Effect of High-Powered Ultrasound on Bioactive Compounds and Microbiological Stability of Juices—Review

Abstract

Extending the shelf life of fruit and vegetable juices plays a key role in improving food security. Ultrasonic technology can be an effective method in the process of replacing traditional heat treatment. It offers a number of benefits, such as better product quality expressed as a higher content of bioactive substances and the preservation of the natural sensory characteristics of the juice: consistency, colour, taste and smell. This paper discusses the methods of ultrasound generation, the equipment used and advances in the construction of ultrasound devices. The influence of ultrasounds on the physical and chemical properties of juices was also analysed, with particular emphasis on bioactive substances and the effectiveness of ultrasonic treatment in the inactivation of microorganisms and enzymes. Prospects and trends in the development of ultrasonic techniques that can effectively increase the efficiency of the ultrasonic juice preservation process are also described. Sonication used alone or with other processing techniques makes it possible to achieve a positive effect on the quality of consumed fruit and vegetable juices.

1. Introduction

Ultrasound has various applications in food processing due to its ability to induce physical, chemical and mechanical changes in food materials. Ultrasound utilizes distinctive physical and chemical phenomena that stand in contrast to those employed in traditional extraction, processing or preservation methods. Ultrasound presents a clear edge in aspects of efficiency, output and specificity, showcasing improved processing durations, heightened quality and decreased chemical and physical risks, all while remaining ecologically conscious [1].

The basic method used in traditional food preservation is heat treatment, which destroys dangerous pathogens or limits the growth of microorganisms [2]. Thermal processing, despite its high effectiveness in inactivating microorganisms, also brings with it undesirable changes that adversely affect the final quality of the processed food. Many food ingredients are thermolabile and decompose when exposed to heat. In addition, heat treatment can also cause adverse changes in the taste, colour and even texture of the processed food. This gave rise to the need to search for new alternative methods of food preservation, which undoubtedly include power ultrasound.

Power ultrasound refers to the use of high-intensity sound waves with a range of 10 to 1000 W/cm² and frequency ranging from 20 to 100 kHz. They induce various physical and chemical effects in liquids, solids and gases. Power ultrasound is characterized by the application of intense acoustic energy, often leading to the phenomenon of cavitation, which involves the creation and collapse of tiny bubbles in the material being treated. These collapses generate locally high temperatures and pressures, leading to mechanical and chemical effects that can be harnessed for different applications.

Ultrasound technology has found applications in juice preservation, offering several benefits in terms of extending shelf life, maintaining nutritional quality and minimizing the need for chemical preservatives. However, it is important to note that ultrasound still has some flows resulting in degradation of primary and secondary metabolites [3]. Therefore, its application in juice preservation should be optimized to achieve the desired effects without negatively impacting sensory attributes or causing undesirable changes.

Different reviews have explored the utilization of ultrasound technology in the preservation and processing of food products [4,5,6,7]. However, a comprehensive review addressing the specific application of power ultrasound in juice preservation has not been previously presented. This article provides a concise overview of the impact of ultrasound on the physical and chemical properties of juices, their microbiological stability and further prospects for using this type of treatment in juice preservation.

2. Ultrasound Generation

Devices for generating ultrasonic waves consist of three basic elements:

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An electrical power generator;

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A transducer(s);

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An emitter.

The electric generator is the source of energy for the ultrasonic system, which must supply the transducer. In general, the electric generator produces an electric current of a certain rated power. Its task is to convert alternating current with a frequency of 50 Hz or 60 Hz into alternating current with the desired frequency, usually from 20 kHz to 40 kHz. Most generators allow the power to be set indirectly by setting the voltage (V) and current (I). The transducer is the central element of each ultrasonic device, and its role is to generate ultrasonic waves. The transducer converts electrical energy into mechanical vibrations at ultrasonic frequencies. The transducer converts, for example, 20 kHz electrical energy from the generator into 20 kHz mechanical vibrations. The aim of the emitter, called a reactor by some and an ultrasonic cell by others, is to radiate an ultrasonic wave from the transducer to the medium. Emitters can also act as an amplification of ultrasonic vibrations during their emission. The two primary types of lab apparatus in the food industry are:

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An ultrasonic cleaning bath;

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A probe system.

Ultrasonic cleaning bath

The ultrasonic cleaner, also known as an ultrasonic bath, is a special device used for cleaning and sanitation in beverage processing operations. It consists of the following elements (Figure 1):

Tank: This is the main part of the ultrasonic cleaner, and is usually made of stainless steel or another chemical-resistant material. The tank has the shape of a bathtub and serves as a working chamber in which the sonication process is carried out. It is available in various sizes to accommodate different object sizes.

Power source with an electric generator: This converts an electric current of typical frequency into an electric current of ultrasonic frequency and delivers it to the transducer.

Transducer: This is the heart of the system. This element converts electrical energy into ultrasonic waves. This is the piezoelectric element attached to the bottom or sides of the tank. The transducer generates high-frequency ultrasonic waves, which create alternating high-pressure and low-pressure zones in the liquid.

Heater: Some ultrasonic baths include a heating element to warm the liquid. This maintains the cleaning solution at an elevated temperature, which can enhance the effectivity of process for certain applications.

Timer: most ultrasonic baths come equipped with a timer that allows the duration of the cleaning cycle to be set.

Drain valve: a drain valve is positioned at the bottom of the tank for easy removal of used liquid once the treatment cycle is complete.

Lid: many models come with a removable or hinged lid to prevent solution evaporation and to reduce noise generated by the ultrasonic waves.

Fan: A cooling fan may be installed on some models to prevent overheating during extended work cycles.

Control panel: on the control panel, the user can adjust the settings of the washer, such as time, treatment temperature and other options.

Advances in ultrasonic bath technology

In the construction and operation of ultrasonic cleaners, several new trends and innovations can be noticed, which are aimed at improving efficiency, precision and comfort of use. Additional features include, among others:

(a)

The ability to choose the frequency of ultrasonic vibrations of two 25/45 or 35/130 kHz, or even three 45-80-100 [8];

(b)

Work at two different ultrasound frequencies simultaneously. The ultrasonic cleaner comprises two types of ultrasonic transducers operating at two equal resonance frequencies, for example, 25 and 40 kHz, which are attached on the bottom and the side, respectively. The result is uniform ultrasonic power distribution [9];

(c)

Power regulation: the ultrasonic power ranges from 10 to 100% of the total power;

(d)

Different working modes:

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Sweep mode—ultrasonic cleaners have several ultrasonic transducers, which, despite the high precision of workmanship, differ in resonant frequency. Therefore, if the generator emits a constant-frequency electric current, some transducers will not work at the resonant frequency, resulting in poor efficiency and rapid overheating. To avoid this, the generator is equipped with a special sweep mode that determines the effective fundamental centre frequency and the range of frequency variation that allows each transducer to achieve resonance. Thanks to these measures, even if for a short time, all transducers operate at their own resonant frequency, which allows for optimization of the energy efficiency of the entire assembly and even distribution of the load on all elements [10].

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Full-wave mode—this is the standard single-frequency ultrasonic treatment mode for all transducers;

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Burst mode—this involves generating periods of power 50–80% higher than normal power followed by equal periods of zero power;

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Degas mode—the intensity of the ultrasound is changed over time to speed up the process of degassing the liquid;

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Impulse mode.

The ultrasonic probe

An ultrasonic probe system, also known as an ultrasonic horn or ultrasonic probe, is a specialized piece of equipment used for applying ultrasonic vibrations directly to a sample. It consists of the following items (Figure 2):

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Generator—the generator produces electrical signals of a specific frequency that are sent to the ultrasonic transducer. The most common frequency is 20 kHz, but other frequencies are also available;

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Ultrasonic converter—this converts electrical energy into mechanical vibrations of fixed frequencies, usually 20 kHz;

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Probe—this transmits ultrasonic energy into a sample;

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Booster (optional)—this is used to increase the intensity of the ultrasonic vibrations.

There are two primary distinctions between an ultrasonic probe and an ultrasonic bath. Firstly, an ultrasonic probe is submerged directly into the solution, while in an ultrasonic bath, the liquid has no direct contact with the ultrasonic emitter and takes the shape of the tank into which it was poured. Secondly, an ultrasonic probe supplies much higher levels of ultrasonic energy than an ultrasonic bath (sometimes even 100 times higher). These significant variations dictate the specific applications of each system. An ultrasonic probe system offers several advantages that make it a valuable tool for various applications. A probe system enables precise targeting and is flexible in use. The probe’s tip can be positioned directly at the point of interest, enabling focused treatment and minimizing the impact on surrounding areas. Ultrasonic probes come in various sizes and shapes, allowing for flexibility in adapting to different sample types and volumes [11].

Another advantage is the possibility of obtaining high ultrasound intensity. This higher intensity allows for more efficient and effective sample processing, especially for challenging tasks such as cell disruption or particle size reduction. Due to its higher energy output, an ultrasonic probe system can achieve desired results in a shorter time. This is particularly advantageous for time-sensitive processes or situations where reducing the exposure of sensitive samples to ultrasonic energy is important. Ultrasonic probes are particularly suitable for small-scale processing, making them ideal for research labs, benchtop setups and applications where larger equipment is not necessary. Another advantage of this system is ease of use. Operating an ultrasonic probe system can be straightforward, especially when using modern systems equipped with user-friendly controls and displays.

Despite these advantages, it is important to note that ultrasonic probe systems also have limitations, such as potential sample heating with prolonged use and the need for careful handling to prevent probe damage. Choosing the right system depends on the specific application requirements and the desired outcomes.

Advances in ultrasonic probe technology

The probe system is constantly being developed and improved, as exemplified by the following trends.

New materials from which tips are made. Currently, tips made of silica glass appear on the market. These probes do not contaminate samples with metal particles, which makes them particularly useful during trace analysis in food and pharmaceutical analysis. Silica glass has a high melting point and is resistant to thermal shock. Silica glass has a relatively low coefficient of thermal expansion, which means that it undergoes minimal dimensional changes under the influence of temperature fluctuations. Silica glass is chemically inert and does not react with most substances. This property makes it valuable for use in corrosive or reactive environments. On the other hand, these types of probes have limited strength and therefore can be used at much lower ultrasonic intensities compared to metal probes [12].

New probe shape. The latest probes are spiral-shaped, and, unlike traditional probes, the ultrasonic power is distributed across the entire surface of a spiral probe (lateral irradiation). They are used in slim reaction vessels [12].

Multiples probes. Multi-element probes increase productivity and minimize repetitive tasks by processing numerous samples simultaneously. Units are available with 4, 8, 16 and 24 tips [13].

Cup horns. Cup horns offer indirect sonication, which means the ultrasonic waves pass through the wall of the sample container. A cup horn can process multiple sealed tubes or vessels at one time without contacting an ultrasonic probe. This method eliminates cross-contamination, sample foaming, overheating and aerosolization, which can all occur when using a probe. Most importantly, a cup horn enables samples under 200μL to be effectively processed [13].

Flow cells. They enable continuous processing of liquid. The process fluid enters through the inlet at the bottom of the unit. As it passes through the cavitation field, it is ultrasonically processed. The fluid exits the unit through an outlet port. The intensity and extent of processing are controlled by adjusting both the flow rate and amplitude setting [13].

3. The Influence of Ultrasound on the Physical and Chemical Properties of Juices

Consumption of fruit, vegetables or fruit and vegetable juices significantly increases ingestion of microelements, mineral salts and vitamins, as well as antioxidants and fibre, which are invaluable in the proper functioning of the body.

Many studies have shown that the substances contained in juices are absorbed much more efficiently by the human body than those that are supplied through various types of preparations (dietary supplements). Therefore, consuming juices that have unique compositions is recommended [14,15]. Experts recommend that you try to consume juices more often than vitamin-mineral tablets. Regular consumption of fruit and vegetable juices allows you to supplement deficiencies of vitamins and bioelements. All the substances contained in the juice are delivered in the easiest form for cells to absorb; additionally, they do not burden the liver or the digestive system and are distributed throughout the body in an extremely fast time [16,17].

The quality and safety of juices are extremely important for the health of consumers. With the increase in public awareness of the importance of nutrition, more and more emphasis is being placed on producers, because they are responsible for providing the highest quality of products. At the same time, they cannot forget about meeting the highest food safety standards [18,19].

Undoubtedly, the raw materials used for the production of juices must be of a high quality. The healthiest are drinks made from fresh fruit and vegetables from controlled crops or organic sources. It is important not to use synthetic substances (preservatives or dyes) that may affect the final quality of the products [20,21]. Careful observance of production processes allows for the preservation of the richness of vitamins, minerals and other nutrients present in raw materials [22,23,24].

It is also very important to adhere to the highest standards of hygiene throughout the production process to avoid contamination and the growth of harmful microorganisms. The use of appropriate pasteurization technologies or other preservation methods is crucial to eliminate potential health risks [25,26].

Certifications such as HACCP (Hazard Analysis and Critical Control Points) or ISO (the International Organization for Standardization) are proof that the manufacturer applies rigorous procedures and complies with food quality and safety requirements [27,28]. Consumers should choose juices with appropriate certificates to be sure that the product meets high standards.

In addition, information on the packaging, such as the expiration date, ingredients, nutritional values and storage recommendations, helps consumers make the right purchasing and best-before decisions.

However, in order for the juices to be safe and fit for consumption for a longer period of time, they must be properly preserved. Heating treatment methods will, however, affect the physical and chemical properties of these types of products to some extent. Therefore, new ways of preserving beverages have recently been sought, including the use of high hydrostatic pressure (HPP), pulsed electric field (PEF) or the increasingly popular ultrasound (US) [29,30].

There are many studies on the effect of the sonication process on the physical and chemical properties of juices.

For example, Gupta et al. [31] carried out sonication of pomelo juice. The scientists examined the chemical composition of the obtained product. In addition, they verified flavour compounds, including octanal, linalool, citral and ethyl butyrate, and as the analyses show, these substances were significantly enhanced, i.e., the taste of the juice was intensified. The hydrolytic activity of rhamnosidase and glucosidase was increased by the use of sonication, which resulted in a lower level of bitterness in pomelo juice. Moreover, the results of the research indicate that properly set parameters of the ultrasonic process in cooperation with degassing agents lead to a significant change in the content of naringin in pomelo juice. And as it is well known, naringin is the main flavonoid, having a wide range of pharmacological activity, including increasing lipolysis, lowering cholesterol and anti-inflammatory, anticancer and antiseptic effects [32,33,34,35].

Improving the effectiveness of new cost-effective technologies in preserving the overall quality characteristics and antioxidant properties of fruit juices is of great interest. The team of Margean et al. [36] examined the effect of pasteurization (80 °C, 2 min) and high-power ultrasonic treatment (50% amplitude—57.5 µm and 70% amplitude—80.5 µm for 5 and 10 min) on the quality of red grape juice. The researchers analysed the processed juice in terms of physicochemical aspects, with particular emphasis on the content of phenolic compounds or L-ascorbic acid. In addition, the authors verified the samples in terms of microbiology. The results of the analyses showed that the samples subjected to pasteurization and sonication for 10 min with an amplitude of 70% had a similar level of total phenol content. On the other hand, the sonicated juice was distinguished by a higher content of L-ascorbic acid compared with the thermally preserved juice. Similar results were obtained for pH, total soluble solids and titratable acidity. The conducted analyses also indicated that the US influenced the inactivation of microorganisms, which may be an alternative method to traditional pasteurization in the juice industry. In addition, it has been proven that, compared to pasteurization, ultrasonic treatments with an amplitude of 70% were more effective than those with an amplitude of 50% in terms of inactivating the microorganisms concerned. In general, the authors of this study determined that to achieve higher levels of flavonoids, such as rutin, quercetin, epicatechins, kaempferol and resveratrol, treatment with an amplitude of 70% for 5 min is recommended. On the other hand, the ultrasonic process conducted at an amplitude of 70% for 10 min turned out to be the most effective in reducing microorganisms and achieving the highest content of L-ascorbic acid and total polyphenols, including hydroxycinnamic acids such as ferulic acid and caffeic acid.

The analyses carried out by Yıkmış [37] present similar results and relationships as in the previously described studies. However, in this case, the aim of the experiment was to examine the effect of ultrasonic treatment (4, 8, 12 and 16 min) and pasteurization on pH, titratable acidity, Brix, colour, lycopene, ascorbic acid, total phenol concentration, total flavonoid concentration, antioxidant activity, sensory properties and microbiological safety in red watermelon juice and yellow watermelon juice. And as the research showed, the Brix values and titratable acidity did not change statistically significantly after ultrasonic treatment (p > 0.05). Interestingly, there was an increase in lycopene content, phenol concentration, flavonoid concentration and antioxidant capacity after 16 min of US treatment. The applied ultrasonic process had an unfavourable effect on the concentration of ascorbic acid, reducing its content in comparison to control samples (not subjected to any treatment). However, these losses were not as high as in the case of pasteurization. Importantly, the organoleptic evaluation showed that the juices after ultrasonic treatment did not lose their sensory attractiveness.

The growing consumer demand for healthy, natural food products prompts scientists and producers to look for new preservation methods that will preserve the high quality and nutritional value of food, including juices. The positive impact of ultrasonic treatment was presented by Ruiz-De Anda et al. [38] in their study, the aim of which was to determine the effect of ultrasound on the physicochemical properties of two types of juices—orange and celery stalk juices. The juice samples were sonicated in the so-called ultrasonic bath (20 kHz, 20 ± 5 °C) for 0, 15, 30, 45, 60, 75 and 90 min. Thereafter, the articles were stored in the dark for 24 h at 4 °C. After this time, the juices were analysed, showing that the ultrasonic treatment had a significant (p < 0.05) effect on the number of bioactive compounds and antioxidant properties of the juices as early as 15 min into the process. The best results were achieved for 45 min of US treatment, where a 13% increase in the total phenol content, a two-fold increase in the content of flavonoids and a 17% increase in antioxidant activity were observed. The method of processing did not affect some physicochemical properties of juices, such as total soluble solids, pH and titratable acidity. Changes in colour after ultrasonic treatment were observed, showing significant correlations with the increase in the content of bioactive compounds and antioxidant activity. To sum up, properly conducted ultrasonic treatment provides the opportunity to obtain juices with increased health-promoting values.

In contrast, a study by Yıkmış et al. [37] showed that sonication, compared to other unconventional methods, still needs to be improved. The researchers’ analyses were aimed at assessing the impact of the use of high-pressure processing (HPP), ultrasound (US) and pulsed electric field (PEF) on changes in the physicochemical and phytochemical properties of strawberry juice. HPP processing at 300 MPa (1 min), US at 55 °C (3 min) and PEF at 35 kV/cm (27 μs) were compared to traditional thermal pasteurization (72 °C, 15 s). No significant differences were observed in total solubles (7.83–8.00°Brix), titratable acidity (0.79–0.84 g/100 mL) or pH (3.45–3.50; except for sonication) between treated and untreated samples (p > 0.05). In contrast, HPP and PEF technologies significantly increased the retention of total TPC phenol content (4–5%), increased TAC total anthocyanin content (15–17%) and RSA free radical scavenging activity (18–19%) compared to unprocessed strawberry juice samples. In the case of the US process, the positive effects of the research were not so noticeable. No changes in TPC were observed, and the content of TAC and RSA increased by less than 9% and 14%, respectively, compared to the control juice. Hence, the conclusion was that the samples treated with HPP and PEF had similar properties in terms of the content of phytochemicals, which was higher in these samples than in sonicated, thermally pasteurized and untreated samples. More research should be carried out, and different parameters of ultrasonic treatment in juice processing should be used to obtain satisfactory analysis results.

Santhirasegaram et al. [39] compared the quality of freshly prepared mango juice after sonication and heat treatment. The product samples were sonicated for 15, 30 and 60 min at 25 °C, 40 kHz (using an ultrasonic cleaning bath: Branson Model 3510 Ultrasonic Cleaner, CT, USA). Pasteurization was carried out in a covered water bath (Memmert, Germany) at a temperature of 90 ± 1 °C for 30 s (mild heat pasteurization) and 90 ± 1 °C for 60 s (high-heat pasteurization). The treatments used did not result in any significant changes in pH, total soluble solids or titratable acidity. Interestingly, sonication lasting 15 and 30 min resulted in a significant improvement in selected quality parameters (increase in the extraction of carotenoids and polyphenols), except for the colour and content of ascorbic acid, compared to freshly squeezed (control) juice. In addition, an increase in radical scavenging activity was observed in all sonicated juice samples regardless of the treatment time. The obtained results support the use of sonication to improve the quality of juices as an alternative to heat treatment.

Table 1. Summary of various analyses on the impact of ultrasound on the physical and chemical properties of various types of juices from the last 5 years.

Type of Juice	Type of Ultrasonic Generator	Process Parameters	Physical and Chemical Properties Discussed	Sonication Effect	References
Carrot–grape juice	Ultrasonic processor (UP400S, Hielscher Ultrasonics GmbH Hielscher USA, Inc.) with 0.5 in. probe at 2 in. depth of the sample	Amount of juice: 250 mL; Frequency: 20 kHz; Duration of sonication: 5 min; Processing temperature: 15 °C.	- Total soluble solids (°Brix); - Determination of pH; - Determination of total phenolics; - Determination of total flavonoids; - Determination of DPPH free radical scavenging activity; - Determination of total antioxidant capacity (TAC); - Reducing power ability.	The results show that sonication does not affect the pH of the juice. It was also observed that the sonication increased the extract content of the juice. Furthermore, sonication had a positive effect on the bioactive compounds, antioxidant activity and reducing power of the juice compared to chemical preservation methods.	Nadeem M. et al. [40] (2018)
Açai (Euterpe oleracea) juice	Ultrasonic processor (model DES500 500W, Unique, Brazil) with a probe 1.3 cm in diameter	Amount of juice: 100 mL; Frequency: 19 kHz; Duration of sonication: 5 min; Processing temperature: 32 ± 1.2 °C.	- pH and titratable acidity; - Cloud value; - Non-enzymatic browning; - Viscosity; - Antioxidant activity (DPPH and ABTS); - Phenolics compounds; - Total anthocyanins; - Peroxidase; - Polyphenoloxidase activity.	The pH and titratable acidity did not change with ultrasound. The cloud values increased with increasing ultrasound energy density. Moreover, the combination of ultrasound together with the application of ozone affected the viscosity of the juices and affected the bioactive compound values, such as antioxidant activity (DPPH and ABST), phenolic compounds and anthocyanin content.	Oliveira et al. [41] (2018)
Guava juice	Ultrasonic processor (ECOSONIC, QR1000 Model, Brazil) with 1.26 cm2 titanium tip	Amount of juice: 200 mL; Frequency: 20 kHz; Duration of sonication: 3, 6 and 9 min; Processing temperature: 25 °C.	- Microstructure; - Pulp sedimentation; - Turbidity (serum cloudiness); - Colour; - Carotenoid extraction; - Identification and quantification of carotenoids via high-performance liquid chromatography (HPLC); - Carotenoid in vitro accessibility.	Ultrasound improved both the in vitro availability of lycopene and the physical properties of guava juice. By reducing the particle size of the pulp, the stability of the physical properties of guava juice was increased.	Campoli S. et al. [42] (2018)
Bayberry juice	Ultrasonic processor (BILON-600Y, Bilon Co., Ltd. Shanghai, China) with a 13 mm diameter probe tip	Amount of juice: 200 mL; Frequency: 20 kHz; Duration of sonication: 1–12 min; Processing temperature: no information.	- Enzyme extraction; - Measurement of enzyme activity; - Measurement of total phenols; - Colour assessment.	A significantly lower inactivation rate of polyphenol oxidase (PPO) and polyphenol peroxidase (POD) was found with ultrasound with cooling (USC) than with ultrasound treatment, indicating that the heat released during ultrasound treatment was beneficial for enzyme denaturation. Furthermore, blueberry POD showed greater resistance to heat and sonication than PPO. In addition, ice cooling during ultrasonic treatment was beneficial for colour and retention of phenolic compounds in blueberry juice.	Cao X. et al. [43] (2018)
Mulberry juice	Ultrasonic bath (Daihan WUC-A10H, Seoul, Korea) containing 3 L of water as the coupling fluid	Amount of juice: 250 mL; Frequency: 40 kHz; Duration and processing temperature: -Group one: the mulberry mash was introduced into the ultrasound at different times of 30, 60, 90 and 120 min. The ultrasonic temperature was fixed at 60 °C; -Group two: the mash was treated with the ultrasound at 30, 45, 60 and 75 °C. The treatment time was fixed at 60 min.	- Total soluble solids; - Titratable acidity; - pH; - Moisture content; - Total phenolic content; - Antioxidant capacity; - Total anthocyanin contents; - L-ascorbic acid content.	Ultrasonic treatment showed significantly increased extraction efficiency for processing the antioxidant-rich mulberry fruit juice. Furthermore, it showed some advantages, such as shorter extraction time and higher extraction efficiency for total ascorbic acid phenolic and anthocyanin content compared to normal processing. Positive correlations between total phenolic content (TPC) and antioxidant capacity of mulberry juice suggest that polyphenols are the main antioxidants in this product.	Nguyen C. L. and Nguyen V. H. [44] (2018)
Cloudy apple juice	Ultrasound probe system (UP 100H, Hielscher Company, Teltow, Germany) with tip diameters of 20 and 10 mm	Amount of juice: 100 mL; Frequency: 30 kHz; Duration of sonication: 2, 6 and 9 min; Processing temperature: less than 41 °C.	- Determination of total phenols (TPs); - Determination of total flavan-3-ols (TFL); - Determination of in vitro antioxidant capacity.	This study showed the negative effect of ultrasound on the processing of cloudy apple juices with regard to the stability of bioactive compounds and antioxidant capacity. TP values and antioxidant capacity in the samples decreased during ultrasonic treatment. Furthermore, TFL showed higher stability than TP with a significant effect of probe diameter and treatment time. In addition, during 7 days of storage at 4 °C, the stability of TP, TFL and antioxidants decreased.	Kovačević D. B. et al. [45] (2019)
Plantlets juice	Ultrasonic (SKYMEN JP-031S, Skymen Cleaning Equipment Shenzhen Co. Ltd., Shenzhen, China) bath cleaner	Amount of juice: 100 mL; Frequency: 40 kHz; Duration of sonication: 20 and 40 min; Processing temperature: (30 °C, 45 °C and 60 °C).	- Total flavonoid contents (TFCs); - Total phenolic contents (TPCs); - The 2;2-Diphenyl-1-Picrylhydrazyl (DPPH) activity; - Total antioxidant capacity (TAC); - Carotenoids; - Chlorophyll contents; - Free amino acids; - Mineral elements; - Electric conductivity (EC); - Brix; - pH; - Cloud value; - Non-enzymatic browning (NEB); - Viscosity - Titratable acidity (TA); - Colour properties.	The results indicate that the use of ultrasound treatments is suitable for the production of better-quality wheat seedling juice with reduced microbial load and increased retention of other bioactive compounds, antioxidants and nutrient profile at a much lower temperature. Therefore, sonication can be successfully used as an alternative processing approach to ensure the microbiological and nutritional quality of wheat juice on an industrial scale.	Ahmed Z. et al. [46] (2019)
Strawberry–apple–lemon juice blend	Ultrasound machine (SCIENTZ-IID, Ningbo Xinzhi Biological Polytron Technologies Inc., Ningbo, China) with a probe diameter of 6 mm and an operating immersion depth of 2 cm	Amount of juice: 50 mL; Power: 376 W; Duration of sonication: 10 min; Processing temperature: 35 °C.	- pH and TSS analysis; - Turbidity analysis; - Instrumental colour assessment; - Ascorbic acid analysis; - Total phenol analysis; - Total anthocyanin analysis; - Determination of antioxidants; - Capacity.	This study showed that total phenols and ascorbic acid (AA) were more effectively retained in the high hydrostatic pressure (HHP)- and ultrasound (US)-treated juice blends than in the heat-treated (HT) blends. Furthermore, they showed a higher antioxidant capacity than the HT-treated samples. The US-treated juice blend was found to have a lower total anthocyanin content compared to the HHP-treated samples.	Feng X. et al. [47] (2020)
Sweet potato juice	Ultrasonic processor (UP200Ht; Hielscher Ultrasound Technology, Germany) attached to 40 mm diameter probe (treatment time of 8 min, ultrasonic intensity of 0.66 W cm2, and a constant frequency of 26 kHz)	Amount of juice: no information; Frequency: 26 kHz; Duration of sonication: 8 min; Processing temperature: no information.	- pH; - Soluble solids; - Titratable acidity; - Extraction and analysis of β-carotene; - In vitro digestion of β-carotene; - Analysis of total polyphenols; - Analysis of antioxidant activity (FTC and ORAC); - Analysis of enzymatic residual activity (polyphenol oxidase and - peroxidase); - Analysis of colour.	This study found that the bioavailability of β-carotene increased after treatment, with sonication being the most effective. All the treatments used effectively preserved the antioxidant properties of the juices. However, not all conditions used for processing were effective in inactivating polyphenol oxidase and peroxidase. After processing, the sweet potato juice samples retained their characteristic colour, and the results indicate that ultrasound was the best processing method.	Rios-Romero E. et al. [48] (2021)
Melon (Cucumis melo) juice	Branson Sonifier SFX- 550 (Branson Ultrasonics Corp, Danbury, CT, USA) equipped with a 1/2-inch tip-horn, operating at 550 W and 20 kHz	Amount of juice: 200 g; Frequency: 20 kHz; Duration of sonication: 10 and 15 min; Processing temperature: 10 °C ± 2 °C.	- Centrifugal sedimentation; - Colour difference; - Analysis of total carotenoids - Total phenolic compounds; - Antioxidant activity; - Radical scavenging according to DPPH assay; - ABTS free radical scavenging assay; - FRAP assay.	The use of high-intensity ultrasound (HIUS) for melon juice processing improved the physical appearance, bioactive compound content and microbiological quality of the juice. HIUS improved the physical appearance of melon juice by reducing sedimentation pulp, minimizing colour change. In addition, the melon juice processed via HIUS was enriched with bioactive compounds due to the increased total carotenoid content and the presence of gallic and syringic acids, increasing the antioxidant capacity of the juice.	Rodríguez-Rico D. et al. [49] (2022)

presents a summary of various analyses on the impact of ultrasound on the physical and chemical properties of various types of juices from the last 5 years.

According to the research results presented above, high-powered ultrasound treatment may be a promising alternative to traditional pasteurization or other processing methods to improve the quality of juices from various fruits and vegetables. It is worth continuing the experiments to determine the optimal process conditions that will meet both national and international requirements not only in terms of extending the shelf life, but also the pro-health value of beverages.

4. The Influence of Ultrasounds on the Microbiological Stability of Fruit, Vegetable and Fruit–Vegetable Juices

Due to the current trends related to a healthy lifestyle and rational nutrition, consumers, when choosing beverages, apart from taste, also take into account their pro-health value. However, in order to preserve the quality of the juice, it is necessary to preserve it before the distribution and storage process. The most popular method is still pasteurization, which involves the use of temperatures below 100 °C. Unfortunately, many substances that have a beneficial effect on the functioning of the human body belong to the group of thermolabile compounds. This means that they break down as a result of too-high temperatures. The pasteurization process has been modernized in recent years, so the losses of these compounds are much smaller. The main focus was on shortening the process time, in some cases even to several tens of seconds (flow pasteurization).

Due to rapid technological development, there are an increasing number of alternative methods. A number of tests are carried out that allow the introduction of innovative treatments that enable the production of products with the desired characteristics and quality, including being microbiologically safe for consumers.

Of particular importance are liquid food decontamination techniques that are inexpensive, simple, reliable and environmentally friendly. Ultrasonics have found application in the fruit and vegetable juice industry due to their desired multifunctional effect: reduced water and energy consumption, minimal loss of flavour and nutrients and increased product homogeneity [50]. Improvements in ultrasound generation technology in recent years have generated interest in reducing microbes. The mechanisms of ultrasonic inactivation of microorganisms are seen in a number of complex physicochemical processes, which are based on fast-changing mechanical stresses, energy dissipation, cavitation with a whole range of derivative phenomena and, in special cases, the so-called cell resonance. Studies show the destructive effect of ultrasound on colonies of microorganisms, manifested in the form of damage to cell elements and tissue structures. Figure 3 shows the classification of the mechanisms responsible for the microbial inactivation effect of sonication.

The microbiological quality of the final product is determined by both the microbiological purity of the raw material and the hygiene of production. The risk of microbiological contamination is associated with the possibility of the development of numerous groups of microorganisms, even in refrigerated storage conditions. The microflora most often infecting fruit or vegetables are bacteria resistant to acidification of the environment, soil or air, lactic acid bacteria, acetic acid bacteria, acid-tolerant fungi and both moulds and yeasts.

Bacteria responsible for spoilage of juices include, among others, Bacillus thermoacidurans, which causes gasless fermentation and acid fermentation. In turn, Alicyclobacillus acidoterrestris causes changes in taste and smell, producing a substance called guaiacol, which is characterized by a medical, disinfectant, smoky smell. And in the presence of Clostridium pasteurianum, large amounts of butyric acid and acetic acid are formed and significant amounts of gases are released. In addition, yeasts are among the dominant microorganisms present in juices (mainly fruit juices) due to their ability to grow at low pH values. The fermentation yeast Saccharomyces cerevisiae produces significant amounts of carbon dioxide and ethanol, making it one of the main spoilage agents.

This type of microbial contamination of beverages can lead not only to the deterioration of their nutritional and sensory properties (functional ingredients, colour, taste, smell), but also contribute to food-borne diseases caused by pathogenic bacteria or toxic fungi. The causes of most diseases in the world, including fatal ones, are mainly Escherichia coli O157:H7, Salmonella and Bacillus cereus, which are found in raw apple and citrus juices. Listeria monocytogenes, although it is not a cause of food poisoning, was isolated from unpasteurized apple juice and beetroot juice [51,52,53,54].

The results reported by Guerrouj et al. [55] indicate that a high level of inactivation of microorganisms present in beverages can be achieved by combining sonication with moderate heat. Freshly squeezed orange juice was sonicated for 1, 10, 20 and 30 min at a frequency of 24 kHz to evaluate its effect on selected microorganisms. Ultrasound was generated continuously using a UP200H processor from Hielscher Ultrasound Technology and an S3 probe (Hielscher). Overheating of the samples during the ultrasound treatment was prevented by circulating ice water through the treatment chamber. The number of aerobic mesophilic bacteria was significantly reduced when the prepared product was sonicated for 30 min, while 20 min of sonication was sufficient to eliminate yeasts and moulds. The results of this study indicate that the applied process conducted at a temperature of 43–45 °C can be used to process orange juice and leads to an improvement in its microbiological safety.

Adekunte et al. [56] used a 1500 W ultrasonic processor (VC 1500, Sonics and Materials Inc., Newtown, USA) for the sonication of tomato juice. Samples whose processing temperature ranged from 32 to 45 °C were processed at a constant frequency of 20 kHz. The performed tests showed the inactivation of yeast (Pichia fermentans) by 5 log, and the level of their reduction depended on the applied amplitude and duration of the process, which were 61 μm and 7.5 min, respectively. The yeast inactivation observed may result from the aforementioned combination of physical and chemical mechanisms occurring during cavitation. The formation of free radicals and H₂O₂ during sonication helps to inactivate microbes. Nevertheless, the physical effects of the process may not be effective enough to inactivate yeast cells that are difficult to disrupt through micro-jet action. Rather, ultrasound may lead to rupture of yeast cells and subsequent release of protein [57].

Satisfactory research results were obtained by Bhat et al. [58] when freshly squeezed kasturi lime fruit juice (Citrus microcarpa) was sonicated for 30 and 60 min at 20 °C and 25 kHz (using an ELMA^® cleaning bath, model T700h W/Acc, Singen, Germany) for microbiological evaluation of the product via analysis of total plate counts, yeast and mould. Sonication of juice samples for 60 min showed a significant reduction in most microorganisms compared to 30 min treatment of samples and control (untreated) samples along with the achievement of final product safety and quality standards through 5-log reductions, which is the mandatory level set by the FDA for fruit and vegetable juices.

In the work of Khandpur and Gogate [59], the effectiveness of ultrasound in the processing of various fruit and vegetable juices obtained from oranges, sweet limes, carrots and spinach was assessed in terms of the growth of microorganisms and changes in quality parameters during storage. Ultrasonic treatment with a constant frequency (20 kHz) for 15 min at a temperature below 30 °C was adopted as the optimal process conditions. The results of the authors’ research unequivocally confirmed the usefulness of sonication to maintain the microbiological safety of beverages characterized by an extended shelf life and excellent quality parameters compared to untreated (control) and thermally treated juices.

The analyses of Choo et al. [60] indicate that regardless of the method of processing used for noni fruit juice (Morinda citrifolia L.), i.e., ultrasonic exposure for 60 min at a constant temperature of 30 °C and frequency of 37 kHz or pasteurization at 90 °C for 60 s, the level of microbes may be within the acceptable range. No yeasts or moulds were detected during 8 weeks of refrigerated storage (4 °C), while the total number of aerobic mesophilic bacteria remained below 10⁴ CFU/mL, indicating that the juice was edible. However, importantly, compared to the fresh sample (control, untreated), the amount of malic acid, ascorbic acid and other beneficial compounds was increased in the sonicated sample but decreased in the pasteurized sample. This study highlights the feasibility of replacing traditional pasteurization with ultrasonic processing to improve the quality of noni juice on an industrial scale.

Several other studies have shown a greater reduction in microbes in juice samples treated with ultrasound at higher temperatures. For example, carrot juice was thermosonicated (24 kHz) at 50 °C, 54 °C and 58 °C for 10 min to evaluate quality parameters and microbial growth after processing and during storage at 4 °C. Control and samples sonicated at 50 °C and 54 °C had 10-, 12- and 14-day shelf lives, respectively. However, juices subjected to ultrasound at 58 °C had the best microbiological quality. In this case, microbial growth remained low at approximately 3 log for mesophiles, 4.5 log for yeasts and moulds and 2 log for Enterobacteria after 20 days of storage. What is more, the juice prepared in this way retained >98% of its carotenoids and 100% of its ascorbic acid. The content of phenolic compounds increased in all processed and stored juices. In this way, the planned experiment confirms that thermosonication is a promising technology ensuring the stability of carrot juice by retarding the growth of microorganisms during storage with minimal changes in the number of bioactive compounds [61].

Bermúdez-Aguirre and Barbosa-Cánovas [62] subjected pineapple, grape and cranberry juices to sonication (24 kHz) at 40 °C, 50 °C and 60 °C for 10 min in continuous and pulsed modes. They observed a reduction in Saccharomyces cerevisiae by 5–7 log units (depending on the product to be fixed) during treatments at 60 °C, with the continuous mode being more effective. In general, the authors of this paper stated that this processing technique seems to be a viable technology for preserving juices, but the process parameters should be controlled, because changes in the pH and colour of products may occur during cavitation. Ultrasound has been reported to increase the sensitivity of microorganisms to heat, high osmotic pressure and low pH due to cavitation and other changes in the outer membrane of the cellular structure.

Basumatary et al. [63] examined the effect of thermal sonication on the microbiological quality of pomelo (Citrus maxima) juice, which they treated for 15, 30, 45 and 60 min at 30, 40 and 50 ± 2 °C. Two ultrasound frequencies, 33 and 44 kHz, were used. Very good test results were obtained, because the total number of microorganisms, yeasts and moulds decreased significantly in relation to the control samples (fresh juice). With the increase in treatment time and temperature, the number of microorganisms under consideration decreased. Subsequent studies therefore indicated that a properly conducted process with the use of ultrasound can be used as a substitute for heat treatment in the processing of pomelo juice due to its positive effect on the quality of the final product (significant increase) in terms of antioxidant activity, with a lower decrease in ascorbic acid content compared to pasteurization.

Subsequent analyses confirmed that ultrasonic treatment may not only be an alternative to pasteurization, but also improve the nutritional value and safety of freshly pressed plum juice. Research by Oladunjoye et al. [64] consisted of pasteurization of the product at 90 °C for 60 s and thermal sonication of the drink for 5, 10, 20 and 30 min at the temperatures of 40, 50 and 60 °C at an ultrasound frequency of 40 kHz, and then determination of the microbiological properties. When evaluating the microflora of pasteurized plum juice, a decrease in all assessed microorganisms to an undetectable level was found, which could be related to cell wall/membrane rupture and nuclear components leading to cell death. However, after thermal sonication at 40 °C, the total number of microorganisms decreased from 3.75 to 1.01 log CFU/mL, and the number of yeasts and moulds decreased from 4.17 to 3.64 log CFU/mL. Raising the ultrasonic treatment temperature to 50 and 60 °C allowed the number of bacteria to be eliminated to undetectable levels, while the number of yeasts and moulds at 50 °C decreased from 3.36 to 1.93 log CFU/mL, with no detectable increase at 60 °C. The incomplete inactivation of yeasts and moulds at 40 °C and partially at 50 °C may be due to the formation of spores that are resistant to these treatment conditions. The inactivation of microorganisms via thermal sonication mainly concerns the acoustic cavitation taking place in the cellular structure, which in turn causes the implosion of bubbles in the liquid and/or the generation of free radicals with antimicrobial properties.

In addition, the high acidity level of plum juice and induced osmotic pressure, together with other reactions, may intensify the effect of cavitation on the structure of microorganisms, resulting in the release of intracellular components, such as protein lipids and nuclear compounds [57,65,66,67,68]. Not without significance is the high quality of ultrasound-treated juice, which is characterized, among other properties, by a higher content of ascorbic acid, carotenoids (2.19–4.30%) and flavonoids (10–16%), as well as antioxidant activity (32, 52–48.5%) compared to the control (untreated juice). Only elevated temperature (60 °C) was not conducive to most of these quality features, which means that the thermosonication process may require further optimization of treatment parameters in order to maintain the quality of the final product to the maximum extent [64].

Due to the biological diversity of individual food products, the structure of ultrasonic generators and the methods of setting and expressing process parameters, it is difficult to clearly indicate the optimal processing conditions for juices pressed from various fruits or vegetables. Based on our own research, we found that the ultrasound intensity needed to inactivate microorganisms ranges widely from 28 to 90 W/cm2 and depends on the initial microbiological condition and the type of juice processed. In the case of tomato juice, we showed that ultrasound sonication at an intensity of 40 W cm−2 for 5 min and 28 W cm−2 for 10 min yielded a microbiologically pure product devoid of microorganisms involved in spoilage even after 10 days of storage [68].

Other research results relating to the effective minimization of the presence of microorganisms contaminating fruit and vegetable juices through the action of ultrasounds were collected, as presented in

Table 2. Other research results relating to the effective minimization of the presence of microorganisms contaminating fruit and vegetable juices through the action of ultrasounds.

Type of Juice	Type of Ultrasound Generator	Process Parameters	Microorganisms Discussed	Sonication Effect	References
Green cactus pear (Opuntia ficus Indica) juice	Ultrasonic processor (VCX1500, Sonics & Materials, Inc. Newtown, CT, USA)	Amount of juice: 250 mL; Frequency: 20 kHz; Duration of sonication: 10, 15 and 25 min (at amplitudes of 40%, 60% and 80%); Processing temperature: 36.1 ± 2.55 °C–76.3 ± 1.16 °C.	Total plate counts; Enterobacteria	In the control juice sample, 4.6 and 4.2 log CFU/mL total plate counts and Enterobacteria were detected, respectively. The organisms considered were below the detection limit when samples were treated at 60% and 80% amplitudes and 15 and 25 min. The temperature reached during the processing of these samples (58 and 76 °C) may explain the reduction in microorganisms.	Cansino et al. [69] (2013)
Carrot juice	Ultrasonic device (Hielscher USA Inc., Ringwood, N.J., U.S.A.) with a probe of 22 mm	Amount of juice: 500 mL; Frequency: 24 kHz; Duration of sonication: 2–10 min; Processing temperature: 50, 54 and 58 °C.	Escherichia coli (ATCC 11755)	A more than 5-log reduction in E. coli ATCC 11755 was achieved in 2 min via the application of ultrasound (24 kHz, 120 μm, 400 W) at 58 °C. Treatment at 54 °C for 10 min was required to achieve the same level of reduction. However, at 50 °C, only a 3.5-log reduction was achieved after 10 min of treatment.	Pokhrel et al. [70] (2017)
Strawberry juice	Ultrasonic cleaning bath (TestLab, Argentine)	Amount of juice: no information Frequency: 40 kHz; Duration of sonication: 10 and 30 min; Processing temperature: 20 ± 1 °C.	Total mesophilic bacteria; total psychrophilic bacteria; yeast and moulds	The storage growth rate of each microbial population was significantly reduced after 30 min of treatment. During 10 days of storage, the reduction in total mesophilic bacteria, total psychrophilic bacteria and yeast and moulds was 0.78, 0.64 and 2.5 log CFU/mL relative to the control.	Tomadoni et al. [71] (2017)
Star fruit (Averrhoa carambola) juice	Ultrasonicator bath (GT Sonic, Guangdong, China)	Amount of juice: 50 mL Frequency: 44 kHz; Duration of sonication: 15, 30, 45, and 60 min; Processing temperature: 25–45 °C.	Total plate counts;yeasts and mould	Regardless of the time and temperature of the process, the complete inactivation of bacterial populations was achieved, and the amount of yeast and mould was reduced from 6.10 (control sample) to about 5 log CFU/mL.	Nayak et al. [72] (2018)
Pumpkin juice	Ultrasonic bath (Elmasonic ultrasonic E-100H, Germany)	Amount of juice: 10 mL Frequency: 37 kHz; Duration and time of ultrasonication: 30 min, 23 °C; duration and time of thermosonication: 30 min, 40, 50 and 60 °C.	Escherichia coli K-12	Ultrasonic treatment was only able to inactivate E. coli K-12 by less than 1 log; however, thermosonication (60 °C, 30 min) reduced the microorganisms by 6.62 ± 0.00 log CFU/mL.	Demir and Kılınç [73] (2019)
Red grape juice	Ultrasonic processor for small- and medium-volume applications (VCX 750, Sonics & Materials, Inc., Newtown, CT, USA) with a 1/2″ (13 mm) probe	Amount of juice: 150 mL; Frequency: 20 kHz; Duration of sonication: 5 and 10 min; Processing temperature: no information.	Total plate counts; Enterobacteriaceae	The lowest total plate counts for Enterobacteriaceae, 1.13 log CFU/mL and 0.53 log CFU/mL, were recorded after sonication for 10 min at 70% amplitude.	Margean et al. [36] (2020)
Red watermelon juice	UP200St- ultrasound device produced by Hielscher Ultrasonics (Berlin, Germany)	Amount of juice: 100 mL; Frequency: 24 kHz; Duration of sonication: 4, 8, 12 and 16 min; Processing temperature: no information.	Total aerobic plate count; yeast and mould	A reduction in total aerobic plate count and yeast and mould was observed from 2.72 to 3.07 log CFU/mL to undetectable levels after 8 min of sonication.	Yıkmış [37] (2020)
Pineapple juice	Ultrasonic processor (UP200S, 200 W, Hielscher, Teltow, Germany) with a 13 mm probe diameter	Amount of juice: 80 mL Frequency: 24 kHz; Duration of sonication: 2–10 min; Processing temperature: 25–65 °C.	Total plate counts; yeast and mould	The average level of contamination of fresh juice samples (day 0) with total plate counts, yeasts and moulds was 4.71 and 4.51 log CFU/mL, and for thermosonicated juice samples, 2.74 and 2.30 log CFU/mL. The applied treatment turned out to be effective in delaying the development of microorganisms during 28 days of storage.	Mala et al. [74] (2021)
Spinach juice	Ultrasonic homogenizer (Scientz-IID, Ningbo, China).	Amount of juice: 100 mL Frequency: 30 kHz; Duration of sonication: 20 min; Processing temperature: 60 °C.	Total plate count; yeasts and mould; Escherichia coli	There was a reduction in total aerobic plate count, yeast and mould as well as E. coli from 4.10, 3.63 and 1.75 log CFU/mL to undetectable levels in the sample treated at 30 kHz, power 600 W, 50% duty cycle, 60 ± 1 °C for 20 min.	Manzoor et al. [50] (2021)
Black carrot juice	Sonicator (UP200S, Hielscher, Germany) with a probe of 7 mm	Amount of juice: 50 mL; Frequency: 24 kHz; Duration of sonication: 4, 8 and 12 min; Processing temperature: 25 °C.	Total plate counts; yeasts and mould	The application of ultrasound for 12 min reduced the total plate count by approximately 2.63, 3.39 and 2.9 log CFU/mL, and for yeast and moulds, nearly 1.29, 2.42 and 2.0 log CFU/mL after 1, 7 and 15 days of storage.	Hasheminya and Dehghannya [75] (2022)

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Koda et al. [76] studied the effectiveness of ultrasound with a frequency of 500 kHz on the inactivation of microorganisms (Gram-negative bacteria Escherichia coli IAM 12058 and Gram-positive bacteria Streptococcus mutans JCM 5175) and concluded that the mechanism of their action is based mainly on chemical effects. Transmission electron microscopy (TEM) studies of bacteria confirmed lethal damage resulting from the interaction of bacterial cells with cavitation bubbles. A significant number of empty cell envelopes as well as their cytoplasmic content have been observed.

According to other scientists, the mode of action of ultrasound on microorganisms in liquids includes hydrodynamic effects (the phenomena of intracellular cavitation and microflux) and the formation of radicals that disrupt the structure of the cell. However, sometimes the use of ultrasound alone is not effective enough to decontaminate juices, especially those contaminated with pathogenic bacterial flora. In addition, high ultrasound power levels may adversely affect the nutritional and sensory properties of food [77,78,79].

Therefore, Kernou et al. [80] evaluated the survival of Escherichia coli ATCC 25922 in orange juice treated with microwaves and/or ultrasounds. Sonication conducted for 60 min at a frequency of 42 kHz had no significant effect on the reduction in survival rates (1.3-log reduction) of the bacteria under consideration. However, it has been established that this method of pretreatment can increase the effectiveness of microwaves to improve the microbiological quality of the juice. More specifically, a significant reduction (by 8.0 log) was achieved via sonication (42 kHz; 20 min) followed by microwave processing (900 W; 30 s) of the orange beverage.

Similar relationships were found by Hosseinzadeh Samani et al. [81] after subjecting cherry juice to microwave–ultrasound treatment. Based on response surface modelling (RSM), the optimal processing conditions were microwave output power, 352.21 W; temperature, 49.94 °C; ultrasound power, 475.13 W; and exposure time, 6 min. Such parameters resulted in a reduction in the number of E. coli bacteria, while maintaining the amount of vitamin C at the level of 142.5 mg per 100 mL. Unfortunately, it was found that each increase in the power of ultrasounds or microwaves and the extension of the exposure time led to a significant reduction in the content of vitamin C due to its increased oxidation, mainly due to the thermal effect. Microwave processing is fast and can significantly reduce the heating time of the juices. And the hydrodynamic effect of ultrasound on microorganisms has great potential to improve the effectiveness of their elimination. This study contributes to the design and control of effective methods for the inactivation of pathogenic microorganisms and food spoilage agents that can be achieved using combined methods.

Satisfactory research results were also obtained through combining ultrasonic and pulsed light treatment in order to obtain good microbiological stability of juice prepared from apples [78]; the use of ultraviolet radiation and ultrasonic treatments to inactivate Alicyclobacillus acidoterrestris spores, also in apple juice [82]; the use of pulsed electric field and ultrasounds on the microbiological quality of grapefruit juice [83]; ultrasound and ozone processing to reduce the number of native microflora in cashew apple juice [84]; high-pressure processing and ultrasound for the preservation of strawberry juice [85]; and ultrasonic treatment with high-voltage cold plasma to ensure better quality and greater stability during carrot juice processing [86].

Ultrasound alone or in combination with mild temperatures (<50 °C in most cases) is a promising alternative technology for juice preservation. Power ultrasound has been shown to be effective against microorganisms contaminating liquid foods and meets the U.S. Food and Drug Administration’s 5-log reduction requirement (pasteurization standards) for certain pathogens contaminating fruit and vegetable juices. This technology has the ability to destroy cell membranes due to the high pressure, temperature and shear forces generated during cavitation. In addition, the inhibitory effect of thermal sonication on the population of microorganisms may result from cavitation and the production of free radicals (H+, O₂, OH, HOO) formed as a result of the sonolysis of water molecules present in juices. Free radicals are therefore co-responsible for causing oxidative damage, disrupting microbial cell walls and inhibiting microbial growth by inactivating the enzymatic activity of mitochondria [87,88,89].

Where ultrasound alone is not sufficiently effective in achieving high microbial reduction, combining it with treatment methods (temperature, pressure, microwave, pulsed light, ultraviolet, HPP, etc.) is recommended. This combination often shows a synergistic effect on more effective inactivation of microbes than when using individual methods separately.

5. Prospects for the Use of Ultrasound in the Future

The application of power ultrasound in juice preservation holds promise for enhancing the quality, safety and shelf life of fruit juices. However, it is important to note that while power ultrasound offers numerous benefits for juice preservation, its effectiveness depends on factors such as equipment design, operating parameters and the specific characteristics of the fruit juice being processed. Progress in the construction of ultrasonic generators has made them more and more efficient, which allows for obtaining higher ultrasonic power with lower energy consumption. Ultrasonic output power is a key factor in scaling up the process. Processing larger volumes of liquid will require more powerful equipment. The ultrasonic power of the devices currently used on an industrial scale varies between 500 W and 16 kW. In typical liquid processing applications, four or more units are clustered together for increased redundancy and capacity customization. For example, a cluster with a power of 60 kW can be used to process up to 50 cubic meters of liquid per hour. It is expected that in the future, the power of ultrasonic characteristics will be further increased, which will significantly increase the throughput of processes in which ultrasound is used. Another factor determining the economy of ultrasonic processes is the conversion of electrical energy into mechanical power. Currently used devices are characterized by an efficiency exceeding 85%. Further progress in the construction of ultrasonic transducers will increase the efficiency of energy conversion and reduce the costs of electricity needed to power ultrasonic generators [90].

New ultrasound generators offer better control of ultrasound parameters such as frequency, amplitude and pulse duration. This will make it possible to adjust the parameters of ultrasonic treatment to specific groups of microorganisms, as well as specific physicochemical properties of the juice. A promising perspective in the preservation of fruit and vegetable juices may also be the construction of devices that include ultrasonic transducers operating at different ultrasound frequencies. This can contribute to a significant increase in the efficiency of the disintegration of microorganisms. Another important trend in preserving juices may be the use of pulsating processing. On the one hand, it significantly reduces the specific energy consumption, and on the other hand, it can be more effective than continuous processing. Another trend will be the use of installations for ultrasonic preserving juices in the flow. Placing ultrasonic transducers inside flowing juice allows for effective and regular transfer of ultrasonic energy. This enables the use of ultrasound on a larger scale, which requires a constant flow of material.

Undoubtedly, an interesting perspective is the combination of ultrasound with other techniques, such as microwave treatment or white pulsating light, which will allow for much better effects in the field of juice preservation and will also affect the preservation, allowing for a better overall quality of the final product. Further development of the application of ultrasound for preserving juices will require adaptation to specific types of fruit juice. Before implementing this technology on an industrial scale, it is necessary to conduct appropriate research, tests and analyses in order to determine the optimal conditions and ultrasound parameters for a given application.

6. Conclusions

Ultrasound has great potential in preserving fruit and vegetable juices. This method makes it possible to reduce the activity of microorganisms responsible for product spoilage, minimizing the loss of biohealth, nutritional, taste and aroma values. Juices pasteurized using ultrasound have a higher content of ascorbic acid compared to juices preserved using traditional heat treatments. In some cases, they also show higher antioxidant activity. The impact of ultrasonic treatment on the effectiveness of juice preservation depends on the intensity and frequency of the ultrasounds used as well as the time and temperature of the process. Important parameters include the chemical composition of the juice and the structure, shape and condition of microorganisms and enzymes. Some enzymes, such as glucose oxidase, peroxidase and polyphenoloxidase, can be effectively inactivated by ultrasound. Still, many microorganisms and enzymes are resistant to ultrasound, which makes it necessary to combine it with technologies such as pulsating light or microwave treatment. The combination of ultrasound with other novel or conventional methods not only overcomes this limitation but also increases the efficiency of the ultrasound-based technique.

Ultrasonic juice preservation is an interesting prospect, but requires careful analysis and case-by-case study to ensure product effectiveness and quality. Future research should focus on scaling up and standardizing ultrasonic machining processes.