Innovative Technologies for Food Preservation: Inactivation of Spoilage and Pathogenic Microorganisms

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Part I

Introduction

Outline

Chapter 1 Conventional Technologies of Food Preservation

Chapter 2 Innovative Technologies for Food Preservation

Chapter 3 Main Groups of Microorganisms of Relevance for Food Safety and Stability: General Aspects and Overall Description

Chapter 1

Conventional Technologies of Food Preservation

Pedro E.D. Augusto¹, Beatriz M.C. Soares² and Nanci Castanha¹,    ¹University of São Paulo (USP), Piracicaba, SP, Brazil,    ²Food Technology Institute (ITAL), Campinas, SP, Brazil

Abstract

The food microbiological stability is achieved by the reduction of the initial microbial load using a suitable procedure (generally a thermal process) followed by maintaining the residual microbial load to acceptable levels.

The factors that affect microbial growth are generally classified into two groups: intrinsic (i.e., of the food) and extrinsic (i.e., of the environment) factors. The main intrinsic factors of food are its water activity (aw), pH and acidity, redox potential (Eh), and the nutrient content. The main extrinsic factors are the temperature and atmospheric composition. The microbial growth will therefore be faster or slower depending on the composition of these factors in relation to the microbial requirements.

Therefore, understanding the factors that affect microbial inactivation and growth is essential to guarantee safe and stable products.

This chapter describes the main conventional technologies for food preservation, as well as their combination through the hurdle technology.

Keywords

Preservation; food preservation; conventional methods; conventional technologies; traditional technologies; food processing

1.1 Thermal Processing

The thermal process is one of the most widely used methods for food preservation, even considering its drawbacks and the development of further nonthermal technologies. It is a unit operation where the food is heated to a certain temperature, maintained for a certain time in order to promote the required microbial and/or enzymatic inactivation, and then cooled. It is generally characterized by a (applied) binomial time and process temperature (t×T), or, even better, an equivalent time at a specific temperature (for example through the F value concept).

The food preservation by thermal processing is based on the use of thermal energy (heat) for microbial and enzymatic inactivation, obtained due to protein denaturation and melting of lipid components, among other effects. However, although microbial and enzymatic inactivation is desirable, the thermal processing also results in other reactions (generally) undesirable, such as sensorial changes and nutrient destruction. The challenge, therefore, is to guarantee a safe and stable product (with the desired microbial and enzymatic inactivation), but with better sensorial and nutritional properties, with lower costs and energy saving consumption.

During processing, the food is heated by a hot fluid to the process temperature, following the heat transfer mechanisms. The food is then kept at this temperature for a certain time, previously calculated in order to optimize the product characteristics (i.e., safety assurance with better sensorial and nutritional attributes), called processing time. The food is then cooled by a cold fluid, interrupting the thermal effects.

The thermal processing is designed based on the process target, i.e., the most thermal-resistant undesirable microorganism or enzyme in the food product. The thermal process design will thus be calculated based on this target, ensuring safety and quality of the processed food.

The thermal process target can be a vegetative cell (as in beer or milk pasteurization), a microbial spore (as in the sterilization processes of low-acid foods—such as milk, corn, and tuna), a microbial toxin (as in the pasteurization of palm heart), or an enzyme (as some resistant pectinolytic enzymes in fruit products). The process target must be chosen aiming firstly food safety, but also considering the nutritional and sensorial characteristics of the final product and the economics.

1.1.1 Thermal Processing Main Characteristics

The main thermal processes used in food preservation are commonly described as commercial sterilization (usually referred as, simply, sterilization) and pasteurization.

The commercial sterilization is a more drastic thermal process, where the final product should have no vegetative cells and spores capable of growing under normal conditions of transportation and storage (Codex Alimentarius, 2003). Thus, in addition to safety, it is a process that ensures stability at ambient conditions (in general for periods in the order of months to some years).

It is a preserving method widely applied to low-acid foods such as milk, meat, corn, peas, carrots, and other vegetables that may be contaminated with spores of Clostridium botulinum, which represent a potential risk to the safety of the product and must be inactivated. Sterilized food is generally processed in pressurized systems, at temperatures of about 120–150°C for an appropriate time. The sterilized products show high stability and shelf life from months up to years. In this case, the final shelf life of the product is determined by physico-chemical and/or sensorial changes.

Pasteurization is a milder thermal process, whose main objectives are to ensure safety and prolong food shelf life. It is a method that needs to be used with a complementary technology (such as cooling, acidification, reduction of water activity, and/or use of preservatives) to guarantee stability. It is generally carried out under atmospheric pressure, with temperatures of about 65–100°C for an appropriate time.

Pasteurization is a process widely applied for the preservation of fruit juices, milks, fermented drinks, heart of palm, and other preserves, and its use implies the need for association with another preservation method. Pasteurized products have low to medium stability and variable shelf life, according to their characteristics and complementary conservation method. Their shelf life may vary from few days (as refrigerated pasteurized milk and other low-acid products with high aw) up to months or years (as juices and fruit jams and other high-acid products and/or with low aw).

Furthermore, the thermal processing of food can be performed inside or outside the package (Table 1.1).

Table 1.1

Main Characteristics of Food Thermal Processing Inside and Outside the Packaging

The in-package process is conducted with the product inside the packaging, i.e., the food is packaged in a hermetic package/container and then the product-packaging system is processed. In this way, the product does not come into contact with the environment after thermal processing (i.e., after the microbial inactivation). It is important to note that in this case, the packaging used must be resistant to the process conditions (temperature and pressure) to avoid deformations that compromise it. The thermal processing inside the packaging is often called Appertization, in honor of Nicolas Appert, a French confectioner who developed methods to preserve packaged foods due to thermal processing, winning a prize in 1810 stipulated by Napoleon Bonaparte for the development of a more efficient conservation method.

In the in-package thermal processes, a heating fluid is normally used and the heat is transferred to the packaging, then through the packaging, then from the packaging to the product, and finally across the product. After heating the product, it is maintained at the process temperature for a specified time and then being cooled. As the heat transfer is not instantaneous, each portion of food will have a different thermal history (T=f(t)) throughout the process. In this case, the process must be dimensioned for the region that the process resulted in a less lethal effect called cold spot or slower heating region (Fig. 1.1). In solid foods (conductive heating), the cold spot is typically located in the geometric center region of the product (for symmetrical packaging, for example, in the center of a cylindrical package). In liquid foods (convective heating), due to the natural convection currents formed during the process, such location is more complex, depending on several factors of the package and process (for example, the uniformity of heating) and product (mainly the rheological properties). However, it is in the lower portion of the packaging, generally between 10% and 20% of the height of a cylindrical package. In those cases, therefore, the guarantee of a safe product (process scaled to the cold spot) will result in regions with super-processed food (edges). Finally, considering that the heat transfer by conduction is less effective than by convection, many solid foods are thermal processed with a liquid portion, aiming to guarantee a faster and more uniform heating profile. Consequently the final products are more homogeneous and have lower gradient of undesirable reactions. These products are called particulate food and the main examples are vegetables in brine (corn, peas, pickles, heart of palm, and beans), tuna and sardine in oil or brine, and meat products added by sauces. In these cases, the liquid fraction is heated by convection, and flow through the spaces between the solid fractions results in more uniform heating of the product. However, the convection currents are slow (around some millimeters per second) and lower than the terminal velocity of the solid fraction; consequently, the fluid movement is not enough to move the solid fraction. Therefore the cold spot of the product is established for the worst case, i.e., at the geometrical center of the solid located at the region of slower fluid heating.

Figure 1.1 Representation of the cold spot location (black stars) during in-package food thermal processing using cylindrical cans (longitudinal section): (A) conductive food without headspace, (B) conductive food with headspace, (C) high consistency convective food, (D) low consistency convective food, (E) particulates food with high consistency fluid, and (F) particulates food with low consistency fluid.

When the thermal process is conducted outside the packaging, the product (usually in the liquid form) is processed in heat exchangers and its packaging must be previously sterilized (by physical or chemical methods). Then product and package are put together in an appropriate environment (aseptic).

Although the in-packaged processes result in higher gradient of reactions (such as microbial or enzymatic reactions, loss of nutrients, and physico-chemical reactions), as well as energy consumption (lower efficiency), they are safer as they are being less likely to post-process contamination. For this reason, it is still the most used method for guarantee of food safety and preservation.

The ohmic and microwave heating are two emerging technologies that can enhance the food heating, increasing the final product quality. In the ohmic heating technology, the product is heated by passing an electric current through it. Due to the low electrical conductivity of food, they act as resistors in the electrical circuit formed, dissipating the energy as heat (Joule effect) and heating. In the microwave heating technology, the electric field formed by the microwave radiation stimulates the movement of polar (water) and charged (ions) molecules. The movement of these molecules dissipates mechanical energy in the form of heat (Joule effect), heating the food. These two technologies are better described in Chapter 2, Innovative Technologies for Food Preservation.

1.1.2 Microbial Inactivation Kinetics

As other chemical and biochemical reactions, the microbial inactivation rate has an exponential relationship with the temperature. In most cases, especially for bacteria, the inactivation can be described by a first-order kinetics (Eq. 1.1). This kinetics shows that the microbial reduction rate at a fixed temperature (T) is a function of the microbial load (C) in the food at a determined moment. It means that the relative microbial reduction (e.g., in percentage) is always equal for same time intervals (Eq. 1.1). By developing Eq. (1.1), Eq. (1.2) is obtained, relating the microbial inactivation with the first-order kinetics constant at that temperature (kT) as a function of processing time (t):

(1.1)

(1.2)

Traditionally, microbial inactivation is expressed by the decimal reduction time (DT) and defined as the time required, at a fixed temperature, to reduce one logarithmic cycle inactivation (90%) of the microbial load. This parameter has a direct relationship with the constant of microbial inactivation rate (kT), expressed by Eq. (1.3). Then, using Eqs. (1.2) and (1.3), and the logarithm in the base 10 (log10=log), Eq. (1.4) is obtained:

(1.3)

(1.4)

Further, the DT values obtained at different temperatures decrease at each increment of temperature, following a similar trend and then being correlated by the thermal coefficient (z), which represents the "temperature difference required to promote a reduction of one logarithmic cycle (90%) in the DT values." The z value can thus be expressed according to the following equation:

(1.5)

The concepts of DT and z can be easily understood by evaluating the curve of microbial inactivation (that correlates the microbial load as a function of the processing time (t) at a fixed temperature (T)) and the DT values variation with the temperature (T). When the curves are obtained using logarithmic scale (Fig. 1.2), the DT and z values can be clearly identified.

Figure 1.2 Graphic representation of the DT and z concepts using hypothetical microbial inactivation curves fixed temperatures (T, T1, T2, T3).

However, in some cases, the microbial inactivation does not follow the first-order kinetics (Eqs. 1.1–1.4) and other complex models are required to describe it (Peleg, 1999, 2006). The Weibull model is the nonlinear model most used to describe the microbial inactivation (Eq. 1.6). This kinetics is defined by two parameters, b and n, both functions of the temperature (i.e., b(T)=bT and n(T)=nT).

(1.6)

The b is the model proportional parameter, which represents the microbial resistance to the thermal process. As higher the b value is, the less resistant is the microorganism. The parameter n is related to the curve shape. When n=1, the microbial inactivation follows the first-order kinetics (Fig. 1.1). When n<1, the curve has an upward concave shape, showing that part of microbial population is thermal resistant and also that only a maximum inactivation can be reached. It is often called as tailing behavior. On the other hand, when n>1, the curve has downward concave shape, highlighting an initial resistance to inactivation process, and can also indicate that the continued exposure results in accumulated damage, reducing the survival thermal resistance. It is often called as shoulder behavior. Both tailing and shoulder behavior are often related not only to the difference on the natural resistance distribution in the population, but also to mixed cultures and population with different cell conditions (as at different growth phases). Moreover, the microbial behavior can be a function of other environment conditions (such as pH, pressure, and food composition), and can also change with the temperature (as, for example, inverting the concavity).

Considering that the first-order kinetics is well applied in many cases, the additional information of this chapter is based mainly on the first-order kinetics.

1.1.3 Process Design

During thermal processing, each part of the food has a specific temperature at each time. For this reason, the thermal processes cannot be described only by its temperature or process time, and should be characterized by both parameters associated with others that influence the heat transfer phenomena (type of equipment, packaging, product characteristics, etc.), and, more correctly, by an equivalent time—as the sterilization value (F that describes the total lethal effect of the process).

The thermal processing design must take into consideration the heating and cooling heat transfer characteristics, for the product and through the product.

The thermal processing is carried out to reach an appropriate decimal reduction (γ, Eq. 1.7) of the processing target. This considers the initial concentration of the target on the food (C0) and the final concentration required (Cf):

(1.7)

The initial concentration of the target microorganism on the food (C0) is determined by the raw material history, being necessary to keep it as lower as possible (for example, by following all the good practices). The required final concentration (Cf) can be defined according to literature data or laws and regulations, aiming to guarantee the food safety and stability.

Considering the required γ and the results of heat penetration tests (i.e., the thermal history of a determined point inside the food, obtained by experiments using thermocouples and data logger, following exactly the same condition of processing), the binomial time versus temperature (t×T) can be defined to reach the process lethality. The process temperature is determined based on the microbial resistances, nutritional and sensory food characteristics, and equipment and physical limitations. The process time is determined considering the inactivation at the most difficult case (the slowest heating point), i.e., the cold spot. The process time is usually designed considering only the heating and retention steps; the cooling step is considered as a safety margin.

However, a thermal process cannot be only characterized by its t×T binomial, since the same binomial can result in different decimal reductions due to the food characteristics (physical properties, heat transfer by convection or conduction, dimensions, packages, etc.), heat exchange media (convective heat transfer coefficient—h, contact area), and the target characteristics (DT e z for the evaluated food). Therefore the sterilization value (F—Eq. 1.8) is the best way to characterize the food thermal processing.

The sterilization value (F) represents the equivalent time (in minutes, for example), at the reference temperature (Tref), that the food is submitted during processing. It is, in general, calculated at the cold spot location, from Eq. (1.8) and through Eq. (1.9). The F value can be obtained by monitoring the cold spot thermal history, acquiring temperature data at short time intervals (Δt; as smaller as possible; ≈1–5 s for convective foods or ≈10–60 s for conductive foods). Therefore, applying the trapezoidal rule for solving Eq. (1.9), the F value can be determined using Eq. (1.9):

(1.8)

(1.9)

Therefore the sterilization values associated to the process can be determined using heat penetration studies. Using this data, the time required is established at the process temperature that guarantees the desired decimal reduction (γ).

Considering the importance of C. botulinum for the thermal processing of foods, the thermal resistance of its spores is commonly used to express the sterilization process to ensure minimal safety for commercialization. Using the temperature of 121.1°C and C. botulinum heat resistance, the calculated F of the process (Fp) is then called F0. The F0 value is calculated based on the values of D121°C=0.21 min and minimum reduction of 12 logarithmic cycles (γ=12). Using Eq. (1.8), the minimum F0 for food processing is 2.52 min. However, for safety reasons, (much) higher values of F0 are applied.

It is important to highlight that in some cases, the final process will be considerably more drastic than the required to guarantee the safety of the product. This occurs, e.g., for meat products, where a more severe process is needed to guarantee the correct product cooking.

Further information can be explored at Augusto (2017), Augusto, Tribst, and Cristianini (2014), Fellows (2006), Ordóñez (2005), Pflug (1988), Potter and Hotchkiss (2007), Ramaswamy and Marcotte (2006), Stumbo (1973), and Teixeira (2006).

1.2 Cooling

The cooling/chilling process is widely used in the food industry for ensuring the shelf life extension of products with lower sensory and nutritional changes. It is a unit operation where the food is cooled to a certain temperature and maintained under these conditions throughout its storage, generally used in association with other preservation processes (with the exception of raw materials).

Temperature is a physical measure that relates the internal energy of the system, i.e., the molecular energy level, being directly proportional to their level of agitation and vibration. At lower temperatures, therefore, the molecular energy level is smaller, as will be slower the reactions involving them.

In the cooling process, the food is cooled and stored at temperatures above those necessary for freezing, keeping the water in the liquid state. Thus, although slowly, the physico-chemical, microbiological, and biochemical reactions continue to occur in refrigerated foods.

The reduction in the rate of most reactions can be associated with the concept of Q10 (Eq. 1.10), which defines the variation in the rate of the reactions by changing the temperature at 10°C, describing an exponential reduction in the reactions with decreasing temperature (Fig. 1.3). For most of the reactions involved with food the Q10 value is between 2 and 5, i.e., by reducing the temperature by 10°C, a reduction of ≈2 and 5 times is observed in the rate of the reactions. This concept explains the longer shelf life of refrigerated products:

(1.10)

Figure 1.3 Q10 concept to describe the rate of reaction as a function of temperature (left); enzymatic activity in relation to temperature (right).

However, the behavior of enzymatic and microbial reactions are more complex and cannot be explained only by the Q10 concept.

Fig. 1.3 shows the typical behavior of an enzyme as a function of temperature. Although the concept of Q10 may explain its activity in a certain temperature range, the denaturation occurs due to maintenance at higher temperatures than the optimum result in reduced enzymatic activity.

Similar behavior is observed for microbial growth (since the microbial metabolism is dependent on various enzymatic functions), but with one important difference: each microorganism has a minimum temperature of development, below which it remains in a latent state. Table 1.2 and Fig. 1.4 show the optimal and minimum temperatures of development for the main groups of microorganisms.

Table 1.2

Optimal and Minimum Temperatures of Microbial Growth (Leitão, Hagler, Hagler, & Menezes, 1988)

Figure 1.4 Microbial growth rate (MGR) as a function of temperature.

Thus, in general, the cooling can not only reduce the microbial growth rate, but also prevent the development of certain microorganisms, acting as a selective method. However, it is important to note that this process does not result in microbial inactivation, and, therefore, must be associated with another preservation method for safety assurance in consumption. The thermal pasteurization processes, which result in microbial inactivation, are frequently used with cooling.

Another important factor that should be considered is the thermal history of a product during its refrigerated storage (Fig. 1.5), since frigorific system differs from the ideal ones. Although cooling chambers have heat-insulating coating, such insulation is not perfect, so there is a permanent flow of heat from the neighborhood (hottest) to the system. Thus, when the product is at the desired temperature (or setpoint—SP), it exchanges heat with the environment and is heated. When the product temperature reaches an upper limit (Tup), the refrigeration system is switched on and its temperature begins to descend to a lower limit (Tlow), where the system is shut down. From that moment, the product is heated again by exchanging heat with the neighborhood, in a cyclical behavior.

Figure 1.5 Ideal (A) and real (B) thermal history of a product during refrigerated storage.

Therefore it is important to note that the chamber temperature (and consequently the product temperature) varies continuously along the storage and that the refrigeration system design should be done aiming to minimize the extent of this fluctuation (i.e., Tup and Tlow should be as close as possible from SP), since lower temperatures may result in freezing of the product, while higher temperatures may allow microbial growth.

Further information can be explored at Augusto (2017), Fellows (2006), Ordóñez (2005), and Potter and Hotchkiss (2007).

1.3 Freezing

The freezing process is a unit operation where latent heat is removed from the food, with consequent solidification of the water (Figs. 1.6–1.8). The change in the physical state of the water defines freezing as a very effective method of conservation, being more than a super cooling.

Figure 1.6 Schematic representation of refrigeration and freezing processes (RRR, relative rate of reactions).

Figure 1.7 Frozen water fraction as a function of temperature and freezing thermal history in food and in pure water (constant atmosphere pressure, equilibrium).

Figure 1.8 Schematic representation of the fast and slow freezing processes (black stars represent the ice crystals, while dashed arrows indicate the movement of liquid water).

By solidifying the water, the freezing process drastically reduces the rate of reaction by reducing the molecular mobility and by hindering the interaction between reagents (Reid & Fennema, 2008). Also, the immobilization of water causes a solid concentration in the unfrozen fraction, resulting in reduction on the water activity (aw) in that region (Fellows, 2006; Reid & Fennema, 2008; Strasburg, Xiong, & Chiang, 2008).

Due to the high mobility restrictions, some reactions may even be interrupted, as the microbial growth, for example. When food is frozen, the microorganisms enter into latency state, not, as a general rule, inactivated. Thus, when the microbial load reduction is required, another method of preservation should be used for safety assurance in consumption. Once again the thermal pasteurization processes are frequently used in conjunction with freezing, since it results in microbial inactivation.

However, reactions that do not depend on the water still remain at a relatively high rate even during freezing (Fig. 1.6), as in the case of lipid oxidation. It is observed thereby that oxidation of lipids limits the shelf life of most frozen products.

The safety and quality of frozen foods are strongly influenced by the freezing process, frozen storage, and thawing process.

Although the phase transition is a well-defined phenomenon for pure substances, it is not for solutions, suspensions, emulsions, and complex products, such as foods. For example, the freezing process of pure water and food is shown in Fig. 1.7. For pure water at atmospheric pressure, a decrease in its temperature is observed when the heat is removed from it, until the temperature of 0°C (i.e., the thermal energy is removed in the form of sensible heat). At this point, any amount of energy removed will result in change in physical state of the corresponding amount of water molecules, which will go from liquid to solid, without the temperature change (i.e., thermal energy is removed in the form of latent heat). If further energy is removed after complete freezing, the ice is then cooled (i.e., thermal energy is removed as sensible heat). In the case of pure substances therefore the freezing temperature is well defined (for a given pressure). On the other hand, the freezing of complex products is not defined by a marked change in the physical state of liquid to solid water at a fixed temperature. Instead, because of the colligative properties of the solute and water, the freezing starts at temperature slightly lower than 0°C (generally between −1°C and −5°C). From this condition, if more energy is removed from the food, its behavior is described by a continuous progression of cooling–freezing (Fig. 1.7). As is the pure water what is frozen in the process, the concentration of the solution (unfrozen fraction) increases when part of the water molecules passes from liquid to solid, reducing the freezing point. Consequently, it is impossible to dissociate the phase shift and the cooling in the freezing curve, as is clearly shown in Fig. 1.7. As a result, there is not a fixed freezing temperature (even for the same pressure), but a temperature below which there will always be frozen and unfrozen water fractions.

Another important characteristic of the freezing process is related to the thermodynamic properties of water and food. Unlike the vast majority of substances, the molar volume of solid water is higher than the molar volume of liquid water (which also results in lower density of ice compared to liquid water). As a consequence, the water forms sharp crystals and presents a volume expansion of about 9% during