the final and rate-limiting step of triacylglycerol (TAG) biosynthesis in eukaryotic organisms. DGAT genes have been identified in numerous organisms. Multiple isoforms of DGAT are present in eukaryotes. We previously cloned DGAT1 and DGAT2 genes of tung tree (Vernicia fordii), whose novel seed TAGs are useful in a wide range of industrial applications. The objective of this study was to understand the developmental regulation of DGAT family gene expression in tung tree. To this end, we first cloned a tung tree gene encoding DGAT3, a putatively soluble form of DGAT that possesses 11 completely conserved amino acid residues shared among 27 DGAT3s from 19 plant species. Unlike DGAT1 and DGAT2 subfamilies, DGAT3 is absent from animals. We then used TaqMan and SYBR Green quantitative real-time PCR, along with northern and western blotting, to study the expression patterns of the three DGAT genes in tung tree tissues. Expression results demonstrate that 1) all three isoforms of DGAT genes are expressed in developing seeds, leaves and flowers; 2) DGAT2 is the major DGAT mRNA in tung seeds, whose expression profile is well-coordinated with the oil profile in developing tung seeds; and 3) DGAT3 is the major form of DGAT mRNA in tung leaves, flowers and immature seeds prior to active tung oil biosynthesis. These results suggest that DGAT2 is probably the major TAG biosynthetic isoform in tung seeds and that DGAT3 gene likely plays a significant role in TAG metabolism in other tissues. Therefore, DGAT2 should be a primary target for tung oil engineering in transgenic organisms.

Lead retention by broiler litter biochars in small arms range soil: impact of pyrolysis temperature.

Phosphorus-rich manure biochar has a potential for stabilizing Pb and other heavy metal contaminants, as well as serving as a sterile fertilizer. In this study, broiler litter biochars produced at 350 and 650 °C were employed to understand how biochar's elemental composition (P, K, Ca, Mg, Na, Cu, Pb, Sb, and Zn) affects the extent of heavy metal stabilization. Soil incubation experiments were conducted using a sandy, slightly acidic (pH 6.11) Pb-contaminated (19906 mg kg(-1) total Pb primarily as PbCO(3)) small arms range (SAR) soil fraction (<250 µm) amended with 2-20 wt % biochar. The Pb stabilization in pH 4.9 acetate buffer reached maximum at lower (2-10 wt %) biochar amendment rate, and 350 °C biochar containing more soluble P was better able to stabilize Pb than the 650 °C biochar. The 350 °C biochar consistently released greater amounts of P, K, Mg, Na, and Ca than 650 °C biochar in both unbuffered (pH 4.5 sulfuric acid) and buffered (pH 4.9 acetate) systems, despite 1.9-4.5-fold greater total content of the 650 °C biochar. Biochars, however, did not influence the total extractable Pb over three consecutive equilibration periods consisting of (1) 1 week in pH 4.5 sulfuric acid (simulated leaching by rainfall), (2) 1 week in pH 4.9 acetate buffer (standard solution for toxicity characteristic leaching procedure), and (3) 1 h in pH 1.5 glycine at 37 °C (in vitro bioaccessibility procedure). Overall, lower pyrolysis temperature was favorable for stabilizing Pb (major risk driver of SAR soils) and releasing P, K, Ca, and other plant nutrients in a sandy acidic soil.

Expression and purification of recombinant tung tree diacylglycerol acyltransferase 2.

Diacylglycerol acyltransferases (DGATs) esterify sn-1,2-diacylglycerol with a long-chain fatty acyl-CoA, the last and rate-limiting step of triacylglycerol (TAG) biosynthesis in eukaryotic organisms. At least 74 DGAT2 sequences from 61 organisms have been identified, but the expression of any DGAT2 as a partial or full-length protein in Escherichia coli had not been reported. The main objective of this study was to express and purify recombinant DGAT2 (rDGAT2) from E. coli for antigen production with a minor objective to compare rDGAT2 expression in yeast. A plasmid was engineered to express tung tree DGAT2 fused to maltose binding protein and poly-histidine (His) affinity tags. Immunoblotting showed that rDGAT2 was detected in the soluble, insoluble, and membrane fractions. The rDGAT2 in the soluble fraction was partially purified by amylose resin, nickel-nitrilotriacetic agarose (Ni-NTA) beads, and tandem affinity chromatography. Multiple proteins co-purified with rDGAT2. Size exclusion chromatography estimated the size of the rDGAT2-enriched fraction to be approximately eight times the monomer size. Affinity-purified rDGAT2 fractions had a yellow tint and contained fatty acids. The rDGAT2 in the insoluble fraction was partially solubilized by seven detergents with SDS being the most effective. Recombinant DGAT2 was purified to near homogeneity by SDS solubilization and Ni-NTA affinity chromatography. Mass spectrometry identified rDGAT2 as a component in the bands corresponding to the monomer and dimer forms as observed by SDS-PAGE. Protein bands with monomer and dimer sizes were also observed in the microsomal membranes of Saccharomyces cerevisiae expressing hemagglutinin-tagged DGAT2. Nonradioactive assay showed TAG synthesis activity of DGAT2 from yeast but not E. coli. The results suggest that rDGAT2 is present as monomer and dimer forms on SDS-PAGE, associated with other proteins, lipids, and membranes, and that post-translational modification of rDGAT2 may be required for its enzymatic activity and/or the E. coli protein is misfolded.

Expression of a lipid-inducible, self-regulating form of Yarrowia lipolytica lipase LIP2 in Saccharomyces cerevisiae.

Saccharomyces cerevisiae is frequently used as a bioreactor for conversion of exogenously acquired metabolites into value-added products, but has not been utilized for bioconversion of low-cost lipids such as triacylglycerols (TAGs) because the cells are typically unable to acquire these lipid substrates from the growth media. To help circumvent this limitation, the Yarrowia lipolytica lipase 2 (LIP2) gene was cloned into S. cerevisiae expression vectors and used to generate S. cerevisiae strains that secrete active Lip2 lipase (Lip2p) enzyme into the growth media. Specifically, LIP2 expression was driven by the S. cerevisiae PEX11 promoter, which maintains basal transgene expression levels in the presence of sugars in the culture medium but is rapidly upregulated by fatty acids. Northern blotting, lipase enzyme activity assays, and gas chromatographic measurements of cellular fatty acid composition after lipid feeding all confirmed that cells transformed with the PEX11 promoter-LIP2 construct were responsive to lipids in the media, i.e., cells expressing LIP2 responded rapidly to either free fatty acids or TAGs and accumulated high levels of the corresponding fatty acids in intracellular lipids. These data provided evidence of the creation of a self-regulating positive control feedback loop that allows the cells to upregulate Lip2p production only when lipids are present in the media. Regulated, autonomous production of extracellular lipase activity is a necessary step towards the generation of yeast strains that can serve as biocatalysts for conversion of low-value lipids to value-added TAGs and other novel lipid products.

Expression of tung tree diacylglycerol acyltransferase 1 in E. coli.

BACKGROUND: Diacylglycerol acyltransferases (DGATs) catalyze the final and rate-limiting step of triacylglycerol (TAG) biosynthesis in eukaryotic organisms. Database search has identified at least 59 DGAT1 sequences from 48 organisms, but the expression of any DGAT1 as a full-length protein in E. coli had not been reported because DGAT1s are integral membrane proteins and difficult to express and purify. The objective of this study was to establish a procedure for expressing full-length DGAT1 in E. coli. RESULTS: An expression plasmid containing the open reading frame for tung tree (Vernicia fordii) DGAT1 fused to maltose binding protein and poly-histidine affinity tags was constructed and expressed in E. coli BL21(DE3). Immunoblotting showed that the recombinant DGAT1 (rDGAT1) was expressed, but mostly targeted to the membranes and insoluble fractions. Extensive degradation also occurred. Nonetheless, the fusion protein was partially purified from the soluble fraction by Ni-NTA and amylose resin affinity chromatography. Multiple proteins co-purified with DGAT1 fusion protein. These fractions appeared yellow in color and contained fatty acids. The rDGAT1 was solubilized from the insoluble fraction by seven detergents and urea, with SDS and Triton X-100 being the most effective detergents. The solubilized rDGAT1 was partially purified by Ni-NTA affinity chromatography. PreScission protease digestion confirmed the identity of rDGAT1 and showed extensive precipitation following Ni-NTA affinity purification. CONCLUSIONS: This study reports the first procedure for expressing full-length DGAT1 from any species using a bacterial expression system. The results suggest that recombinant DGAT1 is degraded extensively from the carboxyl terminus and associated with other proteins, lipids, and membranes.

Screening biochars for heavy metal retention in soil: role of oxygen functional groups.

Oxygen-containing carboxyl, hydroxyl, and phenolic surface functional groups of soil organic and mineral components play central roles in binding metal ions, and biochar amendment can provide means of increasing these surface ligands in soil. In this study, positive matrix factorization (PMF) was first employed to fingerprint the principal components responsible for the stabilization of heavy metals (Cu, Ni, Cd, Pb) and the release of selected elements (Na, Ca, K, Mg, S, Al, P, Zn) and the pH change in biochar amended soils. The PMF analysis indicated that effective heavy metal stabilization occurred concurrently with the release of Na, Ca, S, K, and Mg originating from soil and biochar, resulting in as much as an order or magnitude greater equilibrium concentrations relative to the soil-only control. In weathered acidic soil, the heavy metal (especially Pb and Cu) stabilization ability of biochar directly correlated with the amount of oxygen functional groups revealed by the O/C ratio, pH(pzc), total acidity, and by the (1)H NMR analysis. Equilibrium speciation calculation showed minor influence of hydrolysis on the total soluble metal concentration, further suggesting the importance of binding by surface ligands of biochar that is likely to be promoted by biochar-induced pH increase.

Influence of pyrolysis temperature on biochar property and function as a heavy metal sorbent in soil.

While a large-scale soil amendment of biochars continues to receive interest for enhancing crop yields and to remediate contaminated sites, systematic study is lacking in how biochar properties translate into purported functions such as heavy metal sequestration. In this study, cottonseed hulls were pyrolyzed at five temperatures (200, 350, 500, 650, and 800 °C) and characterized for the yield, moisture, ash, volatile matter, and fixed carbon contents, elemental composition (CHNSO), BET surface area, pH, pHpzc, and by ATR-FTIR. The characterization results were compared with the literature values for additional source materials: grass, wood, pine needle, and broiler litter-derived biochars with and without post-treatments. At respective pyrolysis temperatures, cottonseed hull chars had ash content in between grass and wood chars, and significantly lower BET surface area in comparison to other plant source materials considered. The N:C ratio reached a maximum between 300 and 400 °C for all biomass sources considered, while the following trend in N:C ratio was maintained at each pyrolysis temperature: wood≪cottonseed hull≈grass≈pine needle≪broiler litter. To examine how biochar properties translate into its function as a heavy metal (NiII, CuII, PbII, and CdII) sorbent, a soil amendment study was conducted for acidic sandy loam Norfolk soil previously shown to have low heavy metal retention capacity. The results suggest that the properties attributable to the surface functional groups of biochars (volatile matter and oxygen contents and pHpzc) control the heavy metal sequestration ability in Norfolk soil, and biochar selection for soil amendment must be made case-by-case based on the biochar characteristics, soil property, and the target function.

Influence of soil properties on heavy metal sequestration by biochar amendment: 2. Copper desorption isotherms.

Contaminant desorption constrains the long-term effectiveness of remediation technologies, and is strongly influenced by dynamic non-equilibrium states of environmental and biological media. Information is currently lacking in the influence of biochar and activated carbon amendments on desorption of heavy metal contaminants from soil components. In this study, copper sorption-desorption isotherms were obtained for clay-rich, alkaline San Joaquin soil with significant heavy metal sorption capacity, and eroded, acidic Norfolk sandy loam soil having low capacity to retain copper. Acidic pecan shell-derived activated carbon and basic broiler litter biochar were employed in desorption experiments designed to address both leaching by rainfall and toxicity characteristics. For desorption in synthetic rain water, broiler litter biochar amendment diminished sorption-desorption hysteresis. In acetate buffer (pH 4.9), significant copper leaching was observed, unless acidic activated carbon (pH(pzc)=3.07) was present. Trends observed in soluble phosphorus and zinc concentrations for sorption and desorption equilibria suggested acid dissolution of particulate phases that can result in a concurrent release of copper and other sorbed elements. In contrast, sulfur and potassium became depleted as a result of supernatant replacements only when amended carbon (broiler litter biochar) or soil (San Joaquin) contained appreciable amounts. A positive correlation was observed between the equilibrium aluminum concentration and initial copper concentration in soils amended with acidic activated carbon but not basic biochar, suggesting the importance of cation exchange mechanism, while dissolution of aluminum oxides cannot be ruled out.

Influence of soil properties on heavy metal sequestration by biochar amendment: 1. Copper sorption isotherms and the release of cations.

The amendment of carbonaceous materials such as biochars and activated carbons is a promising in situ remediation strategy for both organic and inorganic contaminants in soils and sediments. Mechanistic understandings in sorption of heavy metals on amended soil are necessary for appropriate selection and application of carbonaceous materials for heavy metal sequestration in specific soil types. In this study, copper sorption isotherms were obtained for soils having distinct characteristics: clay-rich, alkaline San Joaquin soil with significant heavy metal sorption capacity, and eroded, acidic Norfolk sandy loam soil having low capacity to retain copper. The amendment of acidic pecan shell-derived activated carbon and basic broiler litter biochar lead to a greater enhancement of copper sorption in Norfolk soil than in San Joaquin soil. In Norfolk soil, the amendment of acidic activated carbon enhanced copper sorption primarily via cation exchange mechanism, i.e., release of proton, calcium, and aluminum, while acid dissolution of aluminum cannot be ruled out. For San Joaquin soil, enhanced copper retention by biochar amendment likely resulted from the following additional mechanisms: electrostatic interactions between copper and negatively charged soil and biochar surfaces, sorption on mineral (ash) components, complexation of copper by surface functional groups and delocalized π electrons of carbonaceous materials, and precipitation. Influence of biochar on the release of additional elements (e.g., Al, Ca) must be carefully considered when used as a soil amendment to sequester heavy metals.