Inactivation of Alicyclobacillus acidoterrestris spores, single or composite Escherichia coli and native microbiota in isotonic fruit-flavoured sports drinks processed by UV-C light.

Pasteurized sports drinks and other fruit-based beverages are susceptible to deterioration due to thermal processing ineffectiveness to inactivate certain spoilage microorganisms, like Alicyclobacillus acidoterrestris. This represents a major challenge for the beverage industry. The goals of this study were to: i) investigate the UV-C inactivation (annular thin film unit, actinometrical delivered fluence: 795-1270 mJ/cm2, 10-15 min, 20 °C, 1.8 L/h, Reh = 391-1067, recirculation mode operation) and the evolution during refrigerated storage of A. acidoterrestris ATCC 49025 spores and single or composite Escherichia coli ATCC 25922 in isotonic sports drinks (ISDs) made from orange (orange-ISD, UVT% = 81) or orange-banana-mango-kiwi-strawberry-lemon juices (multi-fruit-ISD, UVT% = 91), compared to a turbid orange-tangerine juice (OT juice, UVT% = 40); ii) assess the effect of pH, °Brix, A254nm, turbidity, colour and particle size of the ISDs and juice on microbial inactivation, iii) evaluate the evolution of native microbiota during cold storage, iv) investigate the Coroller, biphasic, Weibull, and Weibull-plus-tail models' ability to describe microbial inactivation and v) measure 5-hydroxymethylfurfural (HMF) formation. The modified biodosimetry method was used to calculate the germicidal UV-C fluences. Heat pasteurization (T-coil, 80 °C/6 min) was evaluated as the control treatment. UV-C was highly effective at inactivating E. coli as 4.1-5.1 and 4.5-5.6 log reductions were determined in the multi-fruit-ISD and orange-ISD, respectively, barely impacted by the background microbiota. No significant differences were recorded for the inactivation of E. coli in the UV-C and T-coil systems. Whereas, a significantly higher inactivation of A. acidoterrestris spores was achieved by UV-C (3.7-4.0 log reductions), compared to the negligible one achieved by the thermal treatment. Even though E. coli inactivation curves were similar in shape, UV-C was less effective when a cocktail of other E. coli strains was present. In comparison to the OT juice, the ISDs' inactivation kinetics were markedly different in shape, with a rapid decrease in population during the first minutes of treatment. The germicidal fluence (Hd biod) corresponding to A. acidoterrestris (19.1 mJ/cm2) was selected as it was higher than the one obtained for E. coli (11.0 mJ/cm2). UV-C induced 2.8- or 1.3 and 2.3- or 0.8 log-reductions of total aerobes or moulds and yeasts in the multi-fruit-ISD and orange-ISD, respectively. Compared to the other models, the Coroller and biphasic models showed a better fit and more accurate parameter estimates. UV-C-induced HMF production was not significant in the ISDs. The current study found that the UV-C treatment was more effective than typical heat pasteurization for inactivating A. acidoterrestris spores in isotonic drinks, following a similar trend for E. coli and native microbiota.

Active food additive based on encapsulated yerba mate (Ilex paraguariensis) extract: effect of drying methods on the oxidative stability of a real food matrix (mayonnaise).

The drying process used to obtain active food additives is critical to ensure its functionality. In this study, freeze- and spray-drying techniques were evaluated for encapsulation of extracts with antioxidant activity from yerba mate (Ilex paraguariensis), using maltodextrin (MD) as wall material. Additionally, the oxidative stability in a real food matrix (mayonnaise) was assessed. Both MD addition and drying methods affected the physical properties [moisture content, water activity (aW)] and oxidative stability. MD addition diminished moisture content and prevented polyphenol compounds from degradation. The spray-dried powders displayed the lowest moisture content (1.6 ± 0.3% bs), the highest polyphenol content (135.4 mg GAE/g pure extract), and oxidative stability than the freeze-dried samples. The antioxidant capacity of the encapsulated powder subjected to spray-drying increased the oxidative stability of the mayonnaise (258 ± 32 min) more than the other assayed system (165 ± 5 min). Therefore, a natural spray-dried antioxidant food additive was obtained with potential use in the food industry.

Hurdle processing of turbid fruit juices involving encapsulated citral and vanillin addition and UV-C treatment.

The aim of this study was to evaluate a hurdle strategy for orange-tangerine (OT) and orange-banana-mango-kiwi-strawberry (OBMKS) juices processing based on UV-C treatment assisted or not by mild heat and the addition of natural antimicrobials. Vanillin and citral emulsions were successfully encapsulated using maltodextrin and HI-CAP (5,18,3) and characterized. The susceptibility of Lactobacillus plantarum ATCC 8014, Escherichia coli ATCC 25922, and Saccharomyces cerevisiae KE 162 to binary mixtures of the encapsulated agents was examined in culture media according to the Berenbaum experimental design. The boundary between growth and non-growth as a function of vanillin and citral concentrations was predicted by means of the probabilistic model using logistic regression. Microbial inactivation achieved by pilot-scale UV-C light (0-390 mJ/cm2) on its own, assisted by mild heat (50 °C, UV-C/H) and combined with antimicrobials (1000 ppm vanillin plus 100 ppm citral) addition (UV-C + A/UV-C/H + A) was assessed in OT and OBMKS. Yeast induced damage in a model solution treated by UV-C + A was studied by flow cytometry (FC). All the antimicrobial mixtures resulted in additive effects (FICindex = 1), thus offering through the probabilistic models a range of formulation possibilities with antimicrobial capacity encompassing lower vanillin and citral concentrations compared to those required when used alone (Vrange = 0-1875 ppm plus Crange = 392-0 ppm). UV-C led up to 3.7-3.8, 2.4-3.6 and 1.5-1.6 log-reductions of E. coli, L. plantarum and S. cerevisiae in OT and OBMKS, respectively. A significant increase of 1.7-2.2, 2.1-2.7 and 4.1-5.3 log cycles in microbial inactivation was observed after UV-C/H treatment. Additional inactivation of 0.7-3.1 and 0.5-2.7 log reductions were observed for E. coli and S. cerevisiae, respectively, when UV-C + A and UV-C/H + A were applied in both juices. Therefore, the addition of antimicrobials to the UV-C treated juices, showed additive to synergistic effects on E. coli and S. cerevisiae, respectively along refrigerated storage. A shift from yeast cells with intact membrane and esterase activity in control samples to cells with permeabilized membrane in C + A, UV-C and UV-C + A samples were determined by FC. The shift was more noticeable in UV-C + A samples. Sublethally damaged cells were only detected in C + A and UV-C samples. This study demonstrates that combining a pilot-scale UV-C treatment with the addition of chosen binary mixtures of vanillin and citral, can ensure more than 5 log-reductions of E. coli, L. plantarum and S. cerevisiae in OT and OBKMS juice blends.

Effect of pilot-scale UV-C light treatment assisted by mild heat on E. coli, L. plantarum and S. cerevisiae inactivation in clear and turbid fruit juices. Storage study of surviving populations.

Consumer growing demands for high-quality and safe food and beverages have stimulated the interest in alternative preservation technologies. Short-wavelength ultraviolet light (UV-C, 254 nm) has proven to be useful for the decontamination of a great variety of clear juices while improving their quality compared to traditional thermal treatments. Suspended solids and coloured compounds in turbid juices, diminish light transmission. The use of UV-C under a hurdle approach, may be a promising strategy for their treatment. The purpose of this study was to analyse Escherichia coli ATCC 25922, Saccharomyces cerevisiae KE 162 and Lactobacillus plantarum ATCC 8014 inactivation in clear pear juice (PJ), turbid orange-tangerine (OT) and orange-banana-mango-kiwi-strawberry (OBMKS) juices processed by single UV-C (390 mJ/cm2, 20 °C) and UV-C assisted by mild heat (UV-C/H, 50 °C) at pilot-scale in a coiled tubing unit and stored under refrigeration (5 °C). Inactivation studies were also conducted in peptone water (PW) and model solution (MS). The adequacy of the Coroller, Weibull and Biphasic Plus Shoulder models was studied. UV-C was highly effective in PW, MS and PJ, achieving up to 5.5-6.3-4.7, 4.8-5.1-4.6 and 4.4-5.5 log reductions for L. plantarum, E. coli,and S. cerevisiae, respectively. Whereas, a moderate inactivation by single UV-C was recorded in the turbid blends, reducing up to 2.4-3.8-1.6 and 3.6-3.7-1.3 log-cycles in OT and OBMKS, respectively. When the UV-C/H treatment was applied, high bacterial inactivation was observed achieving 5.2-5.6, 6.3-6.6 and 5.5-6.7 log reductions in OT, OBMKS and PJ, respectively, while 4.6-4.9 log reductions were determined for the yeast in OBMKS and OT, respectively. Thus, additive inactivation effects between UV-C and H were observed. All the models tested gave useful information regarding the existence of microbial subpopulations with varying resistances. However, the cumulative Weibull distribution function was the most versatile one, fitting inactivation curves with different shapes. Additionally, the frequency distributions of resistances showed that UV-C/H not only increased the UV-C microbicidal effect but changed the distribution of inactivation times. Principal component analysis revealed that UV-C effectiveness was associated to low particle size, a⃰, turbidity and high UV-C transmittance. An increase on the inactivation of treated bacterial populations was recorded along storage, while no yeast recovery was observed, thus emphasizing the contribution of refrigerated storage to microbial inactivation. Microbial inactivation in clear and turbid juices achieved by UV-C (390 mJ/cm2) assisted by mild heat (50 °C) and subsequent refrigerated storage may represent an useful alternative for multiple applications in the juice industry.

Effectiveness of UV-C light assisted by mild heat on Saccharomyces cerevisiae KE 162 inactivation in carrot-orange juice blend studied by flow cytometry and transmission electron microscopy.

The aim of this study was to analyze the effectiveness of UV-C light (0-10.6 kJ/m2) assisted by mild heat treatment (50 °C) on the inactivation of Saccharomyces cerevisiae KE 162 in peptone water and fresh carrot-orange juice blend (pH: 3.8; 9.8°Brix; 707 NTU; absorption coefficient: 0.17 cm-1). Yeast induced damage by single UV-C and mild heat (H) and the combined treatment UV-C/H, was investigated by flow cytometry (FC) and transmission electron microscopy (TEM). When studying induced damage by FC, cells were labeled with fluorescein diacetate (FDA) and propidium iodide (PI) to monitor membrane integrity and esterase activity. UV-C/H provoked up to 4.7 log-reductions of S. cerevisiae; whereas, only 2.6-3.3 log-reductions were achieved by single UV-C and H treatments. FC revealed a shift with treatment time from cells with esterase activity and intact membrane to cells with permeabilized membrane. This shift was more noticeable in peptone water and UV-C/H treated juice. In the UV-C treated juice, double stained cells were detected, suggesting the possibility of being sub-lethally damaged, with compromised membrane but still metabolically active. TEM images of treated cells revealed severe damage, encompassing coagulated inner content, disorganized lumen and cell debris. FC and TEM provided additional information regarding degree and type of damage, complementing information revealed by the traditional plate count technique.

Inactivation of Alicyclobacillus acidoterrestris ATCC 49025 spores in apple juice by pulsed light: Influence of initial contamination and required reduction levels / Inactivación de esporas de Alicyclobacillus acidoterrestris ATCC 49025 por luz pulsada en jugo de manzana: Influencia de la contaminación inicial y de las reducciones requeridas

The purpose of this study was to analyze the response of different initial contamination levels of Alicydobadllus acidoterrestris ATCC 49025 spores in apple juice as affected by pulsed light treatment (PL, batch mode, xenon lamp, 3pulses/s, 0-71.6 J/cm²). Biphasic and Weibull frequency distribution models were used to characterize the relationship between inoculum size and treatment time with the reductions achieved after PL exposure. Additionally, a second order polynomial model was computed to relate required PL processing time to inoculum size and requested log reductions. PL treatment caused up to 3.0-3.5 log reductions, depending on the initial inoculum size. Inactivation curves corresponding to PL-treated samples were adequately characterized by both Weibull and biphasic models (R²d j 94-96%), and revealed that lower initial inoculum sizes were associated with higher inactivation rates. According to the polynomial model, the predicted time for PL treatment increased exponentially with inoculum size.

El objetivo del presente trabajo fue evaluar la influencia de la concentración de esporas de Alicyclobacillus acidoterrestris ATCC 49025 en la respuesta de inactivación por acción de la luz pulsada (modo estanco, lámpara de xenón, 3 pulsos/s, 0-71,6 J/cm²) en jugo de manzana comercial. Para caracterizar la relación existente entre la concentración de esporas y el tiempo de tratamiento con las reducciones logarítmicas alcanzadas luego de la exposición a la luz pulsada (LP), se aplicaron 2 modelos: el de Weibull y el bifásico. Adicionalmente, se estimó la relación entre el tiempo de tratamiento con LP y la concentración inicial de inoculo en el jugo con las reducciones logarítmicas logradas mediante regresión múltiple y la metodología de superficie de respuesta (MSR). La inactivación por LP provocó entre 3 y 3,5 reducciones logarítmicas, según la concentración inicial de esporas. Las curvas de inactivación fueron adecuadamente caracterizadas por los modelos matemáticos propuestos (Restado = 94-96%). El análisis por MSR permitió predecir un aumento exponencial del tiempo de tratamiento requerido conforme se incrementa el nivel de contaminación inicial.

Inactivation of Alicyclobacillus acidoterrestris ATCC 49025 spores in apple juice by pulsed light. Influence of initial contamination and required reduction levels.

The purpose of this study was to analyze the response of different initial contamination levels of Alicyclobacillus acidoterrestris ATCC 49025 spores in apple juice as affected by pulsed light treatment (PL, batch mode, xenon lamp, 3pulses/s, 0-71.6J/cm2). Biphasic and Weibull frequency distribution models were used to characterize the relationship between inoculum size and treatment time with the reductions achieved after PL exposure. Additionally, a second order polynomial model was computed to relate required PL processing time to inoculum size and requested log reductions. PL treatment caused up to 3.0-3.5 log reductions, depending on the initial inoculum size. Inactivation curves corresponding to PL-treated samples were adequately characterized by both Weibull and biphasic models (Radj2 94-96%), and revealed that lower initial inoculum sizes were associated with higher inactivation rates. According to the polynomial model, the predicted time for PL treatment increased exponentially with inoculum size.

Impact of a combined processing technology involving ultrasound and pulsed light on structural and physiological changes of Saccharomyces cerevisiae KE 162 in apple juice.

This study analyzed the effect of single ultrasound (US) (600 W, 20 kHz and 95.2 µm wave amplitude, 10 or 30 min at 20 or 44 ± 1 °C), or combined with pulsed light technology (PL) with controlled heat build-up (Xenon lamp, 3 pulses/s, 71.6 J/cm2, temperature ranges: 2-20 ± 1 °C and 44-56 ± 1 °C) on the inactivation of Saccharomyces cerevisiae KE 162 cells in commercial (pH: 3.5 ± 0.1; 12.5 ± 0.1 °Brix) and freshly pressed (pH: 3.4 ± 0.1; 12.6 ± 0.1 °Brix) apple juices. Structural damages were analyzed by transmission electronic microscopy (TEM) and induced damage by flow cytometry (FC). Cells were labeled with fluorescein diacetate (FDA) and propidium iodide (PI) for monitoring membrane integrity and esterase activity. US+PL treatment at the highest heat build-up led up to 6.4 and 5.8 log-cycles of yeast reduction in commercial and freshly apple juices, respectively. TEM images of treated cells revealed severe damage, encompassing loss and coagulated inner content and cell debris. In addition, FC revealed a shift of yeasts cells with esterase activity and intact membrane to cells with permeabilized membrane. This effect was more notorious after single 30-min US and all combined US+PL treatments, as 91.6-99.0% of treated cells showed compromised membrane. Additionally, heat build-up enhanced this shift when applying 10 min US (20 °C) in both juices.

Study of the inactivation of spoilage microorganisms in apple juice by pulsed light and ultrasound.

The aim of this study was to evaluate the effect of ultrasound (US) (600 W, 20 kHz and 95.2 µm wave amplitude; 10 or 30 min at 20, 30 or 44 ± 1 °C) and pulsed light (PL) (Xenon lamp; 3 pulses/s; 0.1 m distance; 2.4 J/cm(2)-71.6 J/cm(2); initial temperature 2, 30, 44 ± 1 °C) on the inactivation of Alicyclobacillus acidoterrestris ATCC 49025 spores and Saccharomyces cerevisiae KE162 inoculated in commercial (pH: 3.5; 12.5 °Brix) and natural squeezed (pH: 3.4; 11.8 °Brix) apple juices. Inactivation depended on treatment time, temperature, microorganism and matrix. Combination of these technologies led up to 3.0 log cycles of spore reduction in commercial apple juice and 2.0 log cycles in natural juice; while for S. cerevisiae, 6.4 and 5.8 log cycles of reduction were achieved in commercial and natural apple juices, respectively. In natural apple juice, the combination of US + 60 s PL at the highest temperature build-up (56 ± 1 °C) was the most effective treatment for both strains. In commercial apple juice, US did not contribute to further inactivation of spores, but significantly reduced yeast population. Certain combinations of US + PL kept on good microbial stability under refrigerated conditions for 15 days.