A comprehensive and critical review of the unstandardized Folin-Ciocalteu assay to determine the total content of polyphenols: The conundrum of the experimental factors and method validation.

The Folin-Ciocalteu method can be considered to be the most widely used in laboratories around the world, to quantify the total polyphenols content. Many different variations found in this assay have been reported in the scientific literature. In this review, the full experimental conditions influencing the Folin-Ciocalteu assay have been comparatively assessed and discussed. Furthermore, few studies relating to the method validation have been evaluated according to the results of selectivity, linearity, precision, trueness, limit of determination, limit of quantification and robustness. In general, the results derived from the reviewed literature are widely variable according to both, the experimental factors selected and the performance parameters reported, making difficult the comparison of the overall results published.

Valorization of strawberry extrudate waste: Recovery of phenolic compounds by direct-hydrothermal treatment and subsequent methane production by mesophilic semi-continuous anaerobic digestion.

Strawberry extrudate (SE) is an underused by-product from strawberry industry. Recovery of the phenolic compounds present in SE would represent a very interesting valorisation option. Two main challenges need to be solved, firstly, the solubilisation and recovery of the phenolic compounds contained in SE, and, after that, the stabilisation of the resulted de-phenolized SE. The present research evaluates the potential of a biorefinery process combining a hydrothermal pre-treatment, followed by a phenolic extraction process and, finally, the anaerobic digestion of the remaining SE for producing energy that will contribute to compensate the energy requirements of the whole system. Following the hydrothermal pre-treatment at 170 °C for 60 min, an extraction of 0.6 ± 0.1 g of gallic acid per kilogram of SE was achieved using an adsorbent resin, representing a recovery rate of 64 %. Long-term semi-continuous anaerobic digestion of de-phenolized SE was evaluated at different organic loading rates to evaluate the stability of the process. The anaerobic digestion of pre-treated SE achieved a stable methane production value of 243 ± 34 mL CH4·g volatile solids-1·d-1 at an organic loading rate (ORL) of 1.25 g volatile solids·L-1·d-1. During the operation at this ORL, the control parameters including pH, alkalinity, soluble chemical organic demand (sCOD), and volatile fatty acid (VFA) remained stable and consistently constant. Specifically, the VFA in the reactor during this stable period achieved a value of 102 ± 128 mg O2/L. Also, an economic balance showed that the minimal price of the generated phenolic extract for having benefited from the proposed biorefinery system was 0.812 €·(g of gallic acid equivalents)-1, a price within the range of phenolic compounds used in the food industry.

Impact of natural degradation of the invasive alga Rugulopteryx okamurae on anaerobic digestion: Heavy metal pollution and kinetic performance.

This study shows, for the first time, how the natural biodegradation of the Phaeophyceae Rugulopteryx okamurae (R.o.) affects its methane yield, by biochemical methane potential assays, and the methane production kinetics. Additionally, a mechanical (zeolite-assisted milling) and a thermal (120 °C, 45 min) pretreatments were assessed. The highest methane yield was obtained from the mechanically pretreated fresh ashore biomass (219 (15) NLCH4 kgVS-1), which presents the use of zeolite during milling as an economical alternative for heavy metal toxicity reduction. Moreover, no significant differences were observed between the other tests (with the exception of the lowest value obtained for the mechanically pretreated fresh R.o.). Low methane yields were linked to the heavy metal content. However, an increase of 28.5 % and 20.0 % in the k value was found for the untreated fresh R.o. biomass and fresh ashore biomass, respectively, when subjected to thermal pretreatment. Finally, an enhancement of 80.5 % in the maximum methane production rate was obtained for the fresh ashore biomass milled with zeolite compared to the untreated fresh ashore biomass.

Effect of metals on mesophilic anaerobic digestion of strawberry extrudate in batch mode.

According to recent studies, the anaerobic digestion of strawberry extrudate is a promising option with potential in the berry industry biorefinery. However, the lack and/or unbalance of concentrations of metals in some agro-industrial residues could hamper methane production during the anaerobic digestion of these kinds of wastes. In this study, a fractional factorial design was applied to screen the supplementation requirements regarding six metals (Co, Ni, Fe, Cu, Mn, and Zn) for methane production from strawberry extrudate (SE). The logistic model was used to fit the experimental data of methane production-time. It allowed identifying two different stages in the anaerobic process and obtaining the kinetic parameters for each step. Maximum methane production obtained in the first (Bmax) kinetic stage, the methane production in the second stage (P), and the maximum methane production rates (Rmax) concluded a statistically significant effect for Ni and Zn. The second set of experiments was carried out with Ni and Zn through a central composite design to study the concentration effect in the anaerobic digestion process of the strawberry extrudate. The parameters P and Rmax demonstrated a positive interaction between Ni and Zn. Although, Bmax did not prove a statistically significant effect between Ni and Zn.

Turning an invasive alien species into a valuable biomass: Anaerobic digestion of Rugulopteryx okamurae after thermal and new developed low-cost mechanical pretreatments.

The invasive alien seaweed Rugulopteryx okamurae (R.o.) has spread quickly through the Mediterranean Sea causing an unprecedented ecological impact. A solution integrated into a circular economy model is needed in order to curb the negative effects of its presence. Anaerobic digestion (AD) is proposed as a feasible process able to transform biomass into renewable energy. Nevertheless, in order to improve the methane yield and surpass the drawbacks associated with AD processes, this research proposes a thermal pretreatment and a new developed method where the macroalgae is mechanically pretreated with zeolite. Chemical and microstructure characterization of the algal biomass after pretreatments involved scanning electron microscopy (SEM), X-ray powder diffraction (XRD) and Fourier-transform infrared spectroscopy (FTIR). The highest methane yields of 240 (28) and 250 (20) NLCH4 kg-1 VSadded were obtained with the new mechanical pretreatment and the thermal pretreatment at 120 °C for 45 min without zeolite, achieving a 35 % improvement against the non-pretreated algae. A direct relationship between the crystallinity index of the samples and methane production was observed. The experimental data of methane production versus time were found to be in accordance with both first-order kinetic and Transference Function mathematical models.

Integral Valorization of Two-Phase Olive Mill Solid Waste (OMSW) and Related Washing Waters by Anaerobic Co-digestion of OMSW and the Microalga Raphidocelis subcapitata Cultivated in These Effluents.

This study evaluates the comprehensive valorization of the byproducts derived from the two-phase olive oil elaboration process [i.e., olive washing water (OWW), olive oil washing water (OOWW), and olive mill solid waste (OMSW)] in a closed-loop process. Initially, the microalga Raphidocelis subcapitata was grown using a mixture of OWW and OOWW as the culture medium, allowing phosphate, nitrate, sugars, and soluble chemical oxygen demand removal. In a second step, the microalgal biomass grown in the mixture of washing waters was used as a co-substrate together with OMSW for an anaerobic co-digestion process. The anaerobic co-digestion of the combination of 75% OMSW-25% R. subcapitata enhanced the methane yield by 7.0 and 64.5% compared to the anaerobic digestion of the OMSW and R. subcapitata individually. This schedule of operation allowed for integration of all of the byproducts generated from the two-phase olive oil elaboration process in a full valorization system and the establishment of a circular economy concept for the olive oil industry.

Assessment of different mechanical treatments for improving the anaerobic biodegradability of residual raspberry extrudate.

Mechanical treatments can be simple and feasible methods for enhancing the anaerobic digestion of lignocellulosic substrates. This work aims to relate the direct effect of five different mechanical treatments, i.e., variation in the size and number of particles, with the variations in the chemical composition and, subsequently, the effect over the anaerobic digestion of residual raspberry extrudate, which was used as a model substrate. A high variation in the number of particles and the particle size distribution was achieved depending on the mechanical treatment applied, reaching the highest number of particles for the treatments with knife mills and mortar (around 8000 particles per gram). The higher number of particles was related to higher solubilisation, including phenolic compounds and sugars. The combination of knife mills and mortar pretreatment, which presented the highest number of particles, resulted in a 66% more of polyphenols in comparison to the raw substrate. However, the presence of anthocyanins was higher in mechanical treatments with less effect. The enhancement of the anaerobic digestion was clearly related to the increment in the number of particles of small size after the mechanical treatments. The highest methane yield coefficient (236 ± 11 mL CH4/g volatile solids) was achieved for the raspberry extrudate treated with knife mills.

Evaluation and modelling of methane production from corn stover pretreated with various physicochemical techniques.

Lignocellulosic by-products from agricultural crops represent an important raw material for anaerobic digestion and clean renewable, which is a key component of the circular economy. Lignocellulose is recalcitrant to biodegradation and pretreatments are required to increase methane yield during anaerobic digestion. In this work, the efficacy of different physicochemical pretreatments was compared using corn stover biomass as substrate. Anaerobic digestion of untreated and pretreated corn stover was performed in batch mode at mesophilic temperature (38°C) and organic matter solubilization of pretreated substrates was also investigated. The highest organic matter solubilization occurred in autoclave pretreatment (soluble chemical oxygen demand = 5630 ± 42 mg O2 L-1). However, the highest methane yield was obtained using alkaline pretreatment (367 ± 35 mL CH4 g-1 VSadded). Alkaline pretreatment increased methane yield by 43.3% compared to untreated control (256 ± 15 mL CH4 g-1 VSadded). Two mathematical models (i.e. first-order kinetics and transfer function) were utilized to fit the experimental data with the aim of assessing anaerobic biodegradation and to obtain the kinetic constants in all cases studied. Both models adequately fit the experimental results. The kinetic constant, k, of the first-order model increased by 92.8% when stover was pretreated with sulphuric acid compared with control. The transfer function model revealed that the maximum methane production rate, Rm, was obtained for the sulphuric acid treatment, which was 63.5% higher compared to control.

Sulfur-based mixotrophic denitrification with the stoichiometric S0/N ratio and methanol supplementation: effect of the C/N ratio on the process.

The impact of the organic carbon to nitrate ratio (C/N ratio) on mixotrophic denitrification rate has been scarcely studied. Thus, this work aims to investigate the effect of the C/N ratio on the mixotrophic denitrification when methanol is used as a source of organic matter and elemental sulfur as an electron donor for autotrophic denitrification. For this, two initial concentrations of NO3--N (50 and 25 mg/L) at a stoichiometric ratio of S0/N, and four initial C/N ratios (0, 0.6, 1.2, and 1.9 mg CH3OH/mg NO3- -N) were used at 25 (±2) °C. The results showed that when using a C/N ratio of 0.6, the highest total nitrogen removal was obtained and the accumulation of nitrites was reduced, compared to an autotrophic system. The most significant contribution to nitrate consumption was through autotrophic denitrification (AuDeN) for a C/N ratio of 0.6 and 1.2, while for C/N = 1.9 the most significant contribution of nitrate consumption was through heterotrophic denitrification (HD). Finally, organic supplementation (methanol) served to increase the specific nitrate removal rate at high and low initial concentrations of substrate. Therefore, the best C/N ratio was 0.6 since it allowed for increasing the removal efficiency and the denitrification rate.

Comparative effect of biochar and activated carbon addition on the mesophilic anaerobic digestion of piggery waste in batch mode.

A comparative study of the batch mesophilic anaerobic digestion of piggery waste was carried out with the addition of 5% biochar and 5% activated carbon. The results obtained showed that the bioreactors amended with biochar increased cumulative methane production, the kinetic constant for methane production and the COD removal efficiency compared to the control reactors and reactors with activated carbon addition. The maximum methane production and the kinetic constant were 6.9% higher in the reactors with biochar addition compared to the controls; while the COD removal efficiency was 3% higher in the case of biochar addition. In the case of activated carbon, only a slight improvement in anaerobic digestion performance was observed compared to the control.

Effect of organic loading rate on the anaerobic digestion of swine waste with biochar addition.

The aim of this study was to investigate the impact of biochar addition on the mesophilic semi-continuous anaerobic digestion of swine waste with a focus on the effects of the organic loading rate (OLR) on biogas production, methane yield, total volatile fatty acids (TVFA), alkalinity, ammonium, volatile solids (VS) removal efficiency and process stability. Four reactors, two with amended biochar (R1 and R2) and two without biochar addition as controls (R3 and R4), were operated at OLRs in the range of 2-7 g VS/(L d), which corresponded to hydraulic retention times (HRTs) in the range of 7-2 days, respectively. The addition of biochar initially caused an increase in the generation of biogas and methane when compared to the control reactors when the process operated at OLRs of 2 and 3 g VS/(L d). This behaviour could be attributed to the presence of several trace elements (such as Fe, Co, Ni and Mn) in the biochar, which are involved in the action of acetyl-CoA synthase and methyl coenzyme M reductase to catalyse key metabolic steps, especially the methanogenic stage. The pH, alkalinity, TVFA and TVFA/Alkalinity ratio values for the effluents remained within the optimal ranges for the anaerobic digestion process. It was also found that the increase in OLR in the range of 2-5 g VS/(L d) determined a proportional increase in the VS removal rate. However, when the OLR increased up to 7 g VS/(L d), a drastic decrease in the VS removal rate was found for the control reactors. Biochar amendment contributed to a more balanced state of the anaerobic process, preventing biomass washout.

Modeling of the effect of zeolite concentration on the biological nitrification process in the presence of sulfide and organic matter.

The evaluation of the nitrification kinetics in the simultaneous presence of sulfide and organic matter using zeolite as improver was the main goal of this work. According to the sensitivity and collinearity analyses, five parameters were the most sensitive in the model, whose calibrated values were: µ max, AOB = 0.02642 ± 0.002 h-1; µ max, NOB = 0.3307 ± 0.416 h-1; K S,NOB = 1.65·10-6 ± 2.85·10-6 mgHNO2-N/L; k S2 = 0.8213 ± 0.076 and n = 0.6537 ± 0.030. A good fit between the experimental data and the model's results including the effect of zeolite on the kinetic parameters was obtained, with Theil inequality coefficient values between 0.109 and 0.007 for all the variables studied, with all of these values lower than 0.3. Thus, the model proposed is robust and can simulate the nitrification process in the presence of sulfide and organic matter when zeolite was used as improver.

Mesophilic Semi-Continuous Anaerobic Digestion of Strawberry Extrudate Pretreated with Steam Explosion.

The production of strawberry concentrate produces a side stream after extrusion that is commonly landfilled. This strawberry extrudate (SE), of lignocellulosic character, contains valuable bioactive compounds such as sugars and phenols. Thermal treatments, such as steam explosion, are currently used for the valorisation of agricultural lignocellulosic wastes due to their ability to impact the structure of the lignocellulose and hemicellulose present in these wastes, favouring the disruption of fibrous material. Steam explosion has already been shown as a promising technology for phenol recovery from SE. Biogas is an additional valuable resource that might be produced from thermally pretreated and de-phenolised SE. This study assessed the influence of a steam-explosion pretreatment and the subsequent recovery of phenolic compounds from the long-term operation of a semi-continuous anaerobic digester of pretreated SE. The anaerobic digestion of SE steam exploded at 220 °C for 5 min and de-phenolised was stable at an OLR of 0.5 g of volatile solids (VS)/(L·d), which permitted a specific production rate of 135 ± 11 mL of CH4/(g of VS·d). The system was not able to operate at an OLR of 1 g of VS/(L·d), which resulted in a failure of the process. Despite the inhibition threshold of phenolic compounds not being achieved, the inhibition of the anaerobic digestion process at an OLR of 1 g of VS/(L·d) was most likely due to the overloading of the system. This was indicated by the accumulation of soluble organic matter and volatile fatty acids. The increase in the propionic acid concentration up to 1300 mg/L when operating at OLRs higher than 0.5 g of VS/(L·d) could be the main factor responsible for the inhibition. An economic evaluation showed that the proposed approach (steam explosion, phenol recovery, and anaerobic digestion) would offer positive benefits, taking into account the high phenolic recovery (0.90 g of gallic acid equivalents/kg of extrudate) and the low sales price of the phenol extract, i.e., EUR 0.610/g of gallic acid equivalents, needed to reach zero net profit.

High-Value-Added Compound Recovery with High-Temperature Hydrothermal Treatment and Steam Explosion, and Subsequent Biomethanization of Residual Strawberry Extrudate.

This study was on the comparison of hydrothermal treatments at 170 °C (steam injection) and 220 °C (steam explosion) to solubilize the organic matter contained in residual strawberry extrudate, focusing on phenolic compounds that were susceptible to be extracted and on sugars. After the extraction step, the remaining strawberry extrudate phases were subjected to anaerobic digestion to generate biogas that would compensate the energy requirements of the suggested hydrothermal treatments and to stabilize the remaining waste. Hydrothermal treatment at 220 °C allowed the recovery of 2053 mg of gallic acid eq. per kg of residual strawberry extrudate. By contrast, after hydrothermal treatment at 170 °C, only 394 mg of gallic acid eq. per kg of residual strawberry extrudate was recovered. Anaerobic digestion processes were applied to the de-phenolized liquid phase and the solid phase together, which generated similar methane productions, i.e., around 430 mL CH4/g volatile solids, after both 170 °C and 220 °C hydrothermal treatments. Considering the latest observation, hydrothermal treatment at 220 °C is a preferable option for the valorization of residual strawberry extrudate (RSE) due to the high solubilization of valuable phenolic compounds that can be recovered.

Fly ash as stimulant for anaerobic digestion: effect over hydrolytic stage and methane generation rate.

Thermoelectric fly ash was used as a micronutrient source for microorganisms in the anaerobic digestion process of thermally pretreated (1 hour, 120 °C) secondary sludge. The obtained results not only suggest that fly ash improves methane generation in the conversion of volatile fatty acids into methane, but also show a new observation, that the fly ash contributes in the particulate organic solubilization. The maximum methane production rate increased from 6.52 mL/L/d to 22.59 mL/L/d when fly ash was added at a dosage of 150 mg/L in biochemical methane potential tests compared with tests with no added ash. Additionally, the kinetic constants of the hydrolysis of particulate organic matter were obtained in both cases (with and without added ash) in batch reactors using a first-order kinetic model; in the case of no addition, the first-order kinetic parameter was 0.019 ± 0.002 d-1, while with ashes this value increased to 0.045 ± 0.000 d-1. Therefore, the addition of fly ash improves methane generation and hydrolytic kinetics in different orders of magnitude.

Anaerobic digestion of wastewater rich in sulfate and sulfide: effects of metallic waste addition and micro-aeration on process performance and methane production.

This work explores the effect of two metallic wastes (mining wastes, MW; fly ashes, FA) and micro-aeration (MA) on the anaerobic digestion of wastewater which is rich in sulfate and sulfide. Two initial COD concentrations (5,000 and 10,000 mg/L) were studied under both conditions in batch systems at 35 °C, with a fixed COD/SO42- ratio = 10, with 100 mg/L of S2-. It was observed that the use of MW and FA in the assays with an initial COD concentration of 10,000 mg/L resulted in a simultaneous increase in COD removal, sulfate removal, sulfide removal and methane generation, while MA only improved the COD and sulfide removals in comparison with the control system. On the contrary, the use of MW, FA or MA in systems with initial COD concentrations equal to or lower than 5,000 mg/L did not show any improvement with respect to the control system in terms of COD removal, sulfate removal or methane generation, with only sulfide removal being positively affected by MW and FA.

Performance evaluation of mesophilic semi-continuous anaerobic digestion of high-temperature thermally pre-treated olive mill solid waste.

The aim of the present work was to evaluate the effects of a thermal pre-treatment of olive mill solid waste (OMSW) and phenol extraction process on the semi-continuous anaerobic digestion of this pre-treated waste during a prolonged operational period (275 days) in order to assess the organic loading rates (OLR) of 1 ad 2 g Volatile Solids (VS)/(L·d). The anaerobic digestion of thermally pre-treated and de-phenolized OMSW was stable at an OLR of 1 g VS/(L·d), which permitted a specific production rate of 172 ±â€¯60 mL CH4/(g VS·d). However, the system was not able to operate at an OLR of 2 g VS/(L·d), which resulted in the total failure of the process. Regardless of the applied OLR, the phenolic compounds were effectively degraded and the inhibition thresholds were not reached. The inhibition of the anaerobic digestion process at an OLR of 2 g VS/(L·d) was probably due to the overloading of the system, indicated by the accumulation of organic matter and volatile fatty acids. The operation of the anaerobic digester under stable conditions allowed for high profitability for the proposed bio-refinery concept, which would still be profitable at a phenol extract price above 51.8 €/kg, which is 90% lower than the current price of 520 €/kg.

Valuable Compound Extraction, Anaerobic Digestion, and Composting: A Leading Biorefinery Approach for Agricultural Wastes.

In a society where the environmental conscience is gaining attention, it is necessary to evaluate the potential valorization options for agricultural biomass to create a change in the perception of the waste agricultural biomass from waste to resource. In that sense, the biorefinery approach has been proposed as the roadway to increase profit of the agricultural sector and, at the same time, ensure environmental sustainability. The biorefinery approach integrates biomass conversion processes to produce fuels, power, and chemicals from biomass. The present review is focused on the extraction of value-added compounds, anaerobic digestion, and composting of agricultural waste as the biorefinery approach. This biorefinery approach is, nevertheless, seen as a less innovative configuration compared to other biorefinery configurations, such as bioethanol production or white biotechnology. However, any of these processes has been widely proposed as a single operation unit for agricultural waste valorization, and a thoughtful review on possible single or joint application has not been available in the literature up to now. The aim is to review the previous and current literature about the potential valorization of agricultural waste biomass, focusing on valuable compound extraction, anaerobic digestion, and composting of agricultural waste, whether they are not, partially, or fully integrated.

Phenols recovery after steam explosion of Olive Mill Solid Waste and its influence on a subsequent biomethanization process.

A promising source of high added value compounds is the Olive Mill Solid Waste (OMSW). The aim of this research was to evaluate the viability of a biorefinery approach to valorize OMSW through the combination of steam explosion, phenols extraction, and anaerobic digestion. Steam explosion treatment increased the total phenol content in the steam exploited OMSW, which was twice than that the total phenol content in raw OMSW, although some undesirable compounds were also formed. Phenol extraction allowed the recovery of 2098mg hydroxytyrosol per kg of OMSW. Anaerobic digestion allowed the partial stabilization of the different substrates, although it was not improved by the steam explosion treatment. The economic suitability of the proposed biorefinery approach is favorable up to a phenol extract price 90.7% lower than the referenced actual price of 520€/kg.

Sequential adaptation of Nannochloropsis gaditana to table olive processing water.

The main objective of this study was to evaluate the suitability of Nannochloropsis gaditana to grow by sequential adaptation to TOPW (Table olive processing water) at increased substrate concentrations (10-80%). Sequential adaptation allows growing Nannochloropsis gaditana up to 80% TOPW, although the maximum microalgae biomass productions were achieved for percentages of 20-40%, i.e. 0.308 ± 0.005 g VSS (Volatile Suspended Solids)/L. In all growth experiments, proteins were the majority compound in the grown microalgae biomass (0.44 ± 0.05 g/g VSS), whereas phenols were retained up to a mean concentration of 12.1 ± 1.9 mg total phenols/g VSS. The highest microalgae biomass production rate at rate of 80% TOPW took place in the first two days when most nutrients were also removed. Average removal efficiencies at this percentage of TOPW were 69.1%, 50.9%, 54.3% and 71.8% for total organic carbon, total soluble nitrogen, phosphate and total phenols, respectively. Sequential adaptation can ensure the obtaining of a sustainable microalgae culture as a treatment method for TOPW.