Biorefineries serve to efficiently utilize biomass and their by-products. Algal biorefineries are designed to generate bioproducts for commercial use. Due to the high carbohydrate content of algal biomass, biorefinery to generate biofuels, such as bioethanol, is of great interest. Carrageenan is a predominant polysaccharide hydrocolloid found in red macroalgae and is widely used in food, cosmetics, and pharmaceuticals. In this study, we report the biorefinery of carrageenan derived from processing of experimental strains of the red macroalgae Kappaphycus alvarezii. Specifically, the chemical composition and enzymatic hydrolysis of the residue produced from carrageenan extraction were evaluated to determine the conditions for efficient generation of carbohydrate bioproducts. The productivity and growth rates of K. alvarezii strains were assessed along with the chemical composition (total carbohydrates, ash, sulfate groups, proteins, insoluble aromatics, galacturonic acid, and lipids) of each strain. Two strains, brown and red, were selected based on their high growth rates and productivity and were treated with 6 % KOH for extraction of carrageenan. The yields of biomass from treatment with 6 % KOH solution of the brown and red strains were 89.3 and 89.5 %, respectively. The yields of carrageenan and its residue were 63.5 and 23 %, respectively, for the brown strain and 60 and 27.8 %, respectively, for the red strain. The residues from the brown and red strains were assessed to detect any potential bioproducts. The galactan, ash, protein, insoluble aromatics, and sulfate groups of the residue were reduced to comparable extents for the two strains. However, KOH treatment did not reduce the content of glucan in the residue from either strain. Glucose was produced by enzymatic hydrolysis for 72 h using both strains. The glucan conversion was 100 % for both strains, and the concentrations of glucose from the brown and red strains were 13.7 and 11.5 g L−1, respectively. The present results highlight the efficiency of generating a key bioproduct from carrageenan residue. This study demonstrates the potential for glucose production using carrageenan residue. Thus, the biorefinery of K. alvarezii can be exploited not only to produce carrageenan, but also to generate glucose for future use in biofuel production.

In this study, trypsin (Enzyme Comission 3.4.21.4) was immobilized in a low cost, lignocellulosic support (corn cob powder—CCP) with the goal of obtaining peptides with bioactive potential from cheese whey. The pretreated support was activated with glyoxyl groups, glutaraldehyde and IDA-glyoxyl. The immobilization yields of the derivatives were higher than 83%, and the retention of catalytic activity was higher than 74%. The trypsin-glyoxyl-CCP derivative was thermally stable at 65 °C, a value that was 1090-fold higher than that obtained with the free enzyme. The trypsin-IDA-glyoxyl-CCP and trypsin-glutaraldehyde-CCP derivatives had thermal stabilities that were 883- and five-fold higher, respectively, then those obtained with the free enzyme. In the batch experiments, trypsin-IDA-glyoxyl-CCP retained 91% of its activity and had a degree of hydrolysis of 12.49%, while the values for trypsin-glyoxyl-CCP were 87% and 15.46%, respectively. The stabilized derivative trypsin-glyoxyl-CCP was also tested in an upflow packed-bed reactor. The hydrodynamic characterization of this reactor was a plug flow pattern, and the kinetics of this system provided a relative activity of 3.04 ± 0.01 U·g⁻¹ and an average degree of hydrolysis of 23%, which were suitable for the production of potentially bioactive peptides.

Milk whey proteins are well known for their high biological value and versatile functional properties, characteristics that allow its wide use in the food and pharmaceutical industries. In this work, a 24 kDa protein from buffalo cheese whey was analyzed by mass spectrometry and presented homology with Bos taurus beta-lactoglobulin. In addition, the proteins present in buffalo cheese whey were hydrolyzed with pepsin and with different combinations of trypsin, chymotrypsin and carboxypeptidase-A. When the TNBS method was used the obtained hydrolysates presented DH of 55 and 62% for H1 and H2, respectively. Otherwise for the OPA method the DH was 27 and 43% for H1 and H2, respectively. The total antioxidant activities of the H1 and H2 samples with and without previous enzymatic hydrolysis, determined by DPPH using diphenyl-p-picrylhydrazyl radical, was 4.9 and 12 mM of Trolox equivalents (TE) for H2 and H2Dint, respectively. The increased concentrations for H1 and H2 samples were approximately 99% and 75%, respectively. The in vitro gastrointestinal digestion efficiency for the samples that were first hydrolyzed was higher compared with samples not submitted to previous hydrolysis. After in vitro gastrointestinal digestion, several amino acids were released in higher concentrations, and most of which were essential amino acids. These results suggest that buffalo cheese whey is a better source of bioavailable amino acids than bovine cheese whey.

Rhizopus stolonifer was cultivated in wheat bran to produce a cellulase-free alkaline xylanase. The purified enzyme obtained after molecular exclusion chromatography in Sephacryl S-200 HR showed optimum temperature as 45º C and hydrolysis pHs optima as pH 6.0 and 9.0. Xylanase presented higher Vmax at pH 9.0 (0.87 µmol/mg protein) than at pH 6.0 and minor Km at pH 6.0 (7.42 mg/mL) than at pH 9.0.

Cyclodextrin glycosyltransferase (EC 2.4.1.19) is an enzyme that produces cyclodextrins from starch via an intramolecular transglycosylation reaction. An alkalophilic Bacillus strain, isolated from cassava peels, was identified as Bacillus licheniformis. CGTase production by this strain was better when potato starch was used as carbon source, followed by cassava starch and amylopectin. Glucose and amylose, on the other hand, acted as synthesis repressors. When the cultivation was supplemented with sodium ions and had the pH adjusted between 6.0 and 9.0, the microorganism maintained the growth and enzyme production capacity. This data is interesting because it contradicts the concept that alkalophilic microorganisms do not grow in this pH range. After ultrafiltration-centrifugation, one protein of 85.2 kDa with CGTase activity was isolated. This protein was identified in plates with starch and phenolphthalein. Determination of the optimum temperature showed higher activities at 25 degrees C and 55 degrees C, indicating the possible presence of more than one CGTase in the culture filtrate. Km and Vmax values were 1.77 mg/mL and 0.0263 U/mg protein, respectively, using potato starch as substrate

Three strains of Bacillus sp. (BACRP, BACNC-1 and BACAR) were isolated from soil adhered to cassava husk. CGTase specific activity for the three isolated strains was higher when cultivated at 40°C. Potato starch, cassava starch, maltodextrin and glucose were used as carbon source and growth temperatures varied from 25 to 55°C. The three isolates presented higher CGTase specific activity when cultivated with potato starch at 40°C. Isolated BACRP and BACAR presented specific activity of 4.0×10(-3) and 2.2×10(-3) U/mg prot at pH 7.0, respectively, when cultivated in mediums added with NaCl 2%; at pH 10,0 their activities were of 3.4×10(-3) and 3.0×10(-3) U/mg prot, respectively, in the same concentration of NaCl. On the other hand, the isolated BACNC-1 presented activity specific of 2.4×10(-3) U/mg prot when cultivated at pH 7.0 added of NaCl 1%, and at pH 10.0 the specific activity was of 3.4×10(-3) U/mg prot without NaCl addition. This work also showed the presence of cyclodextrins formed during fermentation process and that precipitation with acetone or lyophilization followed by dialysis was efficient at removing CDs (cyclodextrins), thus, eliminating interference in the activity assays. The enzyme produced by the BACAR strain was partially purified and β-CD was liberated as a reaction product.