Michele Vitolo

The influence of pH, temperature and hydrolyzate concentration on the removal of volatile and nonvolatile compounds from sugarcane bagasse hemicellulosic hydrolyzate treated with activated charcoal before or after vacuum evaporation

This paper analyzes the influence of pH, temperature and degree of hydrolyzate concentration on the removal of volatile and nonvolatile compounds from sugarcane bagasse hemicellulosic hydrolyzate treated with activated charcoal before or after the vacuum evaporation process. Furfural and 5-Hydroxymethylfurfural were almost totally removed in all the experiments, irrespective of pH and temperature and whether the charcoal was added before or after the vacuum evaporation process. Adding activated charcoal before the vacuum evaporation process favored the removal of phenolic compounds for all values of pH. Acetic acid, on the contrary, was most effectively removed when the activated charcoal was added after the vacuum evaporation process at an acid pH (0.92) and at the highest degree of hydrolyzate concentration (f=4). However, addition of activated charcoal before or after vacuum evaporation at an acid pH (0.92) and at the highest degree of hydrolyzate concentration (f=4) favored the removal of both acetic acid and phenolic compounds.

Xylitol production by Candida guillermondii as an approach for the utilization of agroindustrial residues

Abstract Different substrates based on hydrolyzed hemicellulosic fractions of agroindustrial residues were used for xylitol production by Candida guilliermondii FTI 20037 under semi-aerobic conditions. Batch fermentation performances were characterized and compared with those attained in a synthetic medium using d -xylose as a major carbon source. For all media tested, simultaneous utilization of hemicellulosic sugars (glucose and xylose) was observed and the highest substrate uptake rate was attained in sugar cane bagasse medium. Increased xylitol concentrations (40 g/litre) were achieved in synthetic and rice straw-media, although the highest xylitol production rate was obtained in sugar cane bagasse hydrolysate. These results show that both hydrolysates can be converted into xylitol with satisfactory yields and productivities.

Characterization of invertase entrapped into calcium alginate beads.

A solution of 10 g/L of sodium alginate (Satialgine® types used [Sanofi trademark]: SG800® and S1100® with manuronic/guluronic ratio of 0.5 and 1.2, respectively) containing invertase (0.08 g of protein/L) was dropped into 0.1 M CaCl2 solution buffered at pH 4.0, 7.0, or 8.0. The beads were left to harden in CaCl2 solution for 24 h. The high immobilization yield of 60% occurred with SG800 at pH8.0. The activity of soluble and insoluble invertase was measured against pH (2.8–8.0), sucrose concentration (4.5–45 mM), and temperature (30–60°C). Both forms presented an optimum pH of 4.6. However, the soluble invertase was stable at the overall pH interval studied, whereas insoluble invertase lost 30% of its original activity at pH > 5.0. At temperatures above 40°C, the insoluble form was more stable than the soluble one. The kinetic constants and activation energies (Ea) for free invertase were KM=41.2 mM, Vmax=0.10 mg of TRS/(min · mL), and Ea 28 kJ/mol for entrapped invertase they were (KM)ap=7.2 mM, (Vmax)ap=0.060 mg of TRS/(min · mL), and (Ea)ap=24 kJ/mol.

Recovery of inulinase using BDBAC reversed micelles

Inulinase from Candida kefyr was extracted into a reversed micelle phase of the cationic surfactant BDBAC in organic solvents (isooctane/hexanol). Factors affecting the efficiency of the reversed micellar forward-extraction including pH, concentration and type of salts, solvent/co-solvent volume ratio, surfactant concentration, buffer concentration and temperature were investigated. A high micelle size (wo ∼ 120 or RM ∼ 18 nm) was necessary to achieve almost complete inulinase solubilization in the micellar phase, which could be generated by raising the temperature to 37°C. Some parameters for optimization of the back-extraction to a fresh aqueous phase were also investigated. Under optimum conditions of forward and backward-extraction, about 90% of the total enzyme was recovered and the specific activity of the inulinase increased some 3-fold.

Environmental parameters affecting xylitol production from sugar cane bagasse hemicellulosic hydrolyzate by Candida guilliermondii

The bioconversion of xylose to xylitol by Candida guilliermondii FTI 20037 cultivated in sugar cane bagasse hemicellulosic hydrolyzate was influenced by cell inoculum level, age of inoculum and hydrolyzate concentration. The maximum xylitol productivity (0.75 g L−1 h−1) occurred in tests carried out with hydrolyzate containing 54.5 g L−1 of xylose, using 3.0 g L−1 of a 24-h-old inoculum. Xylitol productivity and cell concentration decreased with hydrolyzate containing 74.2 g L−1 of xylose.

Fermentation of sugar cane bagasse hemicellulosic hydrolysate for xylitol production: Effect of pH

Candida guilliermondii FTI 20037 was grown in sugar cane bagasse hydrolysate supplemented with (NH4)2SO4 2.0 g l−1, CaCl2 0.1 g l−1 and rice bran 20.0 g l−1, through 45-h batch tests (agitation of 200 min−1 and temperature of 30°C) with initial pH varying from 2.5 to 7.5. Under pH < 4.5 the consumption of glucose, xylose and arabinose as well as the production of xylitol and cells were inhibited. Nevertheless, at pH values ≥ 5.5 the yeast produced xylitol with a yield of 0.75 g g−1 and productivity of 0.57 g l−1 h−1. Moreover, the yeast was also capable of metabolizing the acetic acid, which is always present in media made from hydrolysates of plant material. The inhibition of xylose/xylitol bioconversion could be related to the effects of low pH and undissociated acetic acid concentration over 5.0 g l−1.

Overview of Lactobacillus plantarum as a promising bacteriocin producer among lactic acid bacteria

Chemical preservatives have been traditionally used during the manufacturing of processed products. However, the continuous growing interest of consumers for fresh and natural products makes it necessary to search for alternative compounds. In this context, food industries have been widely using lactic acid bacteria (LAB) as natural preservatives, due to their ability to produce antibacterial compounds such as bacteriocins. Similarly, pharmaceutical industries have improved the use of these bacterial peptides, with antibacterial activity, trying to reduce the indiscriminate use of antibiotics in food products for human and animal consumption. Among LAB, Lactobacillus plantarum can be adapted to various niches thanks to its ability to ferment a wide range of carbohydrates. Additionally, it can be used as starter culture in food fermentations and as an ingredient for probiotic foods, contributing to the organoleptic characteristics of foods at the same time prolonging the shelf-life and safety of these products. The amount of valuable substances obtained from L. plantarum species isolated from different ecological niches is also worth noting, thus proving it to be one of the most important and versatile species among LAB.

Effect of acetic acid present in bagasse hydrolysate on the activities of xylose reductase and xylitol dehydrogenase in Candida guilliermondii

The first two steps in xylose metabolism are catalyzed by NAD(P)H-dependent xylose reductase (XR) (EC 1.1.1.21) and NAD(P)-dependent xylitol dehydrogenase (XDH) (EC 1.1.1.9), which lead to xylose→xylitol→xylulose conversion. Xylitol has high commercial value, due to its sweetening and anticariogenic properties, as well as several clinical applications. The acid hydrolysis of sugarcane bagasse allows the separation of a xylose-rich hemicellulosic fraction that can be used as a substrate for Candida guilliermondii to produce xylitol. However, the hydrolysate contains acetic acid, an inhibitor of microbial metabolism. In this study, the effect of acetic acid on the activities of XR and XDH and on xylitol formation by C. guilliermondii were studied. For this purpose, fermentations were carried out in bagasse hydrolysate and in synthetic medium. The activities of XR and XDH were higher in the medium containing acetic acid than in control medium. Moreover, none of the fermentative parameters were significantly altered during cell culture. It was concluded that acetic acid does not interfere with xylitol formation since the increase in XR activity is proportional to XDH activity, leading to a greater production of xylitol and its subsequent conversion to xylulose.

Protein and glucose 6-phosphate dehydrogenase releasing from baker's yeast cells disrupted by a vertical bead mill.

Abstract Saccharomyces cerevisiae cells were disrupted at 5°C in a 200-ml vertical bead mill using glass beads (diameter: 0.5 mm) as abrasive. The parameters varied were: agitation (1100; 1700; 2300 and 3100 rpm), glass beads-volume (50, 75 and 100 ml) and cell concentration (170, 250 and 325 g/l). Before disruption the cells were thoroughly rinsed in physiological buffer and suspended in 50 mM Tris–HCl (pH 7.5) containing 2 mM MgCl 2 , 2 mM aminocaproic acid, 2 mM dithiothreitol and 1 mM phenylmethylsulfonyl fluoride. After removing the cell debris (centrifugation at 10 000× g /15 min), the total protein concentration and the activity of glucose 6-phosphate dehydrogenase (G6PDH) were measured. The high cell disruption rate (6×10 11 cells/min) occurred 3 min after the beginning of the procedure, carried out at 3100 rpm with 325 g/l of cells and 100 ml of glass beads. Moreover, a significant release of protein and G6PDH required 6 and 3 min of agitation, respectively. Thus G6PDH liberation and the cell disruption are coupled events. Furthermore, this disruption procedure was highly reproductive.

Separation of inulinase from Kluyveromyces marxianus using reversed micellar extraction

Inulinase from K. marxianus was extracted into a reversed micelle phase of the cationic surfactant BDBAC (n-benzyl-n-dodecyl-n-bis(2-hydroxyethyl)ammonium chloride) in isooctane/hexanol. The extractions carried out with cells (5.9 g/l) presented a recovery yield of 87% and a purification factor 2.8. Similar values were found for inulinase recovered from the clarified medium (91% recovery yield and 2.8 purification factor). For scaled-up (400-fold) extractions, the recovery of the initial activity reached 77% and the enrichment factor was 2.8.