J.E. Lozano

Determination of enzymatic activities of commercial pectinases for the clarification of apple juice

Abstract Different methods for testing Polygalacturonase (PG), pectinesterase (PE), and pectinlyase (PL) activities were applied to Rohapect D5S (RHD5) and Pectinol (PA1) commercial enzyme preparations in an apple pectin substrate. The viscometric method for PG activity determination was satisfactory, but foreign proteins could affect the spectrophotometric determination of PL activity in solutions of enzyme preparations. Although 50°C was a well-defined breaking point where enzymes rapidly decrease their activity, rate and range of heat-inactivation were different depending on the activity assayed. While PG activity showed two periods of different thermolability, PL was monophasic and highly sensitive to heat. The pH dependence of the pectic enzyme activities was also studied over the 3–7 range.

Influence of the extraction–purification conditions on final properties of alginates obtained from brown algae (Macrocystis pyrifera)

In this work, three methods (ethanol, HCl, and CaCl(2) routes) of sodium alginate extraction-purification from brown seaweeds (Macrocystis pyrifera) were used in order to study the influence of process conditions on final properties of the polymer. In the CaCl(2) route, was found that the precipitation step in presence of calcium ions followed by proton-exchange in acid medium clearly gives alginates with the lowest molecular weight and poor mechanical properties. It is well known that the acid treatment degrade the ether bonds on the polymeric chain. Ethanol route displayed the best performance, where the highest yield and rheological properties were attained with the lowest number of steps. Although the polymer I.1 showed a molar mass and polydispersity index (M(w)/M(n)) similar to those of commercial sample, its mechanical properties were lower. This performance is related to the higher content of guluronic acid in the commercial alginate, which promotes a more successful calcium chelation. Moreover, the employment of pH 4 in the acid pre-treatment improved the yield of the ethanol route, avoiding the ether linkage hydrolysis. Therefore, samples I.2 and I.3 displayed a higher M(w) and a narrower distribution of molecular weights than commercial sample, which gave a higher viscosity and better viscoelastic properties.

The effect of hydrocolloids on the stability and viscosity of cloudy apple juices

Abstract The influence of xanthan gum (XG) and carboxymethylcellulose (CMC) on the colloidal stability of cloudy apple juice has been studied. Concentrations from 0.4–0.5% completely inhibited juice clarification. At lower gum concentrations, juices with CMC were more stable. Z -potential ( ζ ) was measured by laser doppler electrophoresis. As expected, cloudiness in apple juice increased its stability with | ζ | (the more electro-negative, the more stable) following a behavior common to both food gums. Therefore, stability of cloudy juices with food gums could be predicted only in terms of ζ . Log–log plot of apparent viscosity ( η a ) vs ( γ ), showed a typical shear thinning behavior, with a transition from dilute to concentrated solution at some critical concentration. At low shear rates, XG was more viscous than CMC, demonstrating that the greater stabilizing effect of CMC was basically due to its electro-negativity.

Role of hydrophobicity in adhesion of wild yeast isolated from the ultrafiltration membranes of an apple juice processing plant

The role of cell surface hydrophobicity in the adhesion to stainless steel (SS) of 11 wild yeast strains isolated from the ultrafiltration membranes of an apple juice processing plant was investigated. The isolated yeasts belonged to four species: Candida krusei (5 isolates), Candida tropicalis (2 isolates), Kluyveromyces marxianus (3 isolates) and Rhodotorula mucilaginosa (1 isolate). Surface hydrophobicity was measured by the microbial adhesion to solvents method. Yeast cells and surfaces were incubated in apple juice and temporal measurements of the numbers of adherent cells were made. Ten isolates showed moderate to high hydrophobicity and 1 strain was hydrophilic. The hydrophobicity expressed by the yeast surfaces correlated positively with the rate of adhesion of each strain. These results indicated that cell surface hydrophobicity governs the initial attachment of the studied yeast strains to SS surfaces common to apple juice processing plants.

Fractal dimension and mechanism of aggregation of apple juice particles.

Turbidity of freshly squeezed apple juice is produced by a polydisperse suspension of particles coming from the cellular tissue. After precipitation of coarse particles by gravity, only fine-colloidal particles remain in suspension. Aggregation of colloidal particles leads to the formation of fractal structures. The fractal dimension is a measure of the internal density of these aggregates and depends on their mechanism of aggregation. Digitized images of primary particles and aggregates of depectinized, diafiltered cloudy apple juice were obtained by scanning electron microscopy (SEM). Average radius of the primary particles was found to be a = 40±11 nm. Maximum radius of the aggregates, RL, ranged between 250 and 7750 nm. Fractal dimension of the aggregates was determined by analyzing SEM images with the variogram method, obtaining an average value of Df = 2.3±0.1. This value is typical of aggregates formed by rapid flocculation or diffusion limited aggregation. Diafiltration process was found to reduce the average size and polydispersity of the aggregates, determined by photon correlation spectroscopy. Average gyration radius of the aggregates before juice diafiltration was found to be Rg = 629 ± 87 nm. Average number of primary particles per aggregate was calculated to be N = 1174.

Dual-species relations between Candida tropicalis isolated from apple juice ultrafiltration membranes, with Escherichia coli O157:H7 and Salmonella sp.

The objective of this study was to determine the interactions between common spoilage yeast, Candida tropicalis, isolated from ultrafiltration membranes, and Escherichia coli O157:H7 and Salmonella sp. on stainless steel surfaces.

Candida krusei isolated from fruit juices ultrafiltration membranes promotes colonization of Escherichia coli O157:H7 and Salmonella enterica on stainless steel surfaces

To clarify the interactions between a common food spoilage yeast and two pathogenic bacteria involved in outbreaks associated with fruit juices, the present paper studies the effect of the interplay of Candida krusei, collected from UF membranes, with Escherichia coli O157:H7 and Salmonella enterica in the overall process of adhesion and colonization of abiotic surfaces. Two different cases were tested: a) co-adhesion by pathogenic bacteria and yeasts, and b) incorporation of bacteria to pre-adhered C. krusei cells. Cultures were made on stainless steel at 25°C using apple juice as culture medium. After 24 h of co-adhesion with C. krusei, both E. coli O157:H7 and S. enterica increased their counts 1.05 and 1.11 log CFU cm2, respectively. Similar increases were obtained when incorporating bacteria to pre-adhered cells of Candida. Nevertheless C. krusei counts decreased in both experimental conditions, in a) 0.40 log CFU cm2 and 0.55 log CFU cm2 when exposed to E. coli O157:H7 and S. enterica and in b) 0.18 and 0.68 log CFU cm2, respectively. This suggests that C. krusei, E. coli O157:H7, and S. enterica have a complex relationship involving physical and chemical interactions on food contact surfaces. This study supports the possibility that pathogen interactions with members of spoilage microbiota, such as C. krusei, might play an important role for the survival and dissemination of E. coli O157:H7 and Salmonella enterica in food-processing environments. Based on the data obtained from the present study, much more attention should be given to prevent the contamination of these pathogens in acidic drinks.

In situ rheology of yeast biofilms

The aim of the present work was to investigate the in situ rheological behavior of yeast biofilms growing on stainless steel under static and turbulent flow. The species used (Rhodototula mucilaginosa, Candida krusei, Candida kefyr and Candida tropicalis) were isolated from a clarified apple juice industry. The flow conditions impacted biofilm composition over time, with a predominance of C. krusei under static and turbulent flow. Likewise, structural variations occurred, with a tighter appearance under dynamic flow. Under turbulent flow there was an increase of 112 μm in biofilm thickness at 11 weeks (p < 0.001) and cell morphology was governed by hyphal structures and rounded cells. Using the in situ growth method introduced here, yeast biofilms were determined to be viscoelastic materials with a predominantly solid-like behavior, and neither this nor the G’0 values were significantly affected by the flow conditions or the growth time, and at large deformations their weak structure collapsed beyond a critical strain of about 1.5–5%. The present work could represent a starting point for developing in situ measurements of yeast rheology and contribute to a thin body of knowledge about fungal biofilm formation.

Pectin extraction from quince (Cydonia oblonga) pomace applying alternative methods: Effect of process variables and preliminary optimization

The objectives of this study were to introduce alternative methods in the process of pectin extraction from quince pomace, to determine the effect of selected process variables (factors) on the obtained pectin, and to perform a preliminary optimization of the process. A fractional factorial experimental design was applied, where the factors considered were six: quince pomace pretreatment (washing vs blanching), drying method (hot air vs LPSSD), acid extraction condictions (pH, temperature, and time), and pectin extract concentration method (vacuum evaporation vs ultrafiltration). The effects of these factors and their interactions on pectin yield (Y: 0.2–34.2 mg/g), GalA content (44.5–76.2%), and DM (47.5–90.9%), were determined. For these three responses, extraction pH was the main effect, but it was involved in two and three factors interactions. Regarding alternative methods, LPSSD was required for maximum Y and GalA, and ultrafiltration for maximum GalA and DM. Response models were used to predict optimum process conditions (quince blanching, pomace drying by LPSSD, acid extraction at pH 2.20, 80 ℃, 3 h, and concentration under vacuum) to simultaneously maximize Y (25.2 mg/g), GalA (66.3%), and DM (66.4%).

Disinfection efficacy over yeast biofilms of juice processing industries

Membrane separation systems represent a hot - spot for biofilm formation in juice industries. Sodium hypochlorite (NaOCl) has been traditionally the disinfectant of choice; however, its effectiveness over well-established biofilms is limited. In this work the study of biofilm formation on ultrafiltration membranes was proposed. The effectiveness of cleaning and disinfection procedures commonly used in juice industry was tested on the removal and killing of cells. The species used (Rhodotorula mucilaginosa, Candida krusei, Candida kefyr and Candida tropicalis) were isolated from ultrafiltration modules of a clarified apple juice industry. Industrial concentrations of NaOCl (200mgCL∙L-1) showed to be effective against planktonic cultures with >4 log reductions, whereas their overall efficiency against adhered cells was smaller. Recovery of viable cell counts to initial numbers was evidenced regardless of the time of colonization. The topography of the surface showed to have an impact on the efficiency of the disinfectant, presenting membranes smaller log reductions than stainless steel (~1.09-1.53logCFU). At 200mgCl∙L-1 only membranes cross flow recovery was reached with no long-term effect over the attached cells. The overall results demonstrated the recalcitrance of these biofilms to typical cleaning and disinfection process which may confer them with a selective advantage.