Modeling the effect of inoculum size and acid adaptation on growth/no growth interface of Escherichia coli O157:H7

The objective of this study was to model with logistic regression the growth/no growth interface of different initial inoculation levels (10¹, 10³ and 10⁵ CFU/ml; study 1), or nonadapted vs acid-adapted (study 2) Escherichia coli O157:H7 as influenced by pH, NaCl concentration and incubation temperature. Study 1 was conducted with a mixture of four E. coli O157:H7 strains grown (35 °C, 24 h) in tryptic soy broth (TSB). Study 2 was conducted with the same mixture of four E. coli O157:H7 strains grown (35 °C, 24 h) in glucose-free TSB with 1% added glucose (final pH 4.83), or in diluted lactic acid meat decontamination runoff fluids (washings; final pH 4.92), or nonadapted cultures prepared in glucose-free TSB (final pH 6.45), or in water washings (final pH 6.87). Parameters included incubation temperature (10–35 °C), pH (3.52–7.32), and NaCl concentration (0–10% w/v). Growth responses were evaluated for 60 days turbidimetrically (610 nm) every 5 days in 160 (study 1) and 360 (study 2) combinations in quadruplicate samples, with a microplate reader. The lower the initial inoculum the higher were the minimum pH and a_w values permitting growth. Differences in the pH and a_w growth limits among inoculum concentrations increased at 15 and 10 °C. Acid-adapted cultures were able to grow at lower pH than nonadapted cultures, while at temperatures below 25 °C, growth initiation of nonadapted cultures stopped at higher a_w compared to acid-adapted cultures for the whole pH range of 3.52 to 7.32. A comparison with available data indicated that our model for acid-adapted E. coli O157:H7 in different environments may provide representative growth probabilities covering both nonadapted and stress-adapted contaminants.     

Physiological properties of Lactobacillus paracasei, L. danicus and L. curvatus strains isolated from Estonian semi-hard cheese

Growth and survival of Lactobacillus paracasei (six strains), L. danicus sp. nov. (four isolates, two strains) and L. curvatus (two strains) from semi-hard Estonian cheeses were comparatively studied in different environmental conditions of relevance for their growth in cheese and survival in gastric environment. Maximum specific growth rates for L. paracasei strains varied between 0.40 and 0.57 h⁻¹, and all strains were tolerant to low water activities, heating at 60 °C for 30 min and pH 3. The newly discovered genetically distinct species L. danicus was characterized by low maximum growth rates (0.26–0.38 h⁻¹) and low temperature optimum (<30 °C). It was acid and heat sensitive and inhibited at salt concentrations from 4% and water activities below 0.93. L. curvatus was characterized by the highest growth rates (0.65–0.70 h⁻¹), tolerance to high NaCl concentrations, but sensitivity to heating, bile salts and low pH. The study showed that genetically different LAB species isolated from cheese could be distinguished by simple cultivation experiments.     

Prediction of the rheology parameters of ripened semi-hard cheeses using fluorescence spectra in the UV and visible ranges recorded at a young stage

Storage modulus (G′), loss modulus (G″), strain, tan (δ) and complex viscosity (η*) of 20 semi-hard cheeses were measured by dynamic oscillatory analysis after 2, 30 and 60 days of ripening. On the same cheeses and at the same ages, tryptophan and riboflavin fluorescence spectra were recorded. The aim was to predict the rheology parameters of ripened cheeses from spectra recorded on these cheeses at a young stage. Using partial least square, tryptophan fluorescence spectra recorded at 20 °C on 2-days-old cheeses predicted G′, G″, strain, tan (δ) and η* measured at 80 °C on the 60-days-old cheeses with correlation coefficients (R) of 0.98, 0.97, 0.98, 0.98 and 0.97, respectively. Riboflavin fluorescence spectra gave slightly lower correlation coefficients of 0.88, 0.88, 0.92, 0.87 and 0.88, respectively. Dependent only on visible light, the riboflavin fluorescence spectra potentially provide viable and economic prediction of the rheology of ripe cheese.     

Effect of temperature and growth media on the attachment of Listeria monocytogenes to stainless steel

Listeria monocytogenes ATCC 19111 cultivated in nutrient-rich medium (brain heart infusion, BHI) or starved in minimal medium (10% filter sterilized pond water and 90% sterilized distilled water) were investigated for their initial attachment to austenitic stainless steel No. 4 with satin finish at 4 °C, 20 °C, 30 °C, 37 °C, or 42 °C. A droplet (10 μl) containing  10⁷ CFU/ml of L. monocytogenes suspended in BHI or minimal medium was placed on the stainless steel surface. After holding in saturated humidity for 3 h at the desired temperature the surface was washed and prepared for scanning electron microscopy (SEM). Using SEM, attachment of L. monocytogenes was determined by counting cells remaining on the surface. When L. monocytogenes cultivated in BHI were used, with the exception of the number of attached cells being lower at 42 °C than at 37 °C and 30 °C, the number of attached cells increased with increasing temperature (P < 0.05). When L. monocytogenes starved in minimal medium were used, the number of attached cells also increased with increasing attachment temperature (P < 0.05), but the number of attached cells at 42 °C was lower than that at the other temperatures. The attachment of L. monocytogenes to stainless steel surface was greater when cultivated in rich medium of BHI vs starved in the minimal medium.     

Screening of lactic acid bacteria isolates from dairy and cereal products for exopolysaccharide production and genes involved

Scanning electron microscopy (SEM) studies revealed that exposure to 4lethal alkaline stress induced statistically significant (P < 0.05) changes in mean cell length, radius and volume in Listeria monocytogenes and a derived σ^B deficient mutant. Bacterial morphology was altered at pH values above 9.0, to include single filamentous or elongated chain forms. Such filamentation and chain formation was observed in the parent strain and in the σ^B deficient strain, and in buffered and non-buffered media. Giemsa staining revealed that the filaments were multi-nucleate, with nucleoids spaced along the length of the atypical cells. In buffered media, longer alkaline exposure was associated with increases in the frequency and length of filamentation. In non-buffered medium, longer exposure was associated with gradual decline in length and the frequency of observation of filaments. Transfer of alkaline treated cells to neutral conditions was associated with the formation of septa within filaments, cell division, and a rapid return to normal morphology, i.e. within 3 h. The observed effects, and their reversibility, may be important in increasing the alkaline tolerance of this pathogen during phagocytosis within the innate human immune system response, and in adaptation/survival in food environments treated with alkali detergents and/or sanitisers. Such atypical cells may be associated with increased survival of L. monocytogenes in adverse environments and may also contribute to qualitative and quantitative underestimation of this important pathogen in food processing environments, with potential implications in public health.     

Bacterial cells exposed to nanosecond pulsed electric fields show lethal and sublethal effects

Cell suspensions of Escherichia coli K12 and Salmonella typhimurium were exposed to electrical pulses of 32 ns duration at a field intensity of 100 kV/cm and a repetition rate of 30 pulses per second for a total of 300 s. Treated cells were plated onto Tryptone Soya Agar (TSA) and TSA supplemented with NaCl, and cell counts were monitored daily for 3 days. The concentrations of NaCl used were 3 and 4% (w/v) for E. coli and 4 and 5% (w/v) for S. typhimurium. Treatment under these conditions resulted in a 2 log₁₀ reduction for E. coli and approximately a single log₁₀ reduction for S. typhimurium. For both species of bacteria it was discovered that the surviving population was composed of only 1% of uninjured cells. Moreover, the proportion of sublethally injured cells increased more rapidly than the total recoverable population suggesting a process of injury accumulation culminating in death rather than an ‘all or nothing’ mechanism. Sublethal injury manifested itself in a proportion of the injured population of both species by an extended lag phase at longer treatment times. Finally, possible mechanisms by which nanosecond electric pulses inactivate bacteria are discussed.     

Effect of season on color of Hanwoo (Korean native cattle) beef

A total of 1278 head of Hanwoo (Korean native cattle) slaughtered over four seasons were used to evaluate the effect of season on color characteristics of beef longissimus dorsi (LD) muscle. CIE L^*, a^*, b^*, C^* values and hue angle were significantly lower (P<0.05) in cattle slaughtered in the winter season. Meat color was darker in the winter than in the spring and autumn seasons. The L^* values among three average daily temperature (T_a) categories were different (P<0.05) in order of: [5 °CT_a<25 °C] > [T_a25 °C] > [T_a<5 °C], indicating that the meat color of cattle slaughtered at T_a<5 °C was darker. The a^*, b^*, C^* values and hue angle were significantly lower (P<0.05) in cattle slaughtered at T_a<5 °C. Season at slaughter is of great importance for meat color. Namely, meat color of Hanwoo beef was influenced by environmental temperature. Overall, cattle slaughtered in the winter season of T_a<5 °C produced beef with more undesirable meat color properties.     

Modelling the effect of a temperature shift on the lag phase duration of Listeria monocytogenes

The aim of this work is to study and model the effect of a temperature shift on h₀, the product of the growth rate by the lag phase duration (μλ). Our work is based on the data of Whiting and Bagi [Int. J. Food Microbiol. 73 (2002) 291], who studied the influence of both the pre-incubation temperature (T_prior) and the growth temperature (T_growth) on λ values of Listeria monocytogenes. We introduce a new model to describe the evolution of the parameter h₀ as a function of T_prior and T_growth, and compare it to Whiting and Bagi's published polynomial model that describes the influence of T_prior and T_growth on λ independently of μ. For exponential as well as stationary phase cells, h₀ increases almost linearly with the magnitude of the temperature shift. A simple linear model of h₀ turns out to be more suitable to predict λ values than a polynomial model of λ.     

Extension of the shelf life of beef steaks packaged in a modified atmosphere by treatment with rosemary and displayed under UV-free lighting

Fresh beef steaks, either sprayed on the surface with a solution of rosemary and vitamin C or not sprayed, were packaged in 70%O₂₊20%CO₂₊10%N₂ and displayed at 1±1 °C without illumination or illuminated by a standard fluorescent lamp, a low-UV, colour-balanced lamp (Promolux®), or the fluorescent lamp with a UV filter. Metmyoglobin formation, lipid oxidation (2-thiobarbituric acid reactive substances), instrumental colour (CIE L*, a*, b*), psychrotrophic bacterial counts (PCA) and sensory discolouration and off-odour were determined. Results showed that the use of the antioxidant mixture of rosemary and vitamin C together with the absence of UV radiation significantly reduced the rates of metmyoglobin formation and lipid oxidation, as well as microbial growth, and extended the display life from about 10 to about 20 days.     

Effect of temperature and relative humidity on yellowing rate of paddy

The effects of temperature and relative humidity (or water activity) in storage chambers on yellowing rate of paddy were investigated and then an empirical equation for predicting the yellowing rate was developed. Paddy was conditioned using saturated salt solutions at relative humidities ranging from 0.80 to 0.95 and temperatures of 35, 45, 55, 60 and 65°C. The yellowing rate was found to follow the zero order kinetics. The yellowing constant value (k) increased exponentially with temperature and increased linearly with water activity. The magnitude of apparent activation energy varied from 130–145 kJ/mol. A predictive equation for determining yellowing rate was ln k=−δa_w−/T+(γa_w)/T where a_w was water activity (valid from 0.80 to 0.95), T was absolute temperature (valid from 308 to 338 K) and , δ, and γ were constants. The results of variance analysis showed that temperature, water activity and their interaction significantly influenced the yellowing rate of paddy.     

A modified Weibull model for bacterial inactivation

In this paper, a modified Weibull model is proposed to fit microbial survival curves. This model can incorporate shoulder and/or tailing phenomena if they are encountered. We aim to obtain an accurate fit of the “primary” modelling of the bacterial inactivation and to provide a useful and meaningful model for biologists and food industry. A δ parameter close to the classical concept of the D value, established for sterilisation processes, is used in the model. The specific parameterisation of the Weibull model is evaluated for the parameter of interest δ. The goodness-of-fit of the model is compared to the one produced by the model proposed by Geeraerd et al., [Geeraerd, A.H., Herremans, C.H., Van Impe, J.F., 2000. Structural model requirements to describe microbial inactivation during a mild heat treatment. Int. J. Food Microbiol. 59, 185-209.] on experimental data. As our model provides good fits for the different types of survival curves analysed, further research can focus on the development of suitable secondary model types. In this respect, it is interesting to note that the δ parameter is close to the D concept.     

Effect of water activity on inactivation of Listeria monocytogenes and lactate dehydrogenase during high pressure processing

The aim of this study was to investigate the effect of water activity (a_w) on the inactivation of Listeria monocytogenes and lactate dehydrogenase (LDH) during high pressure processing (HPP). For microbial inactivation lyophilized cells of L. monocytogenes 19,115 were left dry or were suspended in 10 ml of 0.1% peptone water, 10 ml of glycerol, or mixtures of glycerol and peptone water. All samples of various a_ws were high pressure (HP) processed at ambient temperature at 600 MPa for 300 s. Following HPP, samples were serially diluted in 0.1% peptone and spread-plated on Tryptic Soy agar supplemented with Yeast Extract. For enzyme inactivation, 4.2 mg of lyophilized LDH was suspended in 2 ml of 100 mM phosphate buffer (pH 7.4), 2 ml of peptone water or glycerol, or in 2 ml mixtures of glycerol and peptone water. A lyophilized sample with no added liquid was also included. All enzyme samples were subjected to HPP as described above. After HPP, LDH was diluted to 0.28 μg/ml in 100 mM phosphate buffer (pH 7.4). LDH activity was assessed by measuring the change in concentration of β-NADH as a function of time. Dynamic light scattering analysis (DLS) was performed to examine the size distribution, polydispersity, and hydrodynamic radius of LDH before and after HPP. No significant difference in CFU/g was observed between lyophilized cells not subjected to HPP and lyophilized cells subjected to 600 MPa for 300 s (P < 0.05). However, lyophilized cells that were suspended in 100% to 60% peptone water showed a ~ 7.5-log₁₀ reduction when subjected to HPP. Survival of L. monocytogenes following HPP significantly increased (P < 0.05) when the peptone water concentration was decreased below 60% (a_w ~ 0.8). DLS results revealed that LDH suspended in buffer underwent aggregation following HPP (600 MPa, 300 s). Inactivation rate constants obtained using a first-order kinetic model indicated that untreated and HP processed lyophilized LDH had similar activities. When LDH was subject to HPP in solutions containing glycerol, enzyme activity decreased as the water content increased (r² = 0.95). Lyophilization completely protected L. monocytogenes and LDH from inactivation by high pressure. Furthermore, enzyme activity and cell survival increased as water activity was decreased. We postulate low a_w results in protein stabilization, which prevents protein denaturation and cell death during HPP.     

A model of the specific growth rate inhibition by weak acids in yeasts based on energy requirements

Zygosaccharomyces bailii, a spoilage yeast, capable of metabolic activity in food environments with low pH, low a_w and in the presence of weak acid preservatives was chosen for a study on the effect of benzoic acid on growth parameters. In batch cultures, under controlled pH, this food preservative inhibited growth, decreasing the specific growth rate (μ) and the yield coefficient (Y_S) on glucose. Data obtained at pH 3.5, 4.0 and 4.5 showed that this inhibition was exclusively promoted by the undissociated form of the acid since the effect was independent of pH when the concentration of the acid was expressed in this form. Moreover, the relationship between the values for μ and Y_S, provided evidence that the specific consumption rate of glucose (q_S) was not affected by benzoic acid, indicating that the inhibition of growth should be completely explained by a decrease of Y_S. The outcome of parallel experiments performed in continuous culture was that the decrease of Y_S was due to an increase of the maintenance coefficient (m), defined as the fraction of q_S diverted from growth to cope with stress, represented in this case by the presence of the preservative. Based on these results a model was built, assuming that m increased hyperbolically with the concentration of benzoic acid, from zero in the absence of the acid up to q_S when growth was completely inhibited. The concentration of the acid, for which m=q_S/2, is a constant (K_W), and represents a measure of the tolerance for a preservative, in this case benzoic acid. The simple equation μ/μ₀=1+W/K_W predicts the value of μ for a concentration (W) of the preservative, requiring the knowledge of two parameters: the specific growth rate in the absence of the preservative (μ₀) and K_W. The equation fitted very well the data of the effect of benzoic acid on the specific growth rate of Z. bailii, having K_W=0.96 mM benzoic acid. The model was also validated with other spoilage yeasts grown in the presence of benzoic acid in microtiter plates in an automated spectrophotometer. The values obtained for K_W under these conditions confirm Z. bailii as the most tolerant (K_W=2.1 mM) followed by Pichia sp. (K_W=0.78 mM), Saccharomyces cerevisiae (K_W=0.53 mM) and Debaryomyces hansenii (K_W=0.11 mM).     

Effects of different heat treatments on the furosine content in fresh filled pasta

The effect of different heat treatments on the furosine content in fresh filled pasta was studied and the rate of furosine increase after treatment has been found to be influenced by the initial furosine content in the mixture and by the treatment design. A mathematical model to describe the experimental data has been developed. The reaction of furosine formation appears to follow a pseudo-zero kinetic order. The estimated value of activation the energy is roughly 111 KJ mole⁻¹ and the z value has been estimated around 22.9 °C. The “furosine increase” (If) parameter has been calculated from processing data in order to compare samples obtained with different time–temperature processing combinations. Calculated values were correlated with F₇₀¹⁰ values (pasteurising effect) and results indicate a good linear relationship between the furosine increase and F₇₀¹⁰ values (r²=0.963; P<0.01). The comparison of the pasteurisation curve with the furosine increase curve shows that an optimal process design for thermal processing of fresh filled pasta under our experimental conditions is obtained at temperatures between 95 and 99 °C for times from 6 to 2 min.     

Bioactivity of the essential oil extracted from Evodia rutaecarpa Hook f. et Thomas against the grain storage insects, Sitophilus zeamais Motsch. and Tribolium castaneum (Herbst)

The toxic, repellent and feeding deterrent activities of the essential oil extracted from Evodia rutaecarpa Hook f. et Thomas, were evaluated against Sitophilus zeamais adults and Tribolium castaneum larvae and adults. Contact toxicity assayed by topical application showed that S. zeamais adults were significantly more susceptible (LD₅₀=0.043 μg/mg body wt) to the essential oil than T. castaneum adults (LD₅₀=0.118 μg/mg body wt) and larvae (LD₅₀=0.093–0.126 μg/mg body wt). However, in the fumigation assays, S. zeamais (LC₅₀=41 μg/L air) was less susceptible to the essential oil than T. castaneum (LC₅₀=11.7 μg/L air). When compared with larvae of various ages, T. castaneum adults were more susceptible to the fumigant toxicity of the essential oil. Also, in the treated filter paper repellency test, the essential oil was more repellent to T. castaneum than to S. zeamais. A flour disk bioassay demonstrated that the essential oil of E. rutaecarpa had a weaker feeding deterrent action against T. castaneum adults than against T. castaneum larvae and S. zeamais adults. The reduction in growth rate of T. castaneum larvae and S. zeamais adults was mainly due to a behavioural (feeding deterrent) action rather than to post-ingestive toxicity of the oil.     

Maintaining muscles at a high post-mortem temperature induces PSE-like meat in turkey

The aim of the present study was to validate an experimental model which surely generates pale, soft, exudative (PSE) turkey meat. Immediately after exsanguination, Pectoralis major (n=15) were kept at various temperatures (4, 20 or 40 °C) for 6 h. All the muscles were then stored at 4 °C for 9 days. They had the same rate of pH fall. L^* values were higher in the 40 °C treatment muscles than in the two other treatment muscles between 1 and 9 h. Drip loss of the 40 °C treatment muscles was higher than in the two other treatment muscles. However, thawing and cook loss were not significantly different between treatments. Cooked meat from the 40 °C treatment muscle was tougher than the two other treatment muscles. Napole yield was lower for these muscles. Myofibrillar protein extractability was lower in the 40 °C treatment muscle whatever the considered time. We showed that the 40 °C treatment muscles were similar to PSE muscles.     

Modeling the combined effects of pH, temperature and ascorbic acid concentration on the heat resistance of Alicyclobacillus acidoterrestis

In this study, thermal inactivation parameters (D- and z-values) of Alicyclobacillus acidoterrestris spores in McIlvaine buffers at different pH, apple juice and apple nectar produced with and without ascorbic acid addition were determined. The effects of pH, temperature and ascorbic acid concentration on D-values of A. acidoterrestris spores were also investigated using response surface methodology. A second order polynomial equation was used to describe the relationship between pH, temperature, ascorbic acid concentration and the D-values of A. acidoterrestris spores. Temperature was the most important factor on D-values, and its effect was three times higher than those of pH. Although the statistically significant, heat resistance of A. acidoterrestris spores was not so influenced from the ascorbic acid within the concentration studied. D-values in apple juice and apple nectars were higher than those in buffers as heating medium at similar pH. The D-values ranged from 11.1 (90 °C) to 0.7 min (100 °C) in apple juice, 14.1 (90 °C) to 1.0 min (100 °C) in apple nectar produced with ascorbic acid addition, and 14.4 (90 °C) to 1.2 min (100 °C) in apple nectar produced without ascorbic acid addition. However, no significant difference in z-values was observed among spores in the juices and buffers at different pH, and it was between 8.2 and 9.2 °C. The results indicated that the spores of A. acidoterrestris may survive in fruit juices and nectars after pasteurization treatment commonly applied in the food industry.     

Influence of temperature and surfactant on Escherichia coli inactivation in aqueous suspensions treated by moderate pulsed electric fields

This research employed a conductometric technique to estimate the inactivation kinetics of Escherichia coli cells in aqueous suspensions (1 wt.%) during simultaneous pulsed electric fields (PEF) and thermal treatments. The electric field strength was E = 5 kV/cm, the effective PEF treatment time t_PEF was within 0–0.2 s, the pulse duration t_i was 10^− 3 s, the medium temperature was 30–50 °C, and the time of thermal treatment t_T was within 0–7000 s. The damage of E. coli was accompanied by cell size decrease and release of intracellular components. The synergy between PEF and thermal treatments on E. coli inactivation was clearly present. The non-ionic surfactant Triton X-100 additionally improved its inactivation. The characteristic damage time followed the Arrhenius law within the temperature range 30–50 °C with activation energies W = 94 ± 2 kJ mol^− 1 and W = 103 ± 5 kJ mol^− 1 with and without the presence of surfactant, respectively. Relations between cell aggregation, cell ζ-potentials and presence of surfactant were analysed.     

Behaviour of Listeria monocytogenes in raw sausages (merguez) in presence of a bacteriocin-producing lactococcal strain as a protective culture

The effectiveness of a bacteriocin produced by Lactococcus lactis subsp. lactis M in reducing population level and growth of Listeria monocytogenes ATCC 7644 in fermented merguez sausage was examined. Two different formulas (with or without added nitrites) were assayed and predetermined numbers of Listeria (ca 10⁶ cfu g⁻¹) were added to sausage mixture. The effect of in situ production of the bacteriocin by Lactococcus lactis M on Listeria monocytogenes ATCC 7644 during fermentation and storage of merguez sausages at room (ca 22 °C) or at refrigeration (ca 7 °C) temperature was tested. Results indicated that counts of Listeria monocytogenes were decreased during fermentation of merguez samples fermented with either the bacteriocin-producing Lactococcus lactis M (Bac⁺) or a nonbacteriocin-producing Lactococcus lactis J (Bac⁻). However, reduction in Listeria cfu's was greater in samples fermented with the Bac⁺ than in those fermented with the Bac⁻ starter. In merguez sausage made without nitrites addition, the Bac⁺ starter induced further decrease in Listeria counts by 1.5 log cycles compared with that induced by the Bac⁻ starter. While in merguez samples with added nitrites (0.4%), the effect of the bacteriocin produced in situ was less important than in those made without nitrites addition.     

Temperature and water activity effects on growth and temporal deoxynivalenol production by two Argentinean strains of Fusarium graminearum on irradiated wheat grain

The ability of six strains of Pichia anomala, four strains of Pichia kluyveri and two strains of Hanseniaspora uvarum predominant during coffee processing to produce polygalacturonase (PG), pectin esterase (PE) and pectin lyase (PL) in yeast polygalacturonic acid medium (YPA) and in coffee broth (CB) was studied. For comparison, a reference strain of Kluyveromyces marxianus CCT 3172 isolated from cocoa and reported to produce high amount of PG was included.

Initial screening of PG activity using YPA medium showed that K. marxianus CCT 3172, P. anomala S16 and P. kluyveri S13Y4 had the strongest activity. Enzymatic assays showed that the four yeast species secreted PG, but none of the yeasts investigated was found to produce PE or PL. P. anomala S16 and P. kluyveri S13Y4 were found to produce higher amounts of PG when grown in CB than in YPA. When K. marxianus CCT 3172, P. anomala S16 and P. kluyveri S13Y4 were grown in YPA broth adjusted to pH of 3.0–8.0 and incubated at temperatures of 15–40 °C, the three yeast species secreted the highest amount of PG at pH 6.0 and at 30 °C. For PG secreted by K. marxianus CCT 3172 and P. anomala S16, the optimum pH and temperature for the enzymatic activity were 5.5 and 40 °C, respectively. On the other hand, PG produced by P. kluyveri S13Y4 showed the highest activity at pH 5.0 and 50 °C.

Significant differences in the extracellular activity of PG were found between the yeasts species as well as between strains within same species. High amounts of PG were produced by two strains of P. anomala and P. kluyveri. It is therefore likely that strains of those two species may be involved in the degradation of pectin during coffee fermentation.