JP6921838B2 - Method for liquid food preservation using pulsed electric field treatment - Google Patents

Description

The present invention relates to a method for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating. The present invention further relates to a method in which the liquid product is preheated before the liquid product is subjected to the method.

Pulsed electric field (PEF) is used as a technique for inducing electroporation of cell membranes by applying short-term pulses by a high-intensity external electric field. The most widely accepted theory of this phenomenon is that applying an external electric field to the biological membrane induces local instability in the lipid bilayer, ultimately leading to pore formation. Pore formation (electroporation) increases the permeability through the membrane (electroporation) and is a reversible process or when applied at high voltage, depending on the strength of the applied electric field. Is irreversible and results in cell death.

In a continuous flow PEF processing system, the duration of the pulse, the time between the two pulses (pause time), the residence and transit time of the fluid element in the high field region, and the time to leave the high field region before entering the cooling section. Various critical times, including, must be considered (Mastwijk et al., 2007). The total (effective) processing time is defined as the product of the number of pulses and the time per pulse received by the fluid element under high field conditions when pumped through the processing equipment. Currently, total processing time and electric field strength are known to be important factors in determining the efficiency of irreversible electroporation (Saulis and Wouters, 2007). Irreversible electroporation is effective against vegetative microorganisms with electric field strengths in the range of 10-20 kV / cm when a pulse with a duration of 2 microseconds is used for a total processing time of 100-400 microseconds (Figure). 1). Reynard and co-workers (1998) investigated the critical effect on pulse duration of a single pulse for gene transfer. They found that a minimum pulse time of about 1 ms was required for orientation and used a pulse with a field strength of 1-2.7 kV / cm and a duration of 24 ms for 3-5 ms. The critical response time for permeation is shown.

In contrast to direct current (DC) pulses, alternating current (AC) currents are used to create high field conditions in liquids. The AC current with a fixed frequency (f) can be seen as a pulse with a duration of 1 / f, but the characteristic pulse shape considered here is a rectangle, which is the repetition frequency of the pulse. Means that is less than the bandwidth (1 / pulse duration). U.S. Pat. No. 2010/0297313 considers AC currents at frequencies above 1 MHz (or pulse durations less than 1 microsecond).

The choice of specific treatment conditions is relevant to the various applications and purposes of electroporation. Reversible electroporation is a procedure frequently used in molecular biology and clinical biotechnology to introduce small or large molecules such as drugs, oligonucleotides, antibodies, and plasmids into cells, where cells The goal is to stay alive. Irreversible electroporation can be used to extract molecules from cells or to inactivate cells. In the present invention, we aim for irreversible electroporation as a non-thermal preservation method, where the maximum temperature and retention time obtained by PEF processing is lower than conventional heat pasteurization. This provides, for example, other beneficial aspects in better preservation of the fresh taste and nutritional value of the product.

When targeting microbial inactivation, the processing conditions selected for pulsed electrical treatment depend on several factors, but should be divided into three groups: processing parameters, microbial properties, and processing medium properties. Can be done.

In addition to field strength and treatment time, temperature is believed to be important for the effectiveness of microbial inactivation by PEF (Rasso et al., 2014). Increased field strength and processing time lead to increased PEF lethality. As a result of these conditions, more energy will be applied per unit of mass and the product will generate more heat. Typical process conditions used for irreversible electroporation are in the range of short pulses of microseconds at high voltage (5-80 kV / cm).

The degree of microbial inactivation by PEF is enhanced by increasing the temperature of the medium (eg, liquid food product) prior to PEF treatment, even in a temperature range that is not lethal to the microorganism. Although not bound by theory, this preheating effect affects the phospholipid bilayer structure of cell membranes, making cells more vulnerable to the PEF process (Wouters et al., 1999).

Microbial properties affect the effectiveness of microbial inactivation by PEF. In general, it has been reported that relatively large microorganisms are more sensitive to PEF than smaller microorganisms and Gram-negative bacteria are more sensitive to PEF than Gram-positive bacteria.

The effectiveness of PEF treatment is often studied in liquid media suspended in microorganisms. The properties of this treatment medium have been investigated and it has been reported that pH is very important for the effectiveness of the treatment. That is, PEF is much more effective in low pH media than in neutral pH media.

The commercial application of irreversible PEF processing is aimed at the inactivation of microorganisms in a continuous stream by passing through the processing apparatus once. As described in U.S. Pat. No. 2012/0103831, a circular loop that provides multiple passes through the processing apparatus by mixing the processed product with the unprocessed product complicates the process. It is avoided because of it.

As mentioned above, the application of an external pulse to the product introduces energy into the product, resulting in an increase in the temperature of the product. This temperature rise depends on the process conditions selected and the properties of the product (Heinz et al., 2002). A cooling section was placed between the two processing chambers in some applications to avoid overheating the product (Sharma et al., 2014). However, this approach requires more electrical energy and energy for cooling. To avoid too much heating, another possibility described is to introduce a pause after the application of the pulse or after the pulse train (El Zakhem et al., 2006). However, while the typical time for in-line thermal cold treatment ranges from seconds to minutes, this study (El Zakhem et al., 2007) increased the total treatment time from 5200 seconds to 7800 seconds. This is not possible in commercial applications. Pauses between pulses of at least 1 minute or between a series of pulses are also applied in the batch system described in Canadian Patent No. 27586878.

Commercially applicable treatment conditions are conditions in which PEF is transmitted at an electric field strength of 10-30 kV / cm. This is because the application of higher electric field strength has technical limitations and can cause dielectric breakdown of food materials.

The applied process conditions are suitable for liquid food products with low pH, i.e. highly acidic fruit juices with pH less than about 4.6. These process conditions appear to be suitable for inactivating larger sized microorganisms in liquid food products in some applications. In addition, Gram-negative bacteria can be inactivated more effectively than Gram-positive bacteria. In particular, inactivating small-sized Gram-positive bacteria is most often cumbersome under currently known PEF process conditions. Moreover, current low pH process conditions cannot be applied in a way that is effective for foods with pH higher than about 4.6.

Current PEF processes for liquid food products include electric field strengths that are relatively high, i.e. 5 kV / cm and above, typically 10-30 kV / cm. These relatively high field strengths generally hinder upscaling of PEF processing up to large volumes due to peak power requirements and limitations in the duration of a single pulse due to maximum stored pulse energy. These technical boundaries limit the maximum throughput of a single line for the storage of low conductive acidic fruit juices to 5000 L / h.

Therefore, the following PEF processing conditions are required.
-Preferably inactivate Gram-positive and / or relatively small-sized microorganisms without losing the efficiency of inactivating Gram-negative bacteria and / or inactivating relatively large-sized microorganisms. And / or-inactivates microorganisms and / or spores in liquid food products having a pH higher than about 4.6 and a pH lower than 4.6, and / or-existing Economically more profitable than inactivation technology and / or-applicable, using a single line to scale to a larger throughput volume than the current state-of-the-art conditions in use Has the potential to be up.

U.S. Pat. No. 2010/0297313 U.S. Pat. No. 2012/0103831 Canadian Patent No. 2758678

Mastwijk et al. , 2007 Saulis and Wouters, 2007 Reynard and collaborators (1998) Rasso et al. , 2014 Wouters et al. , 1999 Heinz et al. , 2002 Sharma et al. , 2014 El Zakhem et al. , 2006 El Zakhem et al. , 2007

The present invention is a method for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating to obtain a heated liquid product.
(A) Step of providing a liquid product;
(B) A step of providing an apparatus for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating;
(C) A step of supplying the liquid product to the inlet of the device and flowing the liquid product through the device;
(D) A step of continuously generating an electric current in the liquid product flowing through the apparatus, in which one minimum pulse is applied to each fluid element with a pulse duration of at least 10 microseconds during passage. And the electric field strength is 0.1 to 5 kV / cm;
Including
With respect to the method, the maximum temperature of the liquid product remains autonomously below 92 ° C. during said resistance heating.

It is part of the present invention that the pulse duration of a single pulse is a more important factor than the total effective processing time. The present inventors have found that even though the total effective processing time which is calculated as E ² _· tau were 4 times higher than that of the conventional PEF treatment 20 kV / cm to be used, 2 microseconds (tau) It was found that inactivation was not efficient at pulse duration and field strength (E) of 10 kV / cm. Under conditions of 0.1 to 5 kV / cm, inactivation was found to be effective only for pulse durations greater than 10 microseconds (eg, 100-1000 microseconds).

The methods of the present invention are applicable to liquid food products and liquid feed products, and the PEF processing conditions of the present invention are equally effective in inactivating Gram-negative and Gram-positive bacteria. PEF processing conditions are applicable to liquid food products and liquid feed products, which are effective in both inactivating relatively large microorganisms and inactivating relatively small microorganisms. Is the target. Furthermore, we have surprisingly found that PEF processing conditions are applicable under currently applied conditions at relatively low pH and conditions at higher pH. Finally, we have more than previously possible treatments using methods known in the art for rapidly and uniformly heating liquid products to a given temperature by resistance heating. We have found processing conditions applicable to higher treatment volumes of liquid food products or liquid feed products.

A second aspect of the present invention relates to a liquid product obtained by the method according to the present invention.

Escherichia coli in orange juice at pH 3.8 after various PEF treatment conditions, Listeria monocytogenes, Lactobacillus plantarum, Salmonella semftemberg (Salmonella) -A diagram showing a decrease in the number of living Saccharomyces cerevisiae. The left panel shows the PEF conditions currently in use, and the right panel shows the PEF conditions of the present invention. The various PEF treatment conditions associated with each panel are mentioned below the panel in FIG. 1B. The non-hollow triangle is 10 kV / cm, 2 microseconds, the non-hollow gray diamond shape is 15 kV / cm, 2 microseconds, and the hollow white circle is 20 kV / cm, 2 microseconds, hollow. Non-gray circles are 0.9 kV / cm, 1000 microseconds, black diamonds are 2.7 kV / cm, 1000 microseconds, and hollow white diamonds are 2.7 kV / cm, 100 microseconds. And the broken line is the detection limit. Escherichia coli in orange juice at pH 3.8 after various PEF treatment conditions, Listeria monocytogenes, Lactobacillus plantarum, Salmonella semftemberg (Salmonella) -A diagram showing a decrease in the number of living Saccharomyces cerevisiae. The left panel shows the PEF conditions currently in use, and the right panel shows the PEF conditions of the present invention. The various PEF treatment conditions associated with each panel are mentioned below the panel in FIG. 1B. The non-hollow triangle is 10 kV / cm, 2 microseconds, the non-hollow gray diamond shape is 15 kV / cm, 2 microseconds, and the hollow white circle is 20 kV / cm, 2 microseconds, hollow. Non-gray circles are 0.9 kV / cm, 1000 microseconds, black diamonds are 2.7 kV / cm, 1000 microseconds, and hollow white diamonds are 2.7 kV / cm, 100 microseconds. And the broken line is the detection limit. It is a temperature conductivity profile of orange juice (pH 3.8), coconut water (pH 5.0), and watermelon juice (pH 6.0). E. coli and L. coli in orange juice, coconut water and watermelon juice after PEF treatment at 2.7 kV / cm, 1000 microseconds. It is a figure which shows the decrease of the living number of monocytogenes (L. monocytogenes). FIG. 6 shows microbial analysis (n = 6) of untreated and PEF-treated orange juice, some of which were qualitative (FIG. 4B) and others of which were quantitative (FIG. 4A). It is a figure showing the perceptual evaluation of an orange juice sample stored at 7 ° C. and ambient temperature for the indicated period, and when comparable to freshly squeezed orange juice, the sample is shown as "good" and freshly squeezed. If not comparable to orange juice, it was indicated as "non-good". It is a figure which shows the amount of soluble solid matter (° brix) in orange juice before and after PEF treatment during storage at 7 degreeC and ambient temperature for 3 months. It is a figure which shows the acidity of the orange juice before and after the PEF treatment during storage at 7 degreeC and the ambient temperature for 3 months. It is a figure which shows the pH of the orange juice before and after the PEF treatment during storage at 7 degreeC and the ambient temperature for 3 months. It is a figure which shows the oil content of the orange juice before and after the PEF treatment during storage at 7 degreeC and the ambient temperature for 3 months. It is a figure which shows the vitamin C content of the orange juice before and after the PEF treatment during storage at 7 degreeC and the ambient temperature for 3 months. It is a figure which shows the pectin esterase activity before and after the PEF treatment during storage at 7 degreeC and the ambient temperature for 3 months.

We have found PEF processing conditions applicable to liquid food products and liquid feed products, and found that these conditions are equally effective in inactivating Gram-negative and Gram-positive bacteria. I found it. We have also found PEF processing conditions applicable to liquid food products and liquid feed products, which inactivate relatively large microorganisms and inactivate relatively small microorganisms. It has been found to be effective in both doing. Furthermore, we have surprisingly found that PEF processing conditions are applicable under conditions of relatively low pH and higher pH. Finally, we have found PEF processing conditions that are applicable to larger treatments of liquid food products and liquid feed products than were previously possible with currently available methods. ..

Thereby, the inventors provide a method of addressing many of the drawbacks associated with currently known methods for heating a liquid product to obtain a liquid product with reduced microbial loading.

The present invention is a method for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating to obtain a heated liquid product.
(A) Step of providing a liquid product;
(B) A step of providing an apparatus for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating;
(C) A step of supplying the liquid product to the inlet of the device and flowing the liquid product through the device;
(D) A step of continuously generating an electric current in the liquid product flowing through the apparatus, in which one minimum pulse is applied to each fluid element with a pulse duration of at least 10 microseconds during passage. And the electric field strength is 0.1 to 5 kV / cm;
Including
With respect to the method, the maximum temperature of the liquid product remains autonomously below 92 ° C. during said resistance heating.

According to the present invention, a method of rapidly and uniformly heating a liquid product to a heating temperature by resistance heating provides a heated liquid product with reduced microbial loading.

Heating a liquid product above a certain maximum temperature, eg, a predetermined maximum temperature, results in an unwanted reduction in fresh flavors, vitamins and nutrients present in the fresh (untreated) product and protein. Can cause degeneration of. The degree of component reduction and denaturation is related to the temperature and time at which the product is exposed to the treatment. Various liquid products are exposed to various temperature-time combinations to obtain the desired degree of enzyme and microbial inactivation. Therefore, alternative (non-) thermal processes with reduced temperature and / or time exposure to the product have received much attention, which can better retain the fresh characteristics of the product. Because it can be done. Better product quality can be expected if either the temperature or the exposure time can be reduced (shortened). In the method of the present invention, the exposure time to heat is significantly reduced and, depending on the process conditions selected, the maximum temperature of the liquid product remains autonomously below about 92 ° C. during resistance heating. Preferably, the maximum temperature of the liquid product autonomously remains below the critical temperature during resistance heating by the methods of the invention and, if present in the product, at this temperature of the liquid product, of the heat sensitive constituents. It is not subject to reduction or protein denaturation, but at the same time the microbial load in the liquid product is reduced to the desired acceptable level. Here, the method of the present invention makes it possible to apply processing conditions that prevent overheating of the liquid product while still effectively and efficiently reducing the microbial load of the liquid product.

In the method according to the invention, the pulse duration of a single pulse is an important factor rather than the total effective processing time. When a pulse duration of 2 microseconds and an electric field strength of 10 kV / cm were applied, the total effective processing time was four times that of the electric field strength of 20 kV / cm where conventional PEF treatment was used. It was determined that microbial inactivation was inefficient. Although not bound by theory, the explanation is that a reduced field strength of 10 kV / cm impairs the electroporation effect.

We have surprisingly found that microbial inactivation is conventional thermal pasteurization under conditions of low electric field strength of 0.1-5 kV / cm combined with an extended pulse duration of 100-1000 microseconds. It was found that sterilization or PEF treatment at 10 kV / cm was effective in achieving inactivation at less intensive temperature-time combinations than required. Although not bound by theory, these findings indicate that pulse duration was an important factor in the method according to the invention.

Although not bound by theory, the external electric field applied to the product has an effect on the protein channels and / or lipid domains of the cell membrane of the microorganisms present in the product. , Channels and / or causes conformational changes in the domain. Membrane protein channels open at a membrane potential of 50 mV, well below the 150-400 mV required for pore formation in the lipid bilayer (Tsong, 1992).

Since the opening and closing of many protein channels depends on the membrane potential difference, it is hypothesized that voltage-sensitive protein channels will be opened when electrical treatment is applied. Once these channels are opened, they carry a higher current than the current they are designed for. As a result, these channels can be irreversibly modified by Joule heating and / or electrical modification of their functional groups occurs (Tsong, 1992). Opening and closing of protein channels occurs in the submicrosecond time range, while protein denaturation takes several milliseconds to several seconds (Tsong, 1992).

This is 3 to 8 times less electric field strength than the lipid bilayer is affected, i.e. 20 kV / cm (ie, conventional PEF conditions) required for irreversible damage by electroporation of the lipid bilayer. In comparison, it suggests that protein channels can be affected by electric field strengths of 2.5 kV / cm to 7 kV / cm for inactivation of protein channels. According to the present invention, an electric field strength of 0.1 kV / cm to 5 kV / cm combined with a pulse duration of 10 to 1000 microseconds establishes efficient inactivation of microorganisms in the liquid product. Sufficient and effective with respect to. For example, the methods of the invention are applicable to liquid products in 1 L / h PEF equipment (“PEF system”). For the exemplary liquid product, the pulse duration is set to either 100 microseconds or 1000 microseconds and the selected field strength or "field strength" is either 0.9 kV / cm or 2.7 kV / cm. It was. The number of pulses applied to the liquid product varies between 0 and 35, and the time difference between two consecutive pulses varies between 0.6 ms and 199 ms, resulting in the result. The maximum temperature varied between 36 ° C and 92 ° C. See Example 1, a more detailed overview that demonstrates the efficiency of the methods of the invention below.

We have found that microbial inactivation is particularly efficient for vegetative cells. Presumably, based on this effect of the methods of the invention, the methods according to the invention also provide an efficient mechanism for inactivating spores. Spores contain essential proteins for germination in the endometrium and in the spore coat layer, which are targets of external electrical stimulation.

As a further example of the application of the method of the present invention, for example, a 1200 L / h PEF apparatus for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating is applied, and a batch of the liquid product is the present invention. Processed in a way. The pulse duration is 1000 microseconds and the electric field strength is 2.0 kV / cm. The number of pulses applied to the liquid product is about 5 pulses and the time difference between the two consecutive pulses is 3.8 ms. See also Example 3 below for detailed embodiments of the present invention.

One embodiment of the present invention is the method of the present invention, wherein the pH of the liquid product exceeds pH 1.5 to 9.0, preferably 4.6, preferably 4.8 to 9.0. It is preferably 5.5 to 8.0, more preferably 6.0 to 7.5. One embodiment of the present invention is the method of the present invention, in which the pH exceeds about 5.0, preferably about 6.0.

Further, in one embodiment, the invention comprises a liquid product having a pH of less than 4.6, preferably 1.5 to 4.6, more preferably about 1.5 to about 3.8. Regarding the method by.

In a further embodiment of the invention, in the method of the invention, the pH of the liquid product is higher than 4.6, preferably 4.6-9.0. One embodiment of the present invention is the method of the present invention, wherein the pH of the liquid product is 5.0 to 9.0, preferably 6.0 to 9.0.

Due to the applicability of the methods of the invention, liquid products with a wide range of pH are processed in one and the same way according to the invention. Since the method of the present invention is applicable to the processing of liquid products having such a wide variety of pHs, PEF processing is desirable, and a variety of liquid products that can be selected for processing by the method of the present invention. The sex is very large. Virtually any liquid product, ingredient, or semi-finished product applied, for example, in food processing, is suitable for rapid and uniform heating by the methods of the invention. Due to our surprising finding that the PEF process is effective and efficient over such a wide pH range, processing foods in a PEF-incorporated manner according to the invention is now more widespread than before. It is available.

Further, one embodiment of the present invention measures 0.01 to 10 S / m at 20 ° C., more preferably 0.1 to 3 S / m at 20 ° C., most preferably 20 ° C. It is a liquid product according to the present invention having an electrical conductivity of 0.2 S / m to 0.8 S / m.

Electrical conductivity within these indicated boundaries is typically preferred, as such electrical conductivity aids in rapid and uniform heating of the liquid product according to the methods of the invention. Since most liquid food products have intraboundary electrical conductivity that is applicable to the application of the methods of the invention in liquid food products, the methods of the invention are numerous in which it is desirable to reduce the microbial load. Applicable for processing liquid food products. For example, electrical conductivity at 20 ° C. has been measured for several batches of liquid food products, eg 0.1 S / m for cranberry juice, 0.15 S / m for beer, 0.2 S for apple juice. / M, 0.4 S / m for chocolate milk, 0.45 S / m for whole milk, 0.4 S / m for soy milk, 0.25 S / m for almond milk, 1.0 S / m for carrot juice It was 1.8 S / m for m and tomato sauce. Therefore, the methods of the present invention are applicable to a wide variety of liquid food products.

Although not bound by theory, the PEF processing conditions of the present invention appear to pave the way for new mechanisms for microbial inactivation.

We have found that when the PEF processing conditions of the present invention are applied, the liquid food product is effectively pasteurized (ie, the microorganisms are effectively and efficiently inactivated). .. The conditions are the maximum temperature of the liquid food product at 40 ° C. to 92 ° C., preferably 50 ° C. to 92 ° C., more preferably about 60 ° C. to 85 ° C., from 10 to 1000 microseconds, preferably about 1000 microseconds. More preferably, in combination with a pulse duration of about 100 microseconds, it comprises a surprisingly low field strength of 0.1 to 5 kV / cm, preferably 4 kV / cm or less, more preferably 3 kV / cm or less.

According to the present invention, in pasteurization when applying the method of the present invention, the number of pulses applied to the liquid product continuously flowing during passage through the treatment zone is preferably at least 1 for each fluid element. Is particularly efficient and effective when is 1 to 100, more preferably 5 to 50.

The number of pulses is given by equation 1, where n is the number of pulses, V is the volume of the processing chamber (L), f is the pulse frequency used (Hz), and φ is. , Flow velocity (L / h).

The number of pulses in designing the process, according to the invention, as long as at least one pulse is applied to all fluid elements flowing through the processing chamber of the apparatus for heating the liquid product rapidly and uniformly. Is not an important step. To ensure that at least one pulse is applied to all fluid elements, the system must be designed with the purpose of residence time in the processing chamber greater than 1 / f. The number of the applied pulses, the desired throughput of the liquid product (φ, L / h), conductivity of the liquid product (σ, S / m), the specific heat capacity of the product _(c p, kJ / kg K), product density (ρ, kg / m ³ ), applied electric field strength (E, V / m), pulse duration (τ- _pulse , s), and temperature gradient obtained in the process (ΔT). , ° C) (the difference between the inlet and outlet temperatures of the liquid product) is the result of the process design. The relationship between these parameters is given by equation 2.

As mentioned earlier, and as further illustrated in Example 1 below, pulse duration is important to the effectiveness of PEF treatment by the methods of the invention. Therefore, the application of one relatively long pulse may be more effective than the application of a shorter pulse with a similar total effective processing time.

For example, for a liquid product processed in a 1 L / hr continuous flow 1 L / h PEF device according to the method of the invention, the pulse duration is typically about 100-1000 microseconds and the electric field strength is about. It is 2.7 kV / cm. Then, typically, the number of pulses is about 1-25, depending on the desired temperature increase across the processing chamber, about 0.6-39 ms, which is between two consecutive pulses. The time difference is the difference between the inlet temperature and the maximum temperature.

For example, for a liquid product batch processed in a 1200 L / h PEF apparatus according to the method of the invention, the pulse duration is typically 1000 microseconds and the electric field strength is about 2.0 kV in the method according to the invention. / Cm. Then, typically, the number of pulses is about 5, and the time difference between the two consecutive pulses is about 3.8 ms.

Typically, the methods of the invention are applicable to processing liquid products in a PEF apparatus having a throughput of 30 L / h to 200 L / h according to the present invention.

Typically, the methods of the invention are applicable to processing liquid products in a PEF apparatus having a throughput of about 30,000 L / h according to the invention.

One embodiment of the present invention is the method of the present invention, wherein the apparatus for rapidly and uniformly heating the liquid product to a predetermined temperature is about 1 L / h to about 30,000 L / h, preferably about. It has a processing amount of 1 L / h or about 30 L / h, or about 200 L / h, or about 1200 L / h, or about 30,000 L / h.

Again, without wishing to be bound by theory, these PEF processing conditions of the present invention result in the inactivation of microorganisms according to a theoretical mechanism that inactivates protein channels in cell membranes. These new PEF processing conditions of the present invention provide new opportunities for microbial inactivation compared to the currently used treatment conditions for PEF. Under these new conditions of the invention, all vegetative microorganisms do not prioritize inactivation for relatively large and / or Gram-negative bacteria compared to relatively small and / or Gram-positive bacteria. Inactivated.

Thus, the methods of the invention inactivate Gram-negative bacteria such as Escherichia coli strains, Salmonella species, other Enterobacteriaceae and Acetobacter in the liquid product. Further, in the method of the present invention, for example, Listeria monocytogenes, Lactobacillus plantarum, Leuconostoc strain and Streptococcus-forming liquid-positive Gram-positive Inactivated in the object. Based on the expected theoretical mechanism, the cell membrane of spore-forming bacteria also contains potential-dependent channels, such as Alicyclobacillus and Clostridium, and thus also inactivates spore-forming bacteria. It is expected that it can be done. In addition, the spores themselves contain proteins essential for germination in the endometrium and spore coat layer that can be targeted by externally applied pulses.

In addition, the methods of the invention are suitable for inactivating the liquid products of relatively large microorganisms such as yeast and mold, for example. The method of the present invention is described in, for example, L. It is also suitable for inactivation in relatively small microorganisms and liquid products such as L. monocytogenes. Of course, the methods of the invention are equally suitable for the inactivation of the microorganisms of these illustrated microorganism sizes shown in Example 1 in the liquid product.

Secondly, the PEF processing conditions of the present invention are equally effective in inactivating microorganisms in liquid food products having relatively low pH and in liquid food products having relatively high pH. Under these new conditions of the present invention, the inactivation of microorganisms in all types of (liquid food) products is possible.

Finally, these conditions of the invention are used because the lower peak voltage is used in the present invention due to the lower electric field strength used in the PEF processing conditions of the present invention as compared to the currently applied PEF conditions. Is easily scaled up and the PEF processing conditions of the present invention are applicable to industrial implementation in larger volumes and throughputs. The method of the present invention is a method of the present invention in an apparatus for rapidly and uniformly heating a liquid product to a heating temperature by resistance heating when the residence time of the fluid in a high electric field region is about 17 ms to 2 seconds. Especially suitable for liquid food products served in. The frequency of the pulse is limited to 1 kHz to 50 kHz to avoid metal emission of the electrodes (Mastwijk, 2006). Typically, according to the invention, in the method of the invention, the flow rate of the liquid product is, in this case, from about 1 L / h to 5000 L / h, preferably from about 1000 L / h to 30,000 L / h.

A further embodiment of the invention is a method according to any of the previous embodiments of the invention, wherein the liquid product is a liquid food product or a liquid feed product.

Further, one embodiment of the invention is the method according to the invention, wherein the liquid product is an ingredient, a semi-finished product, or a fruit juice, vegetable juice, infant food, jam, spread or smoothie, alcohol or non-. Final liquid products such as alcoholic beverages, dairy products, vegetable milk products, liquid eggs, soups, or sauces.

In one embodiment of the invention, the dairy product is selected from milk, a milk product, or a liquid composition comprising a milk component or a milk fraction, the liquid product being heated to a predetermined temperature by resistance heating according to the invention. It is a method for heating rapidly and uniformly.

Further, an embodiment of the present invention is the method according to the present invention, wherein the liquid product is a dairy product containing milk, a milk product, a milk component or a milk fraction.

An important aspect of the invention is to keep the temperature of the liquid product below about 92 ° C, or less than about 85 ° C, or less than about 70 ° C, or less than about 60 ° C during PEF processing, according to the invention. In addition, it is a finding that no cooling section is required between the processing chambers of the equipment applied in the method according to the invention.

As mentioned earlier, current state-of-the-art methods add a cooling section between the processing chambers to avoid overheating of the liquid product. As mentioned above, the method of the present invention requires cooling because heating at critical temperature is very fast (residence time in the processing chamber, less than 1 second) and exposure time to maximum temperature is very short. Not being is an important finding within the present invention.

In order to have an efficient method, only exposure to critical temperature (the temperature at which product quality will be affected) is obtained with electrical energy, as described in the present invention. The non-critical temperature domain can be preheated by conventional heating. Therefore, preferably in the method of the invention, the liquid product is preheated to a temperature in the range of 20 ° C. to 70 ° C., preferably 35 ° C. to 65 ° C., more preferably 40 ° C. to 60 ° C. before being fed to the apparatus. NS. One embodiment of the invention is the method according to the invention, wherein the liquid product is 20 ° C. to 70 ° C., preferably 35 ° C. to 65 ° C., more preferably 40 ° C. to 60 ° C. before being fed to the apparatus. It is preheated to a temperature in the range of.

For practical purposes, liquid products such as liquid food products used in the methods according to the invention are immediately cooled below ambient temperature, eg, 2-8 ° C. Since no retention time is required, cooling of the liquid product proceeds immediately after the liquid product leaves the high field region (preferably within 3 seconds). In the current process design of the present invention, the cooling pipe can be installed 0.5 m downstream from the high electric field region. For a 30,000 L / h (= 8.3 L / s) system with a 3 inch pipe diameter, this is the first from 2.2 L volume or 0.27 seconds and maximum temperature before entering the first cooling section. Approximately 1-5 seconds (depending on viscosity) before the critical temperature drop of 10 ° C. is established, followed by (conventional) cooling towards the desired outlet temperature, usually in the range of 4-7 ° C. means.

The term "autonomous" has its usual meaning and here refers to the temperature of a liquid product that reaches a certain value in the methods of the invention during processing without the help of external cooling (or heating).

For example, liquid products such as liquid food products selected from orange juice, dairy products, coconut water, watermelon juice are preheated to, for example, about 40 ° C, about 50 ° C or about 60 ° C. For example, the liquid product is preheated to about 30 ° C. to 65 ° C., preferably 36 ° C. to 59 ° C., in order to efficiently and effectively inactivate the microorganisms in the liquid product when the method of the invention is applied. Will be done.

Preferably, the maximum temperature of the liquid product remains autonomously below about 85 ° C., more preferably less than about 70 ° C., less than about 63 ° C., or less than about 60 ° C. during resistance heating, according to the invention. be. It is part of the invention that the process parameters of the method of the invention are selected such that the maximum temperature of the liquid product remains below the temperature chosen autonomously. When applying the methods of the invention, below selected temperatures, efficient and effective killing of microorganisms present in the liquid product is guaranteed, while in the fresh (untreated) liquid product. Undesirable reduction of existing fresh flavors, vitamins, and nutrients and denaturation of proteins are prevented, or at least most of them are prevented.

By applying the electric field strength according to the invention with the pulse duration according to the invention, we are surprised to find that, according to the invention, the temperature of the liquid product is about 92 ° C., or about 85 ° C. Alternatively, it has been found that it remains below the maximum temperature of about 70 ° C., or about 60 ° C., which makes the methods of the invention particularly suitable for practice in large-scale settings, such as commercial settings.
An example of a commercial application of the method of the present invention is the processing of a liquid food product in an apparatus for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating, according to the invention. The flow of liquid food product through is from about 500 L / h to 30,000 L / h, for example about 1200 L / h.

Therefore, one embodiment of the invention is the method of the invention, in which the temperature of the liquid product is autonomously below 85 ° C., preferably below 75 ° C., more preferably below 60 ° C. during resistance heating. There is up to.

In one embodiment, the invention relates to a method according to the invention, wherein the electric field strength is less than about 5 kV / cm.

One embodiment of the present invention is the method according to the present invention, in which the electric field strength is 0.5 to 5 kV / cm, more preferably 2.5 to 4 kV / cm. In one embodiment, the present invention relates to the method of the present invention, wherein the electric field strength is less than about 3 kV / cm, preferably about 2.7 kV / cm or less, more preferably about 0.9 to about 2.5 kV / cm.

One embodiment of the invention is the method according to the invention, with a pulse duration of at least 10 microseconds, more preferably 10 to 2000 microseconds, even more preferably 50 to 500 microseconds, most preferably 50 to 50 microseconds. 100 microseconds. In one embodiment, the invention relates to a method according to the invention, wherein the pulse duration is from about 100 microseconds to about 1000 microseconds. In a further embodiment, the invention relates to a method according to the invention, wherein the pulse duration is 1000 microseconds or less, preferably about 100 microseconds.

One embodiment of the present invention is the method according to the present invention, in which the applied pulse is a bipolar pulse. Therefore, one embodiment of the present invention is the method according to the present invention, in which the minimum one pulse applied to the liquid product is a pulse applied in the form of a bipolar pulse. It is advantageous to apply bipolar pulses to the liquid product to avoid electrode damage (Loeffler, 1996). Of course, it is part of the invention that other types of pulses are equally applicable to the methods of the invention.

The methods of the present invention are particularly suitable for the inactivation of microorganisms in liquid food products such as juices, sauces and dairy products. That is, examples of such liquid food products are ingredients, semi-finished products, or fruit juices, vegetable juices, infant foods, jams, spreads or smoothies, alcoholic or non-alcoholic beverages, dairy products, vegetable milk products. , Liquid eggs, final liquid products such as soups or sauces.

One embodiment of the present invention is the method according to the present invention, which is a method for inactivating microorganisms in a liquid product. As mentioned above, the method of the present invention has many advantages over current methods for heating liquid products by resistance heating. One of the main advantages achievable with the methods of the invention is pasteurization of liquid products, such as liquid food products, where the microorganisms are small or large and the microorganisms are Gram-negative or Gram-positive. It is a fungus.

For practical purposes, liquid products such as, for example, liquid food products that are subjected to the method according to the invention are below ambient temperature immediately after the method of the invention is applied to the liquid product, eg, 2 It is cooled to ~ 8 ° C.

Therefore, one embodiment of the present invention is a method in which a heated liquid product is cooled immediately after flowing through an apparatus for rapidly and uniformly heating the liquid product to a predetermined temperature by resistance heating. be. Of course, for practical purposes, cooling is properly applied immediately from the moment the liquid product is flushed through the device, eg, as fast as possible, preferably within 3 seconds. Therefore, an embodiment of the present invention is also a method in which a heated liquid product is transferred through a device for rapidly and uniformly heating the liquid product to a heating temperature by resistance heating and then cooled. The method according to the invention is suitable for application using an apparatus for rapidly and uniformly heating a liquid product to a heating temperature by resistance heating, wherein the apparatus is from about 0.5 L / h to about 2000 L / h, preferably about 2000 L / h. About 0.5 L / h to about 2 L / h, more preferably about 1 L / h, or equally preferably about 100 L / h to about 2000 L / h, preferably about 1000 to 1500 L / h, more preferably about 1200 L / h. It is part of the present invention that it is operated at the flow velocity of the liquid product at h. Preferably, the flow velocity is about 30,000 L / h.

As mentioned earlier, inactivation during the process by preheating the liquid product before subjecting it to a method for rapidly and uniformly heating the liquid product to a predetermined temperature by resistance heating. Improved efficiency has been previously established. However, it was a prerequisite to cool the liquid product during the course of the process, as the liquid product was heated to unacceptable values when applying the PEF conditions known in the art. As mentioned above, it is an important finding within the invention that cooling during the process of the invention is not required, as the temperature of the liquid product does not exceed an unacceptable threshold here. Therefore, in the method of the invention, the liquid product does not need to be cooled during the process and is well preheated before being subjected to a process of rapidly and uniformly heating the liquid product to a predetermined temperature by resistance heating. ..

Thus, preferably, in the methods of the invention, the liquid product is up to a temperature in the range of 20 ° C. to 70 ° C., preferably 35 ° C. to 65 ° C., more preferably 40 ° C. to 60 ° C. before being fed to the apparatus. Preheated.

The method according to the invention is very effective in inactivating the microorganisms present in the liquid product used in the method of rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating according to the present invention. And efficient. When the method of the present invention is applied to a liquid product containing microorganisms, the number of microorganisms is reduced by at least 2 log cfu / mL, most preferably 6 log cfu / mL or more.

Therefore, preferably, the present invention reduces the number of microorganisms (colony forming unit cfu) in the liquid product by at least 2 log cfu / mL, preferably at least 5 log cfu / mL, and most preferably by 6 log cfu / mL or more. be. One embodiment of the invention is the method according to the invention in which the number of microorganisms in a liquid product is reduced by at least 4 log cfu / mL, preferably at least 7 log cfu / mL.

We have established that such a reduction in the number of microorganisms in a liquid food product contributes significantly to the preservation of the product quality scale. For example, the taste, odor, color, and appearance of liquid food products used in the methods of the invention are preserved for a longer period of time as compared to untreated liquid food products. For example, in the method of the present invention, the taste and smell of the liquid food product orange juice is preserved for about 60 days or more when the method of the present invention is applied to orange juice when the juice is stored at about 7 ° C. If the orange juice is subjected to the method of the invention and then kept at ambient temperature, it is equally beneficial to enhance the quality of the orange juice for about 23 days.

It can be seen that the method of the present invention is equally applicable to inactivating Gram-positive bacteria in liquid products and inactivating Gram-negative bacteria in liquid products. Moreover, the size of the microorganism does not play a limiting role, which means that smaller or larger sized microorganisms are inactivated by the methods according to the invention. Therefore, until the present invention, small Gram-positive bacteria could not be efficiently inactivated by known methods for rapidly and uniformly heating liquid products to heating temperature by resistance heating. Various aspects and embodiments of the above are important contributions to the art.

Therefore, one embodiment of the present invention is the method of the present invention, wherein the microorganism optionally comprises Gram-positive bacteria.

When the methods of the invention are applied to liquid products, the pH of the liquid products, if any, is modified to a minimum during the course of the process. This is a further beneficial feature of the method according to the invention.

Therefore, in one embodiment, the present invention states that the pH of the liquid product at the end of the process is within 0.5 pH units, preferably within 0.2 pH units, more preferably 0.1 pH from the pH at the start of the process. With respect to the method, within units, most preferably within 0.05 pH units.

The method according to the invention is suitable for liquid products, especially liquid food products.

A second aspect of the present invention relates to a liquid product obtained by the method according to the present invention described above.

The present invention is further described by the following non-limiting examples provided below.

[Example 1]
<E. coli at pH = 3.6 in orange juice, Listeria monocytogenes, Lactobacillus plantarum, Salmonella Semftemberg and Salmonella Sentenberg・ Microbial inactivation of Saccharomyces cerevisiae (1 L / h scale)>
Pathogenic and putrefactive microorganisms were selected based on their morphology and their association and prevalence with fruit juices. In addition, the heat resistance or PEF resistance of the strain was used as a criterion for strain selection. The selected microorganisms are listed in Table 1.

Table 1. Bacterial strain and yeast strain used in this example

Fresh cultures of all five microorganisms listed in Table 1 were prepared by seeding from frozen stock to medium and culturing overnight. Escherichia coli, Listeria monocytogenes, Saccharomyces cerevisiae, and Salmonella entesia subspecies Serotype S. et al. , 2014. Fresh cultures of Lactobacillus plantarum from frozen stock into MRS medium containing 52.2 g of MRS (De Man, Rosoga and Sharp broth, Merck) and 12 g of agar per 1 L of distilled water. Was similarly prepared by sowing. The plates were incubated overnight at 30 ° C. Single colonies were inoculated into 100 mL flasks with 10 mL MRS broth and cultured in a shaking incubator (180 rpm) at 20 ° C. for 24 hours. From this culture, 200 microliters were inoculated into fresh 19.8 mL MRS broth, replenished with 1% glucose (100 mL flask) and incubated at 20 ° C. and 180 rpm for 24 hours.

After culturing, cells were washed and a suspension of selected microorganisms was added to orange juice (Minute Maid®) to a final concentration of approximately 1.0E + 8-1.0E + 9 cfu / mL.

The inoculated suspension was pumped through a 1 L / h PEF system at a flow rate of 13.0 ± 0.5 mL / min and preheated to 36 ° C. prior to electrical treatment. The suspension then entered two vertically arranged co-linear processing chambers where electrical processing was performed. Due to the variety of processing conditions (electric field strength, pulse duration, and number of pulses applied), the juice was autonomously heated to a variable maximum temperature (Table 2 illustrates the conditions used). ). Since no retention section was added, the juice was cooled via a heating spiral immersed in a water bath immediately after leaving the processing chamber (within 3 seconds). At the outlet, samples were collected aseptically. Due to the variety of frequencies selected, the number of pulses applied and the resulting temperature rise leading to the highest temperature varied (Table 2). Samples were collected at various maximum temperatures to determine dynamics with respect to fixed field strength and pulse duration.

The number of viable microbial cells was subdiluted with 100 μL of serially diluted PEF-treated juice in sterile Peptone physiological saline diluent (PSDF) on a suitable agar plate supplemented with 0.1% sodium pyruvate. Determined by enhancing the proliferation of lethally damaged cells (Timers et al., 2014). 25 ° C. (S. cerevisiae), 30 ° C. (L. monocytogenes, L. plantarum), or 37 ° C. (S. Senftenberg (S. cerevisiae)). Surviving cells were counted after 5 days of incubation in Senftenberg), E. coli).

The temperatures before and immediately after the processing chamber were measured using a HYP-OT thermocouple (omega). In addition, the maximum temperature was indirectly measured using an NTC resistor and the maximum temperature was monitored. Square wave bipolar pulses, voltages, and currents in the processing chamber were recorded on a digital oscilloscope (Rigol DS1102E). Electrical energy is obtained by numerical integration of voltage and current traces according to (Mastwijk, 2006) and (Timemers et al. 2014), which is equal to caloric power within the experimental error (10%).

All experiments were performed in duplicate.

Table 2: PEF conditions used in this study

The conditions tested are provided in Table 2. The dimensions of the processing chamber varied to obtain variable field strength. As a result of this variable dimension, the residence time in the processing chamber was not the same for each condition tested. The frequency was adjusted to obtain the desired maximum temperature. Finally, the number of pulses was calculated by taking the product of the residence time used and the frequency. The time between the two pulses was calculated by dividing the residence time by the number of pulses and subtracting the pulse duration.

Inactivation is shown as the logarithm N of the number of surviving microorganisms under the conditions tested, divided by the starting concentration of microorganisms, where N ₀ is _log-10 (N / N ₀ ). In FIG. 1, inactivation is shown as a function of the maximum temperature at the exit of the treatment chamber for the microorganisms tested. The left panel shows inactivation at various field strengths for short pulses (2 microseconds), which shows current state-of-the-art technology, and the right panel shows for long pulses (ie 100 or 1000 microseconds). It exhibits inactivation at various electric field strengths, demonstrating the invention described in this application.

The inactivation of E. coli in orange juice as a function of electric field strength and fluctuating pulse duration is shown in FIG.

The left panel of FIG. 1 (electric field strength is 10 kV / cm, 15 kV / cm or 20 kV / cm, pulse duration is 2 microseconds) and the right panel of FIG. 1 (electric field strength is 0. As can be seen in FIG. 1 from both 9 kV / cm or 2.7 kV / cm with a pulse duration of 1000 microseconds), the increase in electric field strength at a constant pulse duration is optional. It resulted in more inactivation at the highest temperature.

The effect of this increased field strength was also observed with the other microorganisms shown in FIG. 1 and showed more inactivation at higher field strengths.

Further increases in electric field strength under the conditions described in the present invention up to 5 kV / cm are expected to further reduce the maximum temperature while still inactivating the desired number of microorganisms.

Although not bound by theory, this is explained by the excess of the externally applied electric field (E) with respect to the critical electric field strength (E-c) of the film, resulting in more damage and inactivation ( Alvarez et al., 2006). There is a consensus in the art that E-c varies by various microorganisms and that microbial inactivation requires an electric field strength of greater than 5 kV / cm (Rasso et al., 2014). However, here, the present inventors, according to the present invention, 1.0E-7 (N / N) microorganisms in which the electric field strength of less than 5 kV / cm, for example, less than 2.7 kV / cm is inactivated. It is also shown for the first time that it is effective to achieve at a sufficient level below _0). Comparing the left and right panels of FIG. 1, an electric field strength of 2.7 kV / cm with a pulse duration of 100 microseconds or 1000 microseconds is higher than that commonly applied in the art. And it can be seen that it results in a higher degree of microbial inactivation compared to shorter pulse durations.

Although not bound by theory, it has been proposed that pulse duration plays an important role in inactivating microorganisms and is used today with respect to the conditions described in the present invention. Implies another mechanism. It is hypothesized that if the pulse duration is long enough, voltage-sensitive protein channels will be opened and will carry higher currents than designed. As a result, the channels are irreversibly denatured and the cells lose their viability.

The E. coli inactivation data showed no difference in the degree of inactivation at 2.7 kV / cm when a pulse of 100 or 1000 microseconds was used, which is the criticality of the pulse. It implies a duration of less than 100 microseconds.

Furthermore, it can be seen that both Gram-negative and Gram-positive bacteria _{can be inactivated to 1.0E-7 (N / N 0) using this new PEF condition.} Also, microbial size does not play a significant role in the degree of inactivation (right panel) as it does under current state-of-the-art conditions (left panel). Yeast is 2.7 kV / cm and L. L. plantarum and S. It can be inactivated at higher maximum temperatures than S. Senftenberg, but under new PEF conditions E. coli and L. coli. No major differences are found between L. monocytogenes. On the other hand, a larger difference was found in the currently used PEF conditions (left panel).
[Example 2]
<Microbial inactivation of E. coli and Listeria monocytogenes in products with variable properties (1 L / h scale)>

Escherichia coli (ATCC35218) and Listeria monocytogenes NV8 were prepared from frozen stock according to the method described in (Timers et al., 2014) and tried. )) Or in Brain Heart Infusion Broth (L. monocytogenes).

After culturing, the cells are washed and of different pH and conductivity (see Figure 2), orange juice (Minute Maid®), coconut water (Healthy People), or watermelon juice (freshly squeezed). Suspended in either to reach a concentration of about 1.0 E + 8 cfu / mL. Using the same PEF equipment system and setup as described in Example 1, the flow rate was 13.5 ± 0.5 mL / min. However, only one process condition with an electric field strength of 2.7 kV / cm and a pulse duration of 1000 microseconds was used. Due to the different conductivity among the three products (FIG. 2), different frequency settings were required and the number of pulses given during the process to reach similar maximum temperatures for each product was different. Table 3 shows the range of frequencies tested for the three tested juices, the number of pulses, and the maximum temperature after treatment.

Inactivation is shown as the logarithm N of the number of surviving microorganisms under the conditions tested, divided by the starting concentration of microorganisms, where N ₀ is _log-10 (N / N ₀ ).

Table 3: PEF conditions used in this study

E. coli and L. coli at various maximum temperatures in various juices. The inactivation of L. monocytogenes is shown in FIG. E. coli (Fig. 3A) and L. coli. Comparing the inactivation curves of L. monocytogenes (Fig. 3B) for various liquid media, namely liquid products orange juice, coconut water, and watermelon juice, observed differences in the temperature at which inactivation begins. It turns out that it will not be done. In addition, the degree of inactivation is similar for orange juice, coconut water, and watermelon juice. This indicates that the product matrix, i.e. the different pH of the liquid product orange juice, coconut water and watermelon juice, and the different electrical conductivity has no effect on the degree of inactivation. In addition, a comparable inactivation curve showing inactivation at similar temperatures is found in Gram-negative E. coli (approximately 0.7-1.5 micrometer x 2-5 micrometer bacterial cells). Has a size) and Gram-positive L. coli. Found with respect to monocytogenes (having a cell size of approximately 0.4-0.5 micrometers x 0.5-2 micrometers), which differ in terms of the microorganisms used. Indicates that there is no such thing.

This is the same as the method of the invention applied to orange juice, coconut water, and watermelon juice with respect to the inactivation of Gram-positive and Gram-negative bacteria in a variety of different liquid media, i.e. different liquid food products. It shows that it is effective and efficient.
[Example 3]
<Microbial verification on 1200 L / h scale>

One batch of oranges (variety: Natural folha murcha) was commercially compressed to give 4.000 L of orange juice. Since the orange juice was not inoculated with microorganisms, the naturally occurring microbial populations in orange juice were studied to verify the previously determined process conditions at 1 L / h on a large scale (1.200 L / h).

Orange juice was pumped at a flow of 1.200 ± 100 L / h and preheated to 59 ° C. The pulse was applied to three vertically arranged processing chambers to reach a maximum temperature of 70 ° C. The volume of the treatment zone in the central treatment chamber was smaller than the treatment zones (dimension length 14 mm, diameter 12 mm) of the first and third (outer) treatment chambers (dimension length 14 mm, diameter 8 mm). Due to the power connection, this results in twice the field strength in the central processing chamber compared to the outer processing chamber, 1.9 kV / cm in the center and 1.0 kV / cm in the outer processing chamber. there were. The pulse had a fixed duration of 1000 microseconds. The residence time in the central processing chamber is 5.4 ± 0.5 ms, in the outer processing chamber it is 9.5 ± 0.9 ms, and the total number of pulses applied during processing is 207 ±. The repetition rate at 18 Hz was 5.1 ± 0.01.

The juice was cooled within 3 seconds immediately after leaving the processing chamber, minimizing retention time and reducing the heat load of the product. After cooling, the juice was packaged and aseptically and stored.

Microbial samples of untreated and PEF-treated juice were analyzed in duplicate in three different laboratories, with a total of six untreated and PEF samples. Total mesophilic plate numbers, total coliforms, and yeast and mold numbers were analyzed according to the method described by Dows and ITO (2001). Eguchi et al. , (1999), Salmonella was analyzed according to AOAC (2000), Listeria monocytogenes was analyzed according to ISO11290-1 (1996), and lactic acid bacteria were analyzed according to Silva. et al. The analysis was performed according to the method of (2007).

The results of microbial inactivation in untreated orange juice and PEF-treated orange juice are shown in FIGS. 4A and 4B. The initial microbial load in untreated orange juice was relatively high, with approximately 1.0E + 5 cfu / mL present in untreated orange juice (FIG. 4A). After PEF treatment of orange juice, no microorganisms were detected on the plates (total number of plates), and no mold, yeast or coliforms were detected on the plates (all plates show <1 cfu / mL. See FIG. 4A). In FIG. 4B, the qualitative data show that all lactic acid bacteria and thermophilic thermophilic spore-forming bacteria (ATSB) present in the untreated orange juice were also inactivated during the PEF treatment of the orange juice.
[Example 4]
<Effect of PEF conditions of the present invention on quality aspect and microbial shelf life>

PEF-treated orange juice samples prepared and described in Example 3 were analyzed weekly for a shelf life test lasting 3 months. During this period, quality aspects and microbial counts were analyzed. Microbial analysis was similar to the process described in Example 3. With respect to quality analysis, the amount of soluble solids (JBT FoodTech Citrus Systems, 2011-1), acidity (JBT FoodTech Citrus Systems, 2011-2), pH (JBT FoodTech Citrus), according to methods generally known in the art. Systems, 2011-3), oil content (JBT FoodTech Citrus Systems, 2011-4), vitamin C content (JBT FoodTech Citrus Systems, 2011-5), and pectinesterase activity (Rouse and Atkins), 19 It was measured in a short time (Table 2). Samples were stored at both 7 ° C. and room temperature (ambient temperature 20-25 ° C.) to accelerate accelerated shelf life testing.

Microbial evaluation of the sample was performed, and the results are shown in Table 2. The number of microorganisms in the PEF-treated sample is below the detection limit of 1 cfu / mL during the entire shelf life study lasting 104 days, both when the orange juice was stored at 7 ° C. and when the orange juice was stored at ambient temperature. I showed that.

Table 4.7 Results of microbial analysis of PEF-treated orange juice during storage, stored at either 4.7 ° C or ambient temperature.

Perceptual assessment was performed by a trained panel. Samples were evaluated for overall appearance, color, odor, and taste. A sample was rated "good" if it had similar properties to freshly squeezed orange juice, and a sample was rated "non-defective" if it was no longer of high quality freshly squeezed juice.

The results of perceptual analysis during shelf life are shown in FIG. Samples stored at 7 ° C. were shown to resemble freshly squeezed orange juice for a shelf life of 64 days. After this storage time, the fresh smell and taste diminished, but the appearance and color of the juice was considered "good" until 90 days of storage at 7 ° C. (Fig. 5A). PEF-treated juices stored at ambient temperature were only allowed up to 27 days of storage (Fig. 5B). After this time, it was no longer considered fresh juice.

The quality aspect of orange juice was also monitored. For these aspects, the untreated juice was similarly evaluated prior to PEF treatment to assess the effect of the process on these aspects.

Amount of total soluble solids (° Brix) (Fig. 6. Curves of experiments performed at ambient temperature and 7 ° C. match after day 6 of the graph), acidity (Fig. 7), pH (Fig. 8). ), Oil content (FIG. 9) and vitamin C content (FIG. 10) did not change with PEF treatment and storage at 7 ° C. and storage at ambient temperature.

Differences are found in pectinmethylesterase (PME) activity before and after PEF treatment (Fig. 11). Due to the temperature generated during the PEF process, some of the PME enzymes are inactivated, leading to a decrease in enzyme activity. That is, the heat generated during the process according to the invention raised the temperature of the juice to 70 ° C. The juice subjected to the PEF process of the present invention was preheated to 59 ° C. before being introduced into the PEF apparatus under continuous flow. The inactivation of this enzyme is one of the purposes of pasteurization because the residual activity of the enzyme can lead to the disappearance of turbidity and gelation of the citrus juice during storage. The activity of this enzyme is expressed as the release of acid per mL (multiplied by 1.0E + 4) during pectin hydrolysis as a function of time at pH 7.8 and 20 ° C. In general, most juice sterilizers provide juices with pectinesterase activity represented by pectinesterase units (PEUs) having values of 1.0E-6 to 1.0E-4.
The contents of the invention relating to the scope of claims at the time of filing the application of the present application are as follows.
[1]
A method for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating to obtain a heated liquid product.
(A) Step of providing a liquid product;
(B) A step of providing an apparatus for rapidly and uniformly heating a liquid product to a predetermined temperature by resistance heating;
(C) A step of supplying the liquid product to the inlet of the device and flowing the liquid product through the device;
(D) A step of continuously generating an electric current in the liquid product flowing through the apparatus, in which one minimum pulse is applied to each fluid element with a pulse duration of at least 10 microseconds during passage. And the electric field strength is 0.1 to 5 kV / cm.
Including
The maximum temperature of the liquid product remains autonomously below 92 ° C. during said resistance heating.
The above method.
[2]
The pH of the liquid product exceeds pH 1.5 to 9.0, preferably 4.6, preferably 4.8 to 9.0, more preferably 5.5 to 8.0, and more preferably 6. The method according to [1], which is 0 to 7.5.
[3]
The liquid product is 0.01-10 S / m measured at 20 ° C., preferably 0.1 to 3 S / m measured at 20 ° C., more preferably 0.2 S / m measured at 20 ° C. The method according to [1] or [2], which has an electrical conductivity of ~ 0.8 S / m.
[4]
The method according to any one of [1] to [3], wherein the liquid product is a liquid food product or a liquid feed product.
[5]
The liquid product may be an ingredient, a semi-finished product, or a fruit juice, vegetable juice or smoothie, jam, spread, alcoholic or non-alcoholic beverage, dairy product, vegetable milk product, liquid egg, soup, or sauce. The method according to any one of [1] to [4], which is the final liquid product as described above.
[6]
The method of [5], wherein the dairy product is selected from milk, a milk product, or a liquid composition comprising a milk component or milk fraction.
[7]
In any one of [1] to [6], the temperature of the liquid product remains autonomously below 85 ° C., preferably below 75 ° C., more preferably below 60 ° C. during the resistance heating. The method described.
[8]
The method according to any one of [1] to [7], wherein the electric field strength is 0.5 to 5 kV / cm, preferably 2.5 to 4 kV / cm.
[9]
Any one of [1] to [8], wherein the pulse duration is at least 10 microseconds, preferably 10 to 2000 microseconds, more preferably 50 to 500 microseconds, and most preferably 50 to 100 microseconds. The method described in the section.
[10]
The method according to any one of [1] to [9], wherein a bipolar pulse is applied.
[11]
The method according to any one of [1] to [10], which is a method for inactivating microorganisms in the liquid product.
[12]
The method according to any one of [1] to [11], wherein the heated liquid product is cooled immediately after flowing through the apparatus.
[13]
The liquid product is preheated to a temperature in the range of 20 ° C. to 70 ° C., preferably 35 ° C. to 65 ° C., more preferably 40 ° C. to 60 ° C. before being fed to the apparatus, [1] to. The method according to any one of [12].
[14]
Any one of [1] to [13], wherein the number of microorganisms (cfu) in the liquid product is reduced by at least 2 log cfu / mL, preferably at least 5 log cfu / mL, and most preferably 6 log cfu / mL or more. The method described in.
[15]
The liquid product obtained by the method according to any one of [1] to [14].
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Claims (20)

A method for the enzymatic inactivation, microbial inactivation, or combination of liquid products of liquid products by resistance heating.
(A) Step of providing a liquid product;
(B) A step of providing an apparatus for rapidly and uniformly heating the liquid product to a predetermined temperature by resistance heating using a pulsed electric field;
(C) A step of supplying the liquid product to the inlet of the device and flowing the liquid product through the device;
(D) A step of continuously generating a pulse current in the liquid product flowing through the apparatus, with a pulse duration of at least 10 microseconds on each fluid element while a minimum of one pulse is passing. The step applied, wherein the electric field strength is 0.9 to less than 4 kV / cm;
Including
The maximum temperature of the liquid product remains autonomously between 40 ° C. and 85 ° C. during the resistance heating.
The above method. The method according to claim 1, wherein the pH of the liquid product is pH 1.5 to 9.0. The method according to claim 2, wherein the pH of the liquid product is pH 4.6 to 8.0. The method according to claim 2, wherein the pH of the liquid product is pH 1.5 to 4.6. The method according to any one of claims 1 to 4, wherein the liquid product has an electrical conductivity of 0.01 to 10 S / m as measured at 20 ° C. The method of claim 5, wherein the liquid product has an electrical conductivity of 0.2 S / m to 0.8 S / m as measured at 20 ° C. The method according to any one of claims 1 to 6, wherein the liquid product is a liquid food product or a liquid feed product. The method according to any one of claims 1 to 7, wherein the liquid product is a component, a semi-finished product, or a final liquid product. 8. The liquid product of claim 8, wherein the liquid product is selected from fruit juices, vegetable juices or smoothies, jams, spreads, alcoholic or non-alcoholic beverages, dairy products, vegetable milk products, liquid eggs, soups and sauces. Method. The method of claim 9 , wherein the dairy product is selected from milk, a milk product, or a liquid composition comprising a milk component or milk fraction. The method of any one of claims 1-10, wherein the temperature of the liquid product remains autonomously below 75 ° C. during the resistance heating. 11. The method of claim 11, wherein the temperature of the liquid product remains below 60 ° C. during the resistance heating. The method according to any one of claims 1 to 12, wherein the electric field strength is 2.5 to 3 kV / cm. The method according to any one of claims 1 to 13, wherein the pulse duration is at least 100 microseconds. The method according to any one of claims 1 to 14, wherein a bipolar pulse is applied. The method according to any one of claims 1 to 15, wherein the heated liquid product is cooled immediately after flowing through the apparatus. The method of any one of claims 1-16, wherein the liquid product is preheated to a temperature in the range of 20 ° C. to 70 ° C. before being fed to the apparatus. 17. The method of claim 17, wherein the liquid product is preheated to a temperature in the range of 35 ° C. to 65 ° C. before being fed to the apparatus. The method of any one of claims 1-18, wherein the number of microorganisms (cfu) in the heated liquid product is reduced by at least 5 log cfu / mL. The method of any one of claims 1-19, wherein the smallest single pulse applied to each fluid element during passage has a rectangular pulse shape.

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