Investigating the efficiency of a two-stage anaerobic-aerobic process for the treatment of confectionery industry wastewaters with simultaneous production of biohydrogen and polyhydroxyalkanoates.

The scope of the current study was to investigate the efficiency of a two-stage anaerobic-aerobic process for the simultaneous treatment and valorization of selective wastewater streams from a confectionary industry. The specific wastewater (confectionary industry wastewater, CIW) was a mixture of the rinsing eluting during washing of the cauldrons in which jellies and syrups were produced, and contained mainly readily fermentable sugars, being thus of high organic load. The first stage of the process was the dark fermentation (DF) of the CIW in continuous, attached-biomass systems, in which the effect on hydrogen yields and distribution of metabolites were studied for different packing materials (ceramic or plastic), hydraulic retention times, HRTs (12 h-30 h) and feed substrate concentration (20 g COD/L- 50 g COD/L). In the second stage, the effectiveness of the aerobic treatment of the DF effluents was evaluated in terms of the reduction of the organic load and the production of polyhydroxyalkanoates (PHAs) through an enriched mixed microbial culture (MMC). The MMC was developed in a continuous draw and fill system, in which the accumulation potential of PHAs was studied. It was shown that the hydrogen production rates decreased for increasing substrate concentration and HRTs, with a maximum of 12.70 ± 0.35 m3 H2/m3 initial CIW achieved for the lowest HRT and feed concentration and using ceramic beads as packing material. Butyrate, acetate and lactate were the main metabolites generated in all cases, in different ratios. The distribution of metabolites during DF was shown to highly affect the efficiency of the second process in terms of both the reduction of organic load and the PHAs yields. The highest removal of organic load achieved after 48 h of aerobic treatment was 84.0 ± 0.9 %, whereas the maximum PHAs yield was 21.46 ± 0.13 kg PHAs/m3 initial CIW.

Design and Synthesis of Novel 2-Acetamido, 6-Carboxamide Substituted Benzothiazoles as Potential BRAFV600E Inhibitors - In vitro Evaluation of their Antiproliferative Activity.

The oncogenic BRAFV600E kinase leads to abnormal activation of the MAPK signaling pathway and thus, uncontrolled cellular proliferation and cancer development. Based on our previous virtual screening studies which issued 2-acetamido-1,3 benzothiazole-6-carboxamide scaffold as active pharmacophore displaying selectivity against the mutated BRAF, eleven new substituted benzothiazole derivatives were designed and synthesized by coupling of 2-acetamidobenzo[d]thiazole-6-carboxylic acid with the appropriate amines in an effort to provide even more efficient inhibitors and tackle drug resistance often developed during cancer treatment. All derived compounds bore the benzothiazole scaffold substituted at position-2 by an acetamido moiety and at position-6 by a carboxamide functionality, the NH moiety of which was further linked through an alkylene linker to a sulfonamido (or amino) aryl (or alkyl) functionality or a phenylene linker to a sulfonamido aromatic (or non-aromatic) terminal pharmacophore in the order -C6 H4 -NHSO2 -R or reversely -C6 H4 -SO2 N(H)-R. These analogs were subsequently biologically evaluated as potential BRAFV600E inhibitors and antiproliferative agents in several colorectal cancer and melanoma cell lines. In all assays applied, one analog, namely 2-acetamido-N-[3-(pyridin-2-ylamino)propyl]benzo[d]thiazole-6-carboxamide (22), provided promising results in view of its use in drug development.

Enhancement of biosynthesis of polyhydroxyalkanoates (PHA) from Taihu blue algae by adding by-product acetic acid.

As an easily obtained organic waste, by-product acetic acid could be an appropriate co-substrate with blue algae wastes (increase C/N ratio of substrates) for co-fermentation of PHA production. However, there are still acrylic acid and other chemicals in by-product acetic acid, which could cause severe inhibition for fermenting microorganisms during PHA production process. The current study represented that alkali pretreatment (pH level of 12) is a more favorable method compared with thermal pretreatment (80 â„ƒ for 30 min) for breaking cell walls of blue algae. It seemed that there was no synergistic effect of the combination of thermal and alkali pretreatment methods (temperature of 80 â„ƒ and pH level of 12). Optimal parameters during electro-fenton process for removal of inhibitors in by-product acetic acid were under current of 0.5 A, pH level of 3 and reaction time of 120 min. Both the highest dry weight of PHA and PHA concentration were achieved by applying blue algae and by-product acetic acid (after pretreatment) as co-substrates (mixed ratio of 3:1, stirring speed of 200 r/min, 24 h), indicating that using by-product acetic acid (after pretreatment) as co-substrate could increase C/N ratio and promote PHA production successfully. The current study could offer new insights for improving PHA production by co-fermentation.

Recovery of polyhydroxyalkanoates (PHAs) polymers from a mixed microbial culture through combined ultrasonic disruption and alkaline digestion.

PHAs are a form of cellular storage polymers with diverse structural and material properties, and their biodegradable and renewable nature makes them a potential green alternative to fossil fuel-based plastics. PHAs are obtained through extraction via various mechanical, physical and chemical processes after their intracellular synthesis. Most studies have until now focused on pure cultures, while information on mixed microbial cultures (MMC) remains limited. In this study, ultrasonic (US) disruption and alkaline digestion by NaOH were applied individually and in combination to obtain PHAs products from an acclimated MMC using phenol as the carbon source. Various parameters were tested, including ultrasonic sound energy density, NaOH concentration, treatment time and temperature, and biomass density. US alone caused limited cell lysis and resulted in high energy consumption and low efficiency. NaOH of 0.05-0.2 M was more efficient in cell disruption, but led to PHAs degradation under elevated temperature and prolonged treatment. Combining US and NaOH significantly improved the overall process efficiency, which could reduce energy consumption by 2/3rds with only minimal PHAs degradation. The most significant factor was identified to be NaOH dosage and treatment time, with US sound energy density playing a minor role. Under the semi-optimized condition (0.2 M NaOH, 1300 W L-1, 10 min), over 70% recovery and 80% purity were achieved from a 3 g L-1 MMC slurry of approximately 50% PHAs fraction. The material and thermal properties of the products were analyzed, and the polymers obtained from US + NaOH treatments showed comparable or higher molecular weight to previously reported results. The products also exhibited good thermal stability and rheological properties, compared to the commercial standard. In conclusion, the combined US and NaOH method has the potential in real application as an efficient process to obtain high quality PHAs from MMC, and cost-effectiveness can be further optimized.

Lysophosphatidic Acid Is a Proinflammatory Stimulus of Renal Tubular Epithelial Cells.

Chronic kidney disease (CKD) refers to a spectrum of diseases defined by renal fibrosis, permanent alterations in kidney structure, and low glomerular-filtration rate. Prolonged epithelial-tubular damage involves a series of changes that eventually lead to CKD, highlighting the importance of tubular epithelial cells in this process. Lysophosphatidic acid (LPA) is a bioactive lipid that signals mainly through its six cognate LPA receptors and is implicated in several chronic inflammatory pathological conditions. In this report, we have stimulated human proximal tubular epithelial cells (HKC-8) with LPA and 175 other possibly pathological stimuli, and simultaneously detected the levels of 27 intracellular phosphoproteins and 32 extracellular secreted molecules with multiplex ELISA. This quantification revealed a large amount of information concerning the signaling and the physiology of HKC-8 cells that can be extrapolated to other proximal tubular epithelial cells. LPA responses clustered with pro-inflammatory stimuli such as TNF and IL-1, promoting the phosphorylation of important inflammatory signaling hubs, including CREB1, ERK1, JUN, IκΒα, and MEK1, as well as the secretion of inflammatory factors of clinical relevance, including CCL2, CCL3, CXCL10, ICAM1, IL-6, and IL-8, most of them shown for the first time in proximal tubular epithelial cells. The identified LPA-induced signal-transduction pathways, which were pharmacologically validated, and the secretion of the inflammatory factors offer novel insights into the possible role of LPA in CKD pathogenesis.

Sorption of two common antihypertensive drugs onto polystyrene microplastics in water matrices.

Recent studies have shown the widespread occurrence of microplastics in multiple environmental compartments. When discharged into the aquatic environment, microplastics interact with other chemicals acting as vectors of organic and inorganic micropollutants. In the present study, we examined the sorption of two commonly used antihypertensive drugs, valsartan (VAL) and losartan (LOS), onto polystyrene (PS) microplastics and we studied the effects of water matrix, solution's pH, salinity, and microplastics' aging on their sorption. According to the results, the sorption of VAL and LOS onto PS is a slow process that reaches equilibrium after 12 days. The sorption of both target micropollutants was pH-dependent and significantly decreased under alkaline conditions. The removal of VAL was enhanced in the presence of 100 mM of Ca2+ while no statistical significant effects were observed when Na+ was added. The increase of salinity either did not affect or decreased the removal of LOS. Lower sorption of both drugs was observed when aged PS was used despite that the specific surface area for aged PS was 39% higher than pristine. Calculation of the sorption distribution coefficient (Kd) for different water matrices showed that the increase of matrix complexity inhibited target compounds' removal and the sorption rate decreased from bottled water > river water ≈ treated wastewater for the two compounds. For VAL, the Kd values ranged between 795 ± 63 L/kg (bottled water) and 384 ± 88 L/kg (river water), while for LOS between 4453 ± 417 L/kg (bottled water) and 3078 ± 716 L/kg (treated wastewater). Both VAL and LOS sorption onto PS microplastics can be described by hydrophobic and electrostatic interactions. The current results indicate that PS particles could affect the transportation of antihypertensive drugs in the aquatic environment causing potential adverse effects on the environment and public health.

A Comparative Study of Various Pretreatment Approaches for Bio-Ethanol Production from Willow Sawdust, Using Co-Cultures and Mono-Cultures of Different Yeast Strains.

The effect of different pretreatment approaches based on alkali (NaOH)/hydrogen peroxide (H2O2) on willow sawdust (WS) biomass, in terms of delignification efficiency, structural changes of lignocellulose and subsequent fermentation toward ethanol, was investigated. Bioethanol production was carried out using the conventional yeast Saccharomyces cerevisiae, as well as three non-conventional yeasts strains, i.e., Pichia stipitis, Pachysolen tannophilus, Wickerhamomyces anomalus X19, separately and in co-cultures. The experimental results showed that a two-stage pretreatment approach (NaOH (0.5% w/v) for 24 h and H2O2 (0.5% v/v) for 24 h) led to higher delignification (38.3 ± 0.1%) and saccharification efficiency (31.7 ± 0.3%) and higher ethanol concentration and yield. Monocultures of S. cerevisiae or W. anomalus X19 and co-cultures with P. stipitis exhibited ethanol yields in the range of 11.67 ± 0.21 to 13.81 ± 0.20 g/100 g total solids (TS). When WS was subjected to H2O2 (0.5% v/v) alone for 24 h, the lowest ethanol yields were observed for all yeast strains, due to the minor impact of this treatment on the main chemical and structural WS characteristics. In order to decide which is the best pretreatment approach, a detailed techno-economical assessment is needed, which will take into account the ethanol yields and the minimum processing cost.

Modeling of continuous dark fermentative hydrogen production in an anaerobic up-flow column bioreactor.

Dark fermentation (DF) of several types of wastes is a promising process to alleviate environmental pollution as it leads to the production of valuable hydrogen (H2) gas and high added value products, such as volatile fatty acids (VFAs). In this study a kinetic model for fermentative H2 production in an Up-flow column reactor (UFCR) is presented. Τhe model structure includes seven biochemical reactions taking place in a two-phase biofilm-liquid system. The observed difference in the overall stoichiometry of the bioconversion process for different hydraulic retention times (HRTs) is predicted by this model as it is attributed to the difference in the extent of individual bioconversion steps, each of which has a constant stoichiometry but a different rate depending on the HRT. The respective kinetic parameters were estimated through model fitting to the experimental results of the UFCR, which operated at different HRTs (12-2 h) and fed with the soluble fraction of a food industry waste (FIW). A good agreement of the experimental and predicted values of soluble metabolic products and H2 production was obtained, rendering this model as a useful tool for further investigation and prediction of the characteristics of the DF process in attached-biomass growth systems.

In situ biogas upgrading via cathodic biohydrogen using mitigated ammonia nitrogen during the anaerobic digestion of Taihu blue algae in an integrated bioelectrochemical system (BES).

Biohydrogen using migrated ammonia as nitrogen source, and biogas upgrading with hydrogen collected at biocathode in an integrated bioelectrochemical system (BES) were investigated, during the anaerobic digestion of Taihu blue algae. Under an applied voltage of 0.4 V, biohydrogen (202.87 mL) reached 2.34 and 2.90 times than groups with applied voltage of 0 V and 0.8 V, respectively. Moreover, biohydrogen of the group with 1000 mg/L initial ammonia addition (524.16 mL) reached 1.53 times than that the of the control. With 0.25 bar of H2 injected at the beginning (R1), highest methane production (286.62) mL and content (75.73%) were obtained. Comparing to other groups, not only microbial genus responsible for both aceticlastic and hydrogenotrophic methanogens of the group R1 were apparently enriched, but key enzymes related to methane production also acquired better abundances. Therefore, it's promising to conduct the ammonia alleviating, hydrogen producing and biogas upgrading simultaneously using BES.

Designing Efficient Processes for Sustainable Bioethanol and Bio-Hydrogen Production from Grass Lawn Waste.

The effect of thermal, acid and alkali pretreatment methods on biological hydrogen (BHP) and bioethanol production (BP) from grass lawn (GL) waste was investigated, under different process schemes. BHP from the whole pretreatment slurry of GL was performed through mixed microbial cultures in simultaneous saccharification and fermentation (SSF) mode, while BP was carried out through the C5yeast Pichia stipitis, in SSF mode. From these experiments, the best pretreatment conditions were determined and the efficiencies for each process were assessed and compared, when using either the whole pretreatment slurry or the separated fractions (solid and liquid), the separate hydrolysis and fermentation (SHF) or SSF mode, and especially for BP, the use of other yeasts such as Pachysolen tannophilus or Saccharomyces cerevisiae. The experimental results showed that pretreatment with 10 gH2SO4/100 g total solids (TS) was the optimum for both BHP and BP. Separation of solid and liquid pretreated fractions led to the highest BHP (270.1 mL H2/g TS, corresponding to 3.4 MJ/kg TS) and also BP (108.8 mg ethanol/g TS, corresponding to 2.9 MJ/kg TS) yields. The latter was achieved by using P. stipitis for the fermentation of the hydrolysate and S. serevisiae for the solid fraction fermentation, at SSF.

From waste to fuel: Energy recovery from household food waste via its bioconversion to energy carriers based on microbiological processes.

In the present study the bioconversion of dried household food waste (FORBI) to energy carriers was investigated aiming to its sustainable management and valorization. FORBI was either directly fermented towards ethanol and hydrogen or was previously subjected to extraction with water resulting to a liquid fraction (extract) rich in sugars and a solid residue, which were then fermented separately. Subsequently, the effluents were assessed as substrates for methane production via anaerobic digestion (AD). Mono-cultures and co-cultures of C5 and C6 yeasts were used for the alcoholic fermentation whereas for the production of hydrogen, mixed acidogenic consortia were used. Taking into account the optimum yields of biofuels, the amount of recoverable energy was estimated based for each different approach. The maximum ethanol yield was 0.16 g ethanol per kg of FORBI and it was achieved for separate fermentation of liquid and solid fractions of the waste. The highest hydrogen yield that was observed was 210.44 L ± 4.02 H2/kg TS FORBI for 1% solids loading and supplementation with cellulolytic enzymes. Direct AD of either the whole FORBI or its individual fractions led to lower overall energy recovery, compared to that obtained when fermentation and subsequent AD were applied. The recoverable energy was estimated for the different exploitation approaches of the waste. The maximum achieved recoverable energy was 21.49 ± 0.57 MJ/kg.

Biogas Production from Physicochemically Pretreated Grass Lawn Waste: Comparison of Different Process Schemes.

Various pretreatment methods, such as thermal, alkaline and acid, were applied on grass lawn (GL) waste and the effect of each pretreatment method on the Biochemical Methane Potential was evaluated for two options, namely using the whole slurry resulting from pretreatment or the separate solid and liquid fractions obtained. In addition, the effect of each pretreatment on carbohydrate solubilization and lignocellulossic content fractionation (to cellulose, hemicellulose, lignin) was also evaluated. The experimental results showed that the methane yield was enhanced with alkaline pretreatment and, the higher the NaOH concentration (20 g/100 gTotal Solids (TS)), the higher was the methane yield observed (427.07 L CH4/kg Volatile Solids (VS), which was almost 25.7% higher than the BMP of the untreated GL). Comparing the BMP obtained under the two options, i.e., that of the whole pretreatment slurry with the sum of the BMPs of both fractions, it was found that direct anaerobic digestion without separation of the pretreated biomass was favored, in almost all cases. A preliminary energy balance and economic assessment indicated that the process could be sustainable, leading to a positive net heat energy only when using a more concentrated pretreated slurry (i.e., 20% organic loading), or when applying NaOH pretreatment at a lower chemical loading.

On the evaluation of different saccharification schemes for enhanced bioethanol production from potato peels waste via a newly isolated yeast strain of Wickerhamomyces anomalus.

The present study focuses on the exploration of the potential use of potato peels waste (PPW) as feedstock for bioethanol production, using a newly isolated yeast strain, Wickerhamomyces anomalus, via different saccharification and fermentation schemes. The saccharification of PPW was performed via thermal and chemical (acid, alkali) pretreatment, as well as via enzymatic hydrolysis through the use of commercial enzymes (cellulase and amylase) or enzymes produced at lab scale (alpha-amylase from Bacillus sp. Gb67), either separately or in mixtures. The results indicated that the enzymatic treatment by commercial enzymes led to a higher saccharification efficiency (72.38%) and ethanol yield (0.49 g/gconsumed sugars) corresponding to 96% of the maximum theoretical. In addition, acid pretreatment was found to be beneficial for the process, leading also to high hydrolysis and ethanol yields, indicating that PPW is a very promising feedstock for bio-ethanol production by W. anomalus under different process schemes.

Lewis-Brønsted acid catalysed ethanolysis of the organic fraction of municipal solid waste for efficient production of biofuels.

A combined Lewis-Brønsted acid ethanolysis of sugars was thoroughly investigated with the aim of producing ethyl levulinate (EL) in a single step. Ethanolysis carried out at 453 K for 4 h using H2SO4 (1 wt%) and AlCl3·6H2O (30 mol % with respect to sugars) produced a yield of 60 mol % of EL respect to glucose and starch. Such optimised conditions were positively applied directly on different food waste, preliminarily characterised and found to be mainly composed by simple (10-15%) and relatively complex sugars (20-60%), besides proteins (6-10%) and lipids (4-10%), even in their wet form. The catalytic system resulted robust enough to the point that the copresence of proteins, lignin, lipids and mineral salts not only did not negatively affect the overall reactivity, but resulted efficiently converted into soluble species, and specifically, into other liquid biofuels of different nature.

Valorization of kitchen biowaste for ethanol production via simultaneous saccharification and fermentation using co-cultures of the yeasts Saccharomyces cerevisiae and Pichia stipitis.

The biotransformation of the pre-dried and shredded organic fraction of kitchen waste to ethanol was investigated, via co-cultures of the yeasts Saccharomyces cerevisiae and Pichia stipitis (Scheffersomyces stipitis). Preliminary experiments with synthetic media were performed, in order to investigate the effect of different operational parameters on the ethanol production efficiency of the co-culture. The control of the pH and the supplementation with organic nitrogen were shown to be key factors for the optimization of the process. Subsequently, the ethanol production efficiency from the waste was assessed via simultaneous saccharification and fermentation experiments. Different loadings of cellulolytic enzymes and mixtures of cellulolytic with amylolytic enzymatic blends were tested in order to enhance the substrate conversion efficiency. It was further shown that for solids loading up to 40% waste on dry mass basis, corresponding to 170 g.L-1 initial concentration of carbohydrates, no substrate inhibition occurred, and ethanol concentration up to 45 g.L-1 was achieved.

A Leptolyngbya-based microbial consortium for agro-industrial wastewaters treatment and biodiesel production.

A mixed cyanobacterial-mixotrophic algal population, dominated by the filamentous cyanobacterium Leptolyngbya sp. and the microalga Ochromonas (which contributed to the total photosynthetic population with rates of less than 5%), was studied under non-aseptic conditions for its efficiency to remove organic and inorganic compounds from different types of wastes/wastewaters while simultaneously producing lipids. Second cheese whey, poplar sawdust, and grass hydrolysates were used in lab-scale experiments, in photobioreactors that operated under aerobic conditions with different initial nutrient (C, N and P) concentrations. Nutrient removal rates, biomass productivity, and the maximum oil production rates were determined. The highest lipid production was achieved using the biologically treated dairy effluent (up to 14.8% oil in dry biomass corresponding to 124 mg L-1) which also led to high nutrient removal rates (up to 94%). Lipids synthesized by the microbial consortium contained high percentages of saturated and mono-unsaturated fatty acids (up to 75% in total lipids) for all the substrates tested, which implies that the produced biomass may be harnessed as a source of biodiesel.

A novel approach of modeling continuous dark hydrogen fermentation.

In this study a novel modeling approach for describing fermentative hydrogen production in a continuous stirred tank reactor (CSTR) was developed, using the Aquasim modeling platform. This model accounts for the key metabolic reactions taking place in a fermentative hydrogen producing reactor, using fixed stoichiometry but different reaction rates. Biomass yields are determined based on bioenergetics. The model is capable of describing very well the variation in the distribution of metabolic products for a wide range of hydraulic retention times (HRT). The modeling approach is demonstrated using the experimental data obtained from a CSTR, fed with food industry waste (FIW), operating at different HRTs. The kinetic parameters were estimated through fitting to the experimental results. Hydrogen and total biogas production rates were predicted very well by the model, validating the basic assumptions regarding the implicated stoichiometric biochemical reactions and their kinetic rates.

Fungal pretreatment of willow sawdust and its combination with alkaline treatment for enhancing biogas production.

In this study fungal pretreatment of willow sawdust (WSD) via the white rot fungi Leiotrametes menziesii and Abortiporus biennis was studied and the effect on fractionation of lignocellulosic biomass and biochemical methane potential (BMP), was evaluated. Scanning electron microscopy (SEM) and IR spectroscopy were used to investigate the changes in the structural characteristics of the pretreated WSD. Fungal pretreatment results revealed that A. biennis is more attractive, since it resulted in higher lignin degradation and lower holocellulose uptake. Samples of the 14th and 30th d of cultivation (i.e. the middle and the end of the pretreatment experiment) with both fungi were used for BMP tests and the effect of pretreatment duration was also evaluated. BMP increase by 31 and 43% was obtained due to the cultivation of WSD with A. biennis, for 14 and 30 d, respectively. In addition, combination of biological (after 30 d of cultivation) with alkaline (NaOH 20 g/100 gTS) pretreatment was performed, in order to assess the effect of the chemical agent on biologically pretreated WSD, in terms of lignocellulosic content and BMP. Combination of alkaline with fungal pretreatment led to high lignin degradation for both fungi, while the cellulose and hemicellulose removal efficiencies were higher for combined alkaline and L. menziesii pretreatment. The maximum BMP was observed for the combined alkaline and A. biennis pretreatment and was 12.5 and 50.1% higher than the respective alkaline and fungal pretreatment alone and 115% higher than the respective BMP of raw WSD.

2-Oxoamide inhibitors of cytosolic group IVA phospholipase A2 with reduced lipophilicity.

Cytosolic GIVA phospholipase A2 (GIVA cPLA2) initiates the eicosanoid pathway of inflammation and thus inhibitors of this enzyme constitute novel potential agents for the treatment of inflammatory diseases. Traditionally, GIVA cPLA2 inhibitors have suffered systemically from high lipophilicity. We have developed a variety of long chain 2-oxoamides as inhibitors of GIVA PLA2. Among them, AX048 was found to produce a potent analgesic effect. We have now reduced the lipophilicity of AX048 by replacing the long aliphatic chain with a chain containing an ether linked aromatic ring with in vitro inhibitory activities similar to AX048.

Review of feedstock pretreatment strategies for improved anaerobic digestion: From lab-scale research to full-scale application.

When properly designed, pretreatments may enhance the methane potential and/or anaerobic digestion rate, improving digester performance. This paper aims at providing some guidelines on the most appropriate pretreatments for the main feedstocks of biogas plants. Waste activated sludge was firstly investigated and implemented at full-scale, its thermal pretreatment with steam explosion being most recommended as it increases the methane potential and digestion rate, ensures sludge sanitation and the heat needed is produced on-site. Regarding fatty residues, saponification is preferred for enhancing their solubilisation and bioavailability. In the case of animal by-products, this pretreatment can be optimised to ensure sterilisation, solubilisation and to reduce inhibition linked to long chain fatty acids. With regards to lignocellulosic biomass, the first goal should be delignification, followed by hemicellulose and cellulose hydrolysis, alkali or biological (fungi) pretreatments being most promising. As far as microalgae are concerned, thermal pretreatment seems the most promising technique so far.