Algal Sugars Research Articles

Enrichable consortia of microbial symbionts degrade macroalgal polysaccharides in Kyphosus fish.

Coastal herbivorous fishes consume macroalgae, which is then degraded by microbes along their digestive tract. However, there is scarce genomic information about the microbiota that perform this degradation. This study explores the potential of Kyphosus gastrointestinal microbial symbionts to collaboratively degrade and ferment polysaccharides from red, green, and brown macroalgae through in silico study of carbohydrate-active enzyme and sulfatase sequences. Recovery of metagenome-assembled genomes (MAGs) from previously described Kyphosus gut metagenomes and newly sequenced bioreactor enrichments reveals differences in enzymatic capabilities between the major microbial taxa in Kyphosus guts. The most versatile of the recovered MAGs were from the Bacteroidota phylum, whose MAGs house enzyme collections able to decompose a variety of algal polysaccharides. Unique enzymes and predicted degradative capacities of genomes from the Bacillota (genus Vallitalea) and Verrucomicrobiota (order Kiritimatiellales) highlight the importance of metabolic contributions from multiple phyla to broaden polysaccharide degradation capabilities. Few genomes contain the required enzymes to fully degrade any complex sulfated algal polysaccharide alone. The distribution of suitable enzymes between MAGs originating from different taxa, along with the widespread detection of signal peptides in candidate enzymes, is consistent with cooperative extracellular degradation of these carbohydrates. This study leverages genomic evidence to reveal an untapped diversity at the enzyme and strain level among Kyphosus symbionts and their contributions to macroalgae decomposition. Bioreactor enrichments provide a genomic foundation for degradative and fermentative processes central to translating the knowledge gained from this system to the aquaculture and bioenergy sectors.IMPORTANCESeaweed has long been considered a promising source of sustainable biomass for bioenergy and aquaculture feed, but scalable industrial methods for decomposing terrestrial compounds can struggle to break down seaweed polysaccharides efficiently due to their unique sulfated structures. Fish of the genus Kyphosus feed on seaweed by leveraging gastrointestinal bacteria to degrade algal polysaccharides into simple sugars. This study reconstructs metagenome-assembled genomes for these gastrointestinal bacteria to enhance our understanding of herbivorous fish digestion and fermentation of algal sugars. Investigations at the gene level identify Kyphosus guts as an untapped source of seaweed-degrading enzymes ripe for further characterization. These discoveries set the stage for future work incorporating marine enzymes and microbial communities in the industrial degradation of algal polysaccharides.

Unique alcohol dehydrogenases involved in algal sugar utilization by marine bacteria

Marine algae produce complex polysaccharides, which can be degraded by marine heterotrophic bacteria utilizing carbohydrate-active enzymes. The red algal polysaccharide porphyran contains the methoxy sugar 6-O-methyl-d-galactose (G6Me). In the degradation of porphyran, oxidative demethylation of this monosaccharide towards d-galactose and formaldehyde occurs, which is catalyzed by a cytochrome P450 monooxygenase and its redox partners. In direct proximity to the genes encoding for the key enzymes of this oxidative demethylation, genes encoding for zinc-dependent alcohol dehydrogenases (ADHs) were identified, which seem to be conserved in porphyran utilizing marine Flavobacteriia. Considering the fact that dehydrogenases could play an auxiliary role in carbohydrate degradation, we aimed to elucidate the physiological role of these marine ADHs. Although our results reveal that the ADHs are not involved in formaldehyde detoxification, a knockout of the ADH gene causes a dramatic growth defect of Zobellia galactanivorans with G6Me as a substrate. This indicates that the ADH is required for G6Me utilization. Complete biochemical characterizations of the ADHs from Formosa agariphila KMM 3901T (FoADH) and Z. galactanivorans DsijT (ZoADH) were performed, and the substrate screening revealed that these enzymes preferentially convert aromatic aldehydes. Additionally, we elucidated the crystal structures of FoADH and ZoADH in complex with NAD+ and showed that the strict substrate specificity of these new auxiliary enzymes is based on a narrow active site.Key points• Knockout of the ADH-encoding gene revealed its role in 6-O-methyl-D-galactose utilization, suggesting a new auxiliary activity in marine carbohydrate degradation.• Complete enzyme characterization indicated no function in a subsequent reaction of the oxidative demethylation, such as formaldehyde detoxification.• These marine ADHs preferentially convert aromatic compounds, and their strict substrate specificity is based on a narrow active site.

Development of 2,3-Butanediol Production Process from Klebsiella aerogenes ATCC 29007 Using Extracted Sugars of Chlorella pyrenoidosa and Biodiesel-Derived Crude Glycerol

Expectation for renewable energy is increasing due to environmental pollution such as fossil fuel depletion, CO2 emission, and harmful gases. Therefore, in this study, extracted sugars of microalgae, which cause algal blooms and crude glycerol, a biodiesel industry byproduct, were used simultaneously to produce 2,3-BDO. The 2,3-BDO production using only extracted algal sugars was about 4.8 g/L at 18 h, and the production of 2,3-BDO using both extracted algal sugar and crude glycerol was about 7 g/L at 18 h. It was confirmed that the main culture with crude glycerol was increased 1.5-fold compared to the case of using only extracted algal sugars. In addition, four components of the main medium (ammonium sulfate, casein hydrolysate, yeast extract, and crude glycerol) were statistically optimized and the concentrations of the medium were 12, 16, 12, and 13 g/L, respectively. In addition, the final 2,3-BDO production was about 11g/L, which 1.6-fold higher than before the optimization process. As a result, it was confirmed that 2,3-BDO production is possible through the simultaneous use of algal sugars and crude glycerol, which can greatly contribute to the development of zero-waste processes.

Symbiosis extended: exchange of photosynthetic O2 and fungal-respired CO2 mutually power metabolism of lichen symbionts

Lichens are a symbiosis between a fungus and one or more photosynthetic microorganisms that enables the symbionts to thrive in places and conditions they could not compete independently. Exchanges of water and sugars between the symbionts are the established mechanisms that support lichen symbiosis. Herein, we present a new linkage between algal photosynthesis and fungal respiration in lichen Flavoparmelia caperata that extends the physiological nature of symbiotic co-dependent metabolisms, mutually boosting energy conversion rates in both symbionts. Measurements of electron transport by oximetry show that photosynthetic O2 is consumed internally by fungal respiration. At low light intensity, very low levels of O2 are released, while photosynthetic electron transport from water oxidation is normal as shown by intrinsic chlorophyll variable fluorescence yield (period-4 oscillations in flash-induced Fv/Fm). The rate of algal O2 production increases following consecutive series of illumination periods, at low and with limited saturation at high light intensities, in contrast to light saturation in free-living algae. We attribute this effect to arise from the availability of more CO2 produced by fungal respiration of photosynthetically generated sugars. We conclude that the lichen symbionts are metabolically coupled by energy conversion through exchange of terminal electron donors and acceptors used in both photosynthesis and fungal respiration. Algal sugars and O2 are consumed by the fungal symbiont, while fungal delivered CO2 is consumed by the alga.

Continuous production of bioethanol using microalgal sugars extracted from Nannochloropsis gaditana

We developed a continuous production process of bioethanol from sugars extracted from Nannochloropsis gaditana. To improve algal sugar production, the reaction conditions of acid-thermal hydrolysis were investigated based on five different types of acid and their concentrations (1-4%), and the loading ratio of solid/liquid (S/L). As a result, the maximum hydrolysis efficiency (92.82%) was achieved under 2% hydrochloric acid with 100 g/L biomass loading at 121 oC for 15 min. The hydrolysates obtained from N. gaditana were applied to the main medium of Bretthanomyces custersii H1-603 for bioethanol production. The maximum bioethanol production and yield by the microalgal hydrolysate were found to be 4.84 g/L and 0.37 g/g, respectively. In addition, the cell immobilization of B. custersii was carried out using sodium alginate, and the effect of the volume ratio of cell/sodium alginate on bioethanol productivity was investigated in a batch system. The optimal ratio was determined as 2 (v/v), and the immobilized cell beads were applied in the continuous stirred tank reactor (CSTR). Continuous ethanol production was performed using both free cells and immobilized cells at 1 L CSTR. In both groups, the maximum bioethanol production and yield were achieved at dilution rate of 0.04 h-1 (3.93 g/L and 0.3 g/g by free cell, and 3.68 g/L and 0.28 g/g by immobilized cell, respectively).

Pretreatment and fermentation of salt-water grown algal biomass as a feedstock for biofuels and high-value biochemicals

Utilization of halotolerant algal strains and saline water resources is necessary to increase the areas available for algal cultivation and to mitigate fresh water use issues. However, the added salt content from salt-water algae cultivation may impact downstream biomass pretreatment and conversion processes especially when combined with the salt that is generated as part of our Combined Algal Processing (CAP) scheme which includes pretreatment with sulfuric acid for cell disruption and carbohydrate hydrolysis. Here we compared the pretreatment, fermentation, and lipid extraction processes on salt-water versus fresh-water grown algal biomass. Response surface pretreatment plots showed that a broader range of conditions for the salt-water grown algae species yield >90% sugar yield compared to a narrower range for fresh-water species. Despite this, we anticipated that high salt content would inhibit fermentation of algal sugars, a key element the CAP scheme and sought to reduce the formation of additional salt from pretreatment and neutralization by substituting oxalic acid for sulfuric acid. The sugar release response surface for oxalic acid was of a different shape than that of sulfuric acid but achieved >90% sugar release at 2% acid. Fermentation results showed that for Saccharomyces cerevisiae, the added salts had minimal impact on sugar utilization rates or ethanol production. For Actinobacillus succinogenes however, a significant delay was observed due to the additional salts and fermentation was further delayed or inhibited by pretreatment with oxalic acid. This was remedied by removal of the oxalic acid either by the addition of CaCO3 or trioctylamine whereupon fermentation rates recovered. Differences in lipid extraction yields were observed from the fermentation broth between the salt-water grown and fresh-water grown algal biomass and was species dependent.

Multi-omic characterization of laboratory-evolved Saccharomyces cerevisiae HJ7-14 with high ability of algae-based ethanol production.

In this study, an evolved Saccharomyces cerevisiae HJ7-14 with high ability of algae-based ethanol production was characterized by multi-omic approaches. Genome sequencing of the HJ7-14 revealed a point mutation in the GAL83 gene (G703A) involved in the catabolite repression as well as the galactose metabolism. Cultural and transcriptional analyses of a S. cerevisiae mutant with chromosomal GAL83(G703A) indicated that the catabolite repression onto the galactose metabolism was considerably relieved in all cell growth stages. Untargeted metabolomic approach revealed that metabolic phenotypes between the control D452-2 and HJ7-14 strains were clearly discriminated in time-dependent manner. Especially in early growth stage at 6h, the HJ7-14 showed dramatic and coordinated alteration in central carbon and amino acid metabolisms. Through metabolomic re-organization, fold changes in fatty acid metabolism and metabolites related to stress response system were also found upon glucose depletion and active galactose utilization. Multi-omic characterization using genome sequencing, transcription, and metabolome profiling clearly unveiled that the GAL83 gene mutation partially relieved glucose-dependent catabolite repression and allowed the evolved HJ7-14 to efficiently convert algal sugars to ethanol. Our finding could be applicable for engineering of S. cerevisiae able to covert red algal biomass to other biofuels and biochemicals.

Bioethanol Production by Various Hydrolysis and Fermentation Processes with Micro and Macro Green Algae

Algae are considered as a good substrate for bioethanol production. This study was carried out to evaluate the potential chemical and biological hydrolysis of the macrogreen alga Ulva fasciata and the microgreen alga Chlorella vulgaris to pretreatment of cell wall and sugar production. Different concentrations of reduced sugar extracted from U. faciata was studied to obtain the maximum amount of bioethanol. Saccharomyces cerevisiae SH02, Pseudomonas sp. SH03, were used to determine the best microorganisms that affect in fermentation process and hence the production of bioethanol. U. fasciata contains maximum amount of sugar. Chemical hydrolysis is the best methods for pretreatment of algae. S. cerevisiae SH02 was the best microorganisms for fermentation process with sugar obtained from U. faciata by chemical hydrolysis. The highest ethanol production by S. cerevisiae SH02 was 40% with 5% sugar concentration that produced by acid hydrolysis and with increasing sugar ratio ethanol production decrease. The highest ethanol yield with Pseudomonas sp. SH03 was 22% obtained by fermentation of 5% algal sugar.

Utilization of algal sugars and glycerol for enhanced cephalosporin C production by Acremonium chrysogenum M35.

Microalgae are the biomass containing various components, such as carbohydrates, lipids, and amino acids. In this study, carbon sources contained in microalgae were obtained by acid extraction, and cephalosporin C (CPC), a β-lactam antibiotic intermediate, was produced by using Acremonium chrysogenum M35. In addition, the increase of CPC production was not distinct for A.chrysogenum M35 with algal sugars as the only carbon source; therefore, glycerol was added, increasing the CPC production. Thus, cheap residues such as algal sugars form microalgal and glycerol form biodiesel process could be used as the alternative sources for the production of various products.

The Enzymatic Conversion of Major Algal and Cyanobacterial Carbohydrates to Bioethanol

The production of fuels from biomass is categorized as first-, second- or third-generation depending upon the source of raw materials, either food crops, lignocellulosic material, or algal biomass, respectively. Thus far, the emphasis has been on using food crops creating several environmental problems. To overcome these problems, there is a shift toward bioenergy production from non-food sources. Algae, which store high amounts of carbohydrates, are a potential producer of raw materials for sustainable production of bioethanol. Algae store their carbohydrates in the form of food storage sugars and structural material. In general, algal food storage polysaccharides are composed of glucose subunits, however they vary in the glycosidic bond that links the glucose molecules. In starch-type polysaccharides (starch, floridean starch, and glycogen), the glucose subunits are linked together by α-(1→4) and α-(1→6) glycosidic bonds. Laminarin-type polysaccharides (laminarin, chrysolaminarin, and paramylon) are made of glucose subunits that are linked together by β-(1→3) and β-(1→6) glycosidic bonds. In contrast to food storage polysaccharides, structural polysaccharides vary in composition and glycosidic bond. The industrial production of bioethanol from algae requires efficient hydrolysis and fermentation of different algal sugars. However, the hydrolysis of algal polysaccharides employs more enzymatic mixes in comparison to terrestrial plants. Similarly, algal fermentable sugars display more diversity than plants, and therefore more metabolic pathways are required to produce ethanol from these sugars. In general, the fermentation of glucose, galactose, and glucose isomers is carried out by wild type strains of Saccharomyces cerevisiae and Zymomonas mobilis. In these strains, glucose enters glycolysis, where is it converted to pyruvate through either Embden-Meyerhof-Parnas pathway or Entner-Doudoroff pathway. Other monosaccharides must be converted to fermentable sugars before entering glycolysis. In contrast, microbial wild type strains are not capable of producing ethanol from alginate, and therefore the production of bioethanol from alginate was achieved by using genetically engineered microbial strains, which can simultaneously hydrolyze and ferment alginate to ethanol. In this review, we emphasize the enzymatic hydrolysis processes of different algal polysaccharides. Additionally, we highlight the major metabolic pathways that are employed to ferment different algal monosaccharides to ethanol.

Enhancement of hydrolysis of Chlorella vulgaris by hydrochloric acid.

Chlorella vulgaris is considered as one of the potential sources of biomass for bio-based products because it consists of large amounts of carbohydrates. In this study, hydrothermal acid hydrolysis with five different acids (hydrochloric acid, nitric acid, peracetic acid, phosphoric acid, and sulfuric acid) was carried out to produce fermentable sugars (glucose, galactose). The hydrothermal acid hydrolysis by hydrochloric acid showed the highest sugar production. C. vulgaris was hydrolyzed with various concentrations of hydrochloric acid [0.5-10% (w/w)] and microalgal biomass [20-140g/L (w/v)] at 121°C for 20min. Among the concentrations examined, 2% hydrochloric acid with 100g/L biomass yielded the highest conversion of carbohydrates (92.5%) into reducing sugars. The hydrolysate thus produced from C. vulgaris was fermented using the yeast Brettanomyces custersii H1-603 and obtained bioethanol yield of 0.37g/g of algal sugars.

The Production of Ethanol from Micro-Algae Spirulina

This study ascertained the technical potential of producing bio fuel from naturally occurring micro algae spirulina. The algae are grown by BCSIR and the collected algae are examined. The growing environment contains nitrates, phosphates and carbon dioxide that come from atmosphere. The overall research work consisted of manual and mechanical drying of algae, grinding and subjecting the algal matter to acid hydrolysis to extract carbohydrates to form an algal sugar solution. Fermentation of that solution was performed with Saccharomyces cerevisiae to produce ethanol. The presents of ethanol in the solution assures the ultimate desired destination of the work. An average of 15.2g/L of reducing sugar was extracted in the hydrolysate showing that microalgae have significant usable carbohydrates after hydrolysis to conduct this research work. Ethanol percentage varies with the variation of hydrolysis time, concentration of sulfuric acid and fermentation time. The average value of ethanol percentage varies between 0.85-1.0 percentamong the samples. Although the ethanol percentage is not economical for this algae but the main task of establishing the process of bio fuel production from micro algae comes to a successful end. This process gives the assurance and defined way of bio fuel production from other carbohydrate enriched micro algae.

The production of butanol from Jamaica bay macro algae

AbstractThis study ascertained the technical potential of producing biofuel from a naturally occurring macroalgae. The algae examined grow in Jamaica Bay, New York City, on water containing nitrates, phosphates, and carbon dioxide that comes from the atmosphere. The process consisted of manual and mechanical harvesting, drying, grinding, and subjecting the algal matter to acid hydrolysis to extract carbohydrates to form an algal sugar solution. Fermentation of that solution to butanol was performed with butanol ultimately removed by distillation. An average of 15.2 g/L of reducing sugars was extracted in the hydrolysate showing that macroalgae (Ulva lactuca) have significant usable carbohydrates after hydrolysis. It was found necessary to remove the excess solids from the hydrolysate prior to fermentation, as the productivity fell by 75% if this was not done. With the bacterial strains (Clostridium beijerinckii and C. saccharoperbutylacetonicum) and the algal sugar solutions used, an acetone butanol ethanol (ABE) fermentation was used to make butanol. The butanol concentration in the fermentation broth reached about 4 g/L, which is close to the theoretical value for the sugar concentration obtained, and compares well (when adjusted for sugar concentration in the media) with values reported in the literature for other systems. The recovery of reducing sugars in the media during the pilot study was 0.29 g butanol/g sugar. © 2011 American Institute of Chemical Engineers Environ Prog, 2012

The Effect of Puromycin and Cycloheximide on the Infection of Algae‐Free Paramecium bursaria by Symbiotic Chlorellae1

ABSTRACTIt has been suggested that the infection of algae‐free Paramecium bursaria by symbiotic algae involves an induction in the ciliate. Such a process suggests a need for the synthesis of specific proteins. Therefore, an attempt was made to determine the role of protein synthesis during the initial phases of host‐symbiont interaction by examining the capacity of the ciliate to form a stable association with algae when the ciliate is exposed to puromycin (PURO) or cycloheximide (CYC) during the first 1–3 h of algal insestion. Cycloheximide (100 μg/ml) blocked algal but not ciliate growth and protein synthesis while PURO (250 μg/ml) appeared to inhibit these processes in both Puromycin significantly inhibited the infection when presented to the ciliate during the first hour of algal exposure and had little effect when added after that period. Inhibition of ciliate, as compared to the alga, protein synthesis appears to be significant in relationship to those processes leading to infection, as CYC when presented during the first hour of algae‐ciliate exposure has no inhibitory effects. Experiments on algal sugar secretion and ciliate ingestion of algae indicated that neither process was significantly affected by these inhibitors. These results point to a need for host protein synthesis during the initial phase of ingestion of algae which appears to be important to establishment of the symbiotic association.

Glass capillary or fused-silica gas chromatography—mass spectrometry of several monosaccharides and related sugars: Improved resolution

The gas chromatographic separation of several monosaccharides and related sugars derivatized by methoxylation and trimethylsilylation reactions was optimized with glass capillary (SP-2250) and fused silica (SP-2100) columns. Individual sugars included aldoses, ketoses, polyols, acidic forms and N-acetylated amino sugars. Peaks were detected by selected ion monitoring (SIM). The fused silica column gave complete resolution of all peaks (two per hexose and one per hexitol) arising from glucose, galactose, mannose, fructose, sorbitol, mannitol and dulcitol. The resolution of these sugars with the glass capillary column was not as good, but full differentiation was possible on the basis of SIM. Because the fused silica column gave a better resolution of 33 sugars tested and was more easily installed than the glass capillary column, it was utilized for quantitative analysis. A deuterated algal sugar mixture used for quantitation by isotope dilution was found to contain glucose, galactose, mannose, xylose, arabinose, ribose and rhamnose. Full recoveries were obtained of various amounts of glucose, galactose, mannose, fructose and xylose added to human serum.