Applications and opportunities for ultrasound assisted extraction in the food industry A review

Kamaljit Vilkhu a,, Raymond Mawson a , Lloyd Simons a , Darren Bates b

a

Ultrasonics Processing Group, Food Science Australia, 671 Sneydes Road, Werribee, VIC 3030, Australia b Innovative Ultrasonics Pty Ltd, P.O. Box 321, Noosaville, QLD 4566, Australia

Abstract Ultrasound assisted extraction (UAE) process enhancement for food and allied industries are reported in this review. This includes herbal, oil, protein and bioactives from plant and animal materials (e.g. polyphenolics, anthocyanins, aromatic compounds, polysaccharides and functional compounds) with increased yield of extracted components, increased rate of extraction, achieving reduction in extraction time and higher processing throughput. Ultrasound can enhance existing extraction processes and enable new commercial extraction opportunities and processes. New UAE processing approaches have been proposed, including, (a) the potential for modification of plant cell material to provide improved bioavailability of micro-nutrients while retaining the natural-like quality, (b) simultaneous extraction and encapsulation, (c) quenching of the radical sonochemistry especially in aqueous systems to avoid degradation of bioactives and (d) potential use of the radical sonochemistry to achieve targeted hydroxylation of polyphenolics and carotenoids to increase bioactivity.

Keywords: Ultrasound assisted extraction; Cavitation; Particle size; Mass transfer Industrial relevance: The application of ultrasonic assisted extraction (UAE) in food processing technology is of interest for enhancing extraction of components from plant and animal materials. This review shows that UAE technology can potentially enhance extraction of components such as polyphenolics, anthocyanins, aromatic compounds, polysaccharides, oils and functional compounds when used as a pre-treatment step in a unit process. The higher yield obtained in these UAE processes are of major interest from an industrial point of view, since the technology is an add on step to the existing process with minimum alteration, application in aqueous extraction where organic solvents can be replaced with generally recognised as safe (GRAS) solvents, reduction in solvent usage, and shortening the extraction time. The use of ultrasonic for extraction purposes in high-cost raw materials is an economical alternative to traditional extraction processes, which is an industry demand for a sustainable development.

1. Introduction The application of ultrasound as a laboratory based technique for assisting extraction from plant material is widely published. Several reviews have been published in the past to extract plant origin metabolites (Knorr, 2003), flavonoids from foods using a range of solvents (Zhang, Xu, & Shi, 2003) and bioactives from herbs Vinatoru (2001). A limited number of publications have included continuous ultrasonic process development and pilot-scale applications. The range of published

Corresponding author. Tel.: +61 3 9731 3449; fax: +61 3 9731 3250. E-mail address: kamaljit.vilkhu@csiro.au (K. Vilkhu).

extraction applications include herbal, oil, protein and bioactives from plant materials (e.g. flavones, polyphenolics), summarised in Table 1 and outlined in more detail in the following Applications section. Much of the work is empirical in nature and explanations of the mechanisms have been proposed. Some workers also discuss both the mechanisms involved in UAE and the likely issues for potential for scale up. The review by Vinatoru (2001) outlines a program of work where industrial scale up was attempted under an EU Copernicus grant (ERB-CIPA-CT94-02271995). They highlight that while it is relatively easy to achieve extraction on the laboratory bench it is very challenging to attempt extraction on an industrial scale. Several key issues and observations relating to UAE have been identified, as follows, (1) the nature of the tissue being extracted and the location of the

Table 1 List of ultrasound assisted extraction studies from the literature on various food components Product Almond oils Herbal extracts (fennel, hops, marigold, mint) Ginseng saponins Ginger Soy protein Soy isoflavones Rutin from Chinese Scholar Trees Carnosic acid from rosemary Polyphenols, amino acid and caffeine from green tea Pyrethrines from flowers Ultrasound Process Batch, 20 kHz Stirred batch, 20 to 2400 kHz Batch, 38.5 kHz Batch, 20 kHz Continuous, 20 kHz, 3 W per gram Batch, 24 kHz Batch, 20 kHz Batch, 20 and 40 kHz Batch, 40 kHz Batch, 20 and 40 kHz Solvent Supercritical carbon dioxide Water and ethanol Water, methanol and n-butanol Supercritical carbon dioxide Water and alkali (sodium hydroxide) Water and solvent Water and methanol Butanone and ethyl acetate Water Hexane Performance 30% increased yield or extraction time reduction Up to 34% increased yield over stirred 3-fold increase of extraction rate 30% increased yield or extraction time reduction 53% and 23% yield increase over equivalent ultrasonic batch conditions Up to 15% increase in extraction efficiency Up to 20% increase in 30 min Reduction in extraction time Increased yield at 65 C, compared with 85 C Increased yield at 40 C, compared with 66 C Author Riera et al. (2004) Vinatoru (2001) Wu et al. (2001) Balachandran et al. (2006) Moulton and Wang (1982) Rostagno et al. (2003) Paniwynk et al. (2001) Albu et al. (2004) Xia et al. (2006) Romdhane and Gourdan (2002)

components to be extracted with respect to tissue structures, (2) pretreatment of the tissue prior to extraction, (3) the nature of the component being extracted, (4) the effects of ultrasonics primarily involve superficial tissue disruption, (5) increasing surface mass transfer (Balachandran, Kentish, Mawson, & Ashokkumar, 2006; Jian-Bing, Xiang-hong, Mei-qiang, & Zhi-chao, 2006), (6) intraparticle diffusion, (7) loading of the extraction chamber with substrate, (8) increased yield of extracted components and (9) increased rate of extraction, particularly early in the extraction cycle enabling major reduction in extraction time and higher processing throughput (Moulton & Wang, 1982; Caili, Haijun, Quanhong, Tongyi, & Wenjuan, 2006). Living tissues where the desired components are localized in surface glands can be stimulated to release the components by relatively mild ultrasonic stressing (Toma, Vinatoru, Paniwnyk, & Mason, 2001). In tissues where the desired components are located within cells, pre-ultrasound treatment by size reduction to maximise surface area is critical for achieving rapid and complete extraction (Riera, Gols, Blanco, Gallego, Blasco, & Mulet, 2004; Balachandran et al., 2006;Vinatoru, 2001). Where pre-hydration is necessary to achieve extraction, ultrasound effectively accelerates the hydration process (Vinatoru, 2001). Ultrasound induced cavitation bubbles present hydrophobic surfaces within the extraction liquid (Grieser, personnel communication) thereby increasing the net hydrophobic character of the extraction medium. Thus it is possible to extract polar components into otherwise hydrophilic aqueous extraction media, reducing the need for generally undesirable hydrophobic or strongly polar extraction media. The disruption of tissue surface structures is revealed with microscopic examination by Vinatoru (2001), Chemat, Lagha, AitAmar, Bartels, and Chemat (2004), Haizhou, Pordesimo, and Weiss (2004), Balachandran et al. (2006). Several of the authors in the work cited below highlight concerns due to the potential for ultrasonic cavitation to propagate free radicals, in particular hydroxyl radicals. Where the potential oxidative damage is a concern radical production can be quenched by the addition of small amounts of ethanol to lower the

temperatures within the cavitation bubbles and extinguish the chemistry involved (Sun et al. unpublished work in progress). This paper provides a compilation of food-related UAE applications, highlighting the application approaches and performance. Following this, a more detailed discussion is given on UAE mechanisms, process development, equipment design and future opportunities. 2. Applications 2.1. Herbal and oil extraction Ultrasound has been recognised for potential industrial application in the phyto-pharmaceutical extraction industry for a wide range of herbal extracts. Vinatoru (2001) published an overview of the UAE of bioactive principles from herbs. The improvement in extractive value by UAE compared with classic methods in water and ethanol for fennel, hops, marigold and mint was 34%, 18%, 2%, and 3% respectively in water, whereas 34%, 12%, 3%, and 7%, respectively in ethanol. In another study, an aqueous extraction of Geniposide from Gardenia fruit was investigated by Jian-Bing et al. (2006). When ultrasound was applied at 0.15 W cm 2 the extraction yield of Geniposide was increased by 16.5%, in comparison with a static process using 40 ml/g of the solvent volume to fruit weight. The variability in percentage extract yield was mainly due to the individual product structure. Large scale ultrasonic extraction designs were proposed for stirred tank systems with temperature control. In recent years, Albu, Joyce, Paniwnyk, Lorimer, and Mason (2004) investigated the effect of different solvents and ultrasound on the extraction of carnosic acid from rosemary. Using conventional stirred extraction ethanol was significantly less effective then ethyl acetate and butanone. The application of ultrasound improved the relative performance of ethanol such that it was comparable to butanone and ethyl acetate alone.

Thereby ultra-sonication may reduce the dependence on a solvent and enable use of alternative solvents which may provide more attractive (a) economics, (b) environmental and (c) health and safety benefits. Ginsenosides (tri-terpene saponins) are known as the principal ingredients of ginseng roots. Ginseng saponins are associated with traditional herbal medicine and health foods (Tang & Eisenbrand, 1992). UAE of ginseng saponins was approximately 3-times faster than the traditional extraction method involving reflux of boiling solvents in a soxhlet extractor. Furthermore, the UAE technique was achieved at lower temperatures which are more favourable for thermally unstable compounds (Wu, Lin, & Chau, 2001). Similar results were reported on UAE of carvone and Limonene from caraway seeds, which resulted in 2 fold increases in their contents (Chemat et al., 2004). Likewise, anthraquinones from roots of Morinda citrifolia (Noni) are the active compounds which show several therapeutic effects and used in anti-cancer medical applications. Recently, Hemwimol, Pavasant, and Shotipruk (2006) investigated the use of UAE to improve the solvent extraction efficiency of anthraquinones from the roots of M. citrifolia. Ultrasound extraction in an ethanol water system provided a 75% reduction in extraction time and yield comparable with non-sonicated sample. Supercritical fluid extraction (SFE) is an intrinsically capital intensive process where any enhancement of extraction efficiency either in terms of extraction rate or yield is economically attractive. Over a period of many years it has been shown that combined action of ultrasound and supercritical carbon dioxide on extraction could be used to significantly improve extraction rate or yield of amaranth oil from seeds (Bruni, Guerrini, Scalia, Romagnoli, & Sacchetti, 2002), almond oil (Riera et al., 2004), tea seed oil (Rajaei, Barzegar, & Yamini, 2005), gingerols from ginger (Balachandran et al., 2006), operating parameters such as temperature, pressure and CO2 flow for Adlay seed (Coix lachrymal-jobi L. var. Adlay) oil and coixenolide from adlay seed (Ai-jun, Shuna, Hanhua, Tai-qiu, & Guohua, 2006). UAE has been recognised for application in the edible oil industry to improve efficiency and reduce extraction time (Babaei, Jabbari, & Yamini, 2006). This potential was based on UAE increases in oil from soybeans; carvone and limonene from caraway seeds. The ultrasonically induced cavitation was shown to increase the permeability of the plant tissues. Microfractures and disruption of cell walls in soybean flakes (Haizhou et al., 2004) and caraway seeds cell wall (Chemat et al., 2004) provided more evidence for the mechanical effects of ultrasound thus facilitating the release of their contents, in contrast to conventional maceration or extraction. These effects were identified under scanning electron microscopy. Importance was given to the effect of solvent vapour pressure and surface tension on cavitation intensity. The benefit of using ultrasonic pre-treatment before extracting oil from the seeds of Jatropha curcas L., and almond and apricot seeds by aqueous enzymatic oil extraction (AEOE) process was evaluated by Shah, Sharma, and Gupta (2005), Sharma and Gupta (2006). Ultrasonic pre-treatment of the almond and apricot seeds before aqueous oil extraction and aqueous enzymatic oil extraction provided significantly higher

yield with reduction in extraction time. Thus, implementation of ultrasonic pre-treatment reduced oil extraction time that may improve through put in commercial oil production process. 2.2. Protein extraction A small pilot-scale ultrasound batch and continuous soy protein extraction trials were reported by Moulton and Wang (1982). The continuous high-intensity application extracted 54% and 23% more protein for aqueous and alkali extraction respectively, compared with the batch extraction using comparable processing times and volumes. During the trials it was estimated that the continuous process used 70% less energy than the batch system to extract the same amount of protein and sonication efficiency improved with the greater load of thicker slurry, up to 1:10 (flake to solvent) ratio. 2.3. Bioactive extraction from plant materials 2.3.1. Polyphenols Grape marc is the solid waste of the wine-making process. Consisting of skins, seeds, and small amount of leaves, grape marc has long been used for alcohol, tartaric acid and more recently, the recovery of phenolic compound. Phenolic compounds are of particular interest in wine industry as it gives the characteristics colour and flavour in wine, and in pharmaceutical industry for its benefits on human health (Brenna, Buratti, Cosio, & Mannino, 1998). Polyphenols are associated with reduced risk of cardiovascular disease by inhibiting in-vitro oxidation of lowdensity lipoproteins possess anti-ulcer, anti-mutagenic, antiinflammatory activity and anti-carcinogenic properties (Flamini, 2003; Negro, Tommasi, & Miceli, 2003; Bonilla, Mayen, Merida, & Medina, 1998; Palma & Taylor, 1999). Phenolic compounds include tannins and colour pigments, anthocyanins which present at a higher level in red grape marc compared with white grape marc and are more likely to be found on the grape seeds (Springett, 2001; Palma & Taylor, 1999). The application of ultrasound at Food Science Australia has focused on the use of high-powered systems for extraction of bioactives. Principle targets have been polyphenols and carotenoids and in both aqueous and solvent extraction systems. The ultrasound extraction trials have demonstrated improvements in extraction yield ranging from 6 to 35%, as summarised in Table 2. Results of ultrasonically treated Shiraz and Sangiovese grape marc showed 17 and 35% increase in phenolic compounds respectively, However extraction of these compounds yielded much higher recovery from their respective seeds (Vilkhu, Food Science Australia unpublished data). Supercritical carbon dioxide extraction is proposed as a better method than ultrasound assisted extraction of polyphenolic compounds from grape seeds Palma and Taylor (1999). It was believed that the lower catechin (used as a measure of phenolic content) recovery from ultrasound method could be due to the insufficient power of the solvent used (aqueous methanol) or due to the degradation of samples during extraction process. Their study was focused on the efficiency of supercritical fluid extraction (SFE) rather than other methods used in the experiment. The

Table 2 Examples of bioactive ultrasound assisted extraction work completed at Food Science Australia Extract target Beta-carotene Polyphenols Polyphenols Polyphenols Gingerol Product Carrot Red grape marc Black tea Apple Ginger Solvent Aqueous Ethyl-acetate Aqueous Aqueous Aqueous Supercritical carbon dioxide Process Laboratory; 24 kHz, 207 W s ml 1 Laboratory; 24kHz, 2075 W s m 1 Laboratory; 24 kHz, 2075 W s ml 1 Laboratory; 24 kHz, 810 W s ml 1 Laboratory; 40 kHz, 2075 W s ml 1 Laboratory; 20 kHz Processing conditions Ambient Ambient Ambient Hot processing 90 C Hot processing 80 C Pressure 160 bar Improvement range (%) 1525 820 1135 618 6 30

results of catechin recovery using different extraction methods compared to a control (solvent extraction only) was not available, therefore it was not possible to determine whether ultrasound treatment (although having a lower recovery compared to SFE method) contributed to the increase in catechin recovery relative to a control. Most importantly though, the frequency of ultrasound and other extraction conditions (e.g. temperature) was not stated, therefore it was not known whether suitable frequencies or application conditions were used. In recent years it has been shown that pressurized hot water extraction methods offered higher phenolic compound recovery when compared to UAE, hydro-distillation and maceration with 70% ethanol (Ollanketo, Peltoketo, Hartonen, Hiltunen, & Riekkola, 2002). The use of methanol during UAE produced the lowest recovery with results not statistically different from maceration with 70% ethanol. Potential exists for combining ultrasound as an adjunct with the other extraction procedures to improve efficiency and yield. More recently, Tedjo, Eshtiaghi, and Knorr (2002) studied the quality attributes of grape juices for wine-making using non-thermal processes including ultrasound. The non-thermal processes examined offered a suitable gentle-action alternative to other cell breakdown methods with increased grape juice yields. Quality analyses (e.g. sugar, anthocyanins and mineral concentration, acidity, colour) showed that non-thermally processed juices had superior quality to untreated samples and comparable quality to that of enzyme treated grape juices. Likewise significantly enhanced contents of tea polyphenols, amino acid and caffeine in tea infusions were recovered with ultrasound assisted extraction when compared with conventional extraction. The sensory quality of tea infusion with ultrasound assisted extraction was better than that of tea infusion with conventional extraction (Xia, Shi, & Wan, 2006). 2.3.2. Anthocyanins Anthocyanins are enjoying greater prominence due to increasing public concern with the use of synthetic colouring agents. Anthocyanins represent a large group of water-soluble plant pigments based on the 2-phenylbenzophyrylium (flavylium) structure and there are more than 200 compounds in this category (IPCS, 2001). Anthocyanins are the main colour pigments in wild fruits and berries, and predominantly found in the sap of mature cells in grape skin (Springett, 2001). The pigments present in grape skin consist of di-glucosides, mono-glucoside, acylated monoglucosides and acylated di-glucosides of peonidin, malvidin, cyanidin, petunidin and delphinidin. Anthocyanins content in grapes varies from 30750 mg/100 g (Birdle & Timberlake, 1997). The wide variation in amount of these

compounds is greatly dependent upon cultivar, season, growing conditions, degree of ripeness, storage conditions as well as extraction procedures (Cacace & Mazza, 2003). UAE of crushed Shiraz and Merlot grapes by Food Science Australia showed 15 18% increase in total colour in grape juice (unpublished data). A study has been conducted on the potential to use microwave and ultrasound treatments for the extraction of pigments from strawberries. Optimal extraction was achieved using microwaves at 624 W, with a treatment time of 60 s, together with ultrasonic processing for 40 s and a ratio of material and extraction solvent of 1:6. The stability of the pigment extracts was considerably affected by pH, and achieving a maximum at pH 5.0. Addition of sucrose or heating at temperatures up to 80 C had little effect on pigment stability. However, pigment stability and colour were greatly improved by addition of citric acid (Cai, Liu, Li, & An, 2003). 2.3.3. Tartaric acid Tartaric acid occurs naturally in fruits, and found in high concentrations in grapes and tamarind (Springett, 2001). Approximately 90% of the total organic acids in grapes are tartaric and malic acids. Tartaric acid is a by-product in the wine industry since a tremendous amount of tartaric acid from lees has to be removed from the wine after yeast fermentation. Tartaric acid is widely used in bakery operations, wine production, pharmaceutical industry, hardening of gypsum, confectionery processing and in the chemical industry. Palma and Barroso (2002) optimized the UAE conditions for the recovery of tartaric and malic acid from red and white variety grapes for quantitative determination in wine-making by-products. Our studies on UAE of tartaric esters from red grape marc yielded an increase 16 to 23% from two different varieties (Vilkhu, unpublished). 2.3.4. Aroma compounds Over a period of many years it has been shown that ultrasound could be used to extract aromatic chemicals, which impart bouquet to the wines (Cocito, Gaetano, & Delfini, 1995). Solvent mixtures of n-pentane and diethyl-ether (1:2) and dichloromethane were used to study the optimization of the sonication extraction process. This study emphasised that UAE improved extraction efficiency with increased reproducibility of most aroma compounds compared to conventional extraction (Vila, Mira, Lucena, & Fernandez, 1999). An evaluation of UAE of isoflavones from ground soybeans was undertaken by Rostagno, Palma, and Barroso (2003), the efficiency of the extraction was improved by 15% but this was dependent on the organic solvent used. Notably 4060% water

was required to improve the extraction efficiency, which was thought to be due to the relative polarity of the isoflavones and increased ultrasound propagation in aqueous systems. Some aromatic compounds such as rutin from the flower buds of Chinese Scholar Tree (Sophora japonica) have improved considerably with higher levels of organic solvent compared to aqueous conditions. The difference in performance was attributed to hydroxyl radical and hydrogen peroxide formation in aqueous conditions resulting in degradation of the rutin. The application of ultrasound in methanol was considered more effective due to the higher solubility of rutin in methanol and hydrogen peroxide is not formed by ultrasound in methanol (Paniwynk, Beaufoy, Lorimer, & Mason, 2001). In order to extract phycocyanin from Spirulina platensis (Arthrospira platensis) cells, selection of ultrasonic frequency was important (Furuki et al., 2003). The purity of phycocyanin in its crude extract was dependant on ultrasonic frequency. For example, phycocyanin was extracted with higher purity at 28 kHz than at 20 kHz, due to the selective extraction of the active component at these frequencies. It was suggested that rapid and selective extraction of phycocyanin from S. platensis may be possible if an optimized ultrasonic application is developed. 2.3.5. Polysaccharides and functional compounds The extraction of carbohydrates, polysaccharides and other functional compounds has been studied in the recent years. Various extraction procedures with and without a short application of ultrasound at the beginning of the extraction were used to examine the effect of sonication on the extractability of the hemicellulose components of buckwheat hulls (Hromadkova & Ebringerova, 2003), cellulose from sugarcane bagasse (Sun, Sun, Zhao, & Sun, 2004), and xyloglucan from apple pomace (Caili et al., 2006). UAE of these compounds not only accelerates the extraction process but also preserves structural and molecular properties. In sugar cane bagasse hemicellulose extraction processes, UAE improved extractability of hemicelluloses apparently by destruction of cell walls and cleavage of links between lignin and the hemicelluloses (Jing, RunCang, Xiao, & YinQuan, 2004). Whereas ultrasonic aqueous extraction of polysaccharides from edible fungus, Pleurotus tuberregium, resulted in the formation of glycanchitin complexes with higher average molecular weight than compounds obtained by hot water extraction (Mei, Lina, Chi-KeungCheung, & Eng-Choon-Ooi, 2004), which could be due to the sonochemical modification of two polysaccharides. Further improvement in immunological as well as anti-tumour activities of these complexes were reported on animal trials. UAE can enable extraction at lower temperatures, Xu, Zhang, and Hu (2000) have compared UAE with hot water extraction of flavonoids from bamboo leaves. The laboratory scale trials results showed that the optimal conditions for extraction were achieved using UAE at lower temperature, rather than using hot water bath extraction at 80 C. More recently, Rosngela et al. (2007) investigated the chemical composition of Mate tea extracts (leaves of Ilex paraguariensis, a native tree from Brazil). The effect of the ultrasonic treatment resulted in improved mass yield of caffeine and palmitic acid in methanol

solvent. Ultrasound enhanced both the kinetics and yield which was attributed to increase in the internal diffusion that controls the transfer of solute to the solvent and also the destruction of pores in which the solute can be trapped. However the efficiency of the extraction will be dependent on the concentration of the methanol solvent employed Rostagno et al. (2003). 2.4. Bioactive extraction from animal materials There is limited number of publications on UAE from animal material. Attempts were made to extract chitin from fresh water prawn shells (Kjartansson, Zivanovic, Kristbergsson, & Weiss, 2006) and lutein from egg yolk (Xiaohua, Zhimin, Witoon, & Joan, 2006) by using sonication. In chitin studies from prawn shells, it was found that the chitin yield decreased during sonication, this loss was attributed to depolymerization of extracted chitin in the wash water. Subsequently, the degree of acetylation of chitins was unaffected by sonication, but the degree of acetylation of chitosans produced from sonicated chitin decreased. Egg yolk is one of the major lutein sources in our foods (Johnson, 2004). Lutein in egg yolk is highly bio-available, compared with other sources. It was reported that egg yolk intake significantly increased plasma lutein (Handelman, Nightingale, Lichtenstein, Schaefer, & Blumberg, 1999). Recently, Xiaohua, et al. (2006) have reported higher extraction yield of luetin when ultrasonic used in combination of saponificated organic solvent. Further to their report, compared with the traditional saponification solvent extraction method, the UAE extraction method was more effective in extracting lutein from the sample matrix, presumably by avoiding degradation reactions. 3. Extraction mechanisms and process development Extraction enhancement by ultrasound has been attributed to the propagation of ultrasound pressure waves, and resulting cavitation phenomena. High shear forces cause increased mass transfer of extractants (Jian-Bing et al., 2006). The implosion of cavitation bubbles generates macro-turbulence, high-velocity inter-particle collisions and perturbation in micro-porous particles of the biomass which accelerates the eddy diffusion and internal diffusion. Moreover, the cavitation near the liquid solid interface sends a fast moving stream of liquid through the cavity at the surface. Cavitation on the product surface causes impingement by micro-jets that result in surface peeling, erosion and particle breakdown. This effect provides exposure of new surfaces further increasing mass transfer. This phenomenon was confirmed by performed scanning electron micrography on peppermint plant leaves and trichomes. After these were ultrasonically treated for menthol extraction, microscopy results indicated that there were two mechanisms involved in extraction: (a) the diffusion of product through the cuticle of peppermint glandular trichomes and (b) the exudation of the product from broken and damaged trichomes (Shotipruk, Kaufman, & Wang, 2001). Acceleration in the extraction kinetics and improved extraction yield of pyrethrine from pyrethrum was largely

attributed to ultrasonics increasing the intra-particular diffusion of the solute, considered the rate limiting step (Romdhane & Gourdon, 2002). If the substrate is dry then ultrasound may be used to facilitate swelling and hydration and cause enlargement of the pores of the cell wall (Vinatoru, 2001). Diffusion through the plant cell walls, disruption and washing out of the cell contents were also attributed to improved extraction performance. The corresponding reduction in the size of the vegetal material particles by ultrasound disintegration will increase the number of cells directly exposed to extraction by solvent and ultrasonic cavitation (Vinatoru, 2001). Intensive ultra-sonication can also serve the purpose of reducing the particle size in tomato juice (Food Science Australia unpublished data). As large amplitude ultrasound waves pass through a mass media, cavitational bubble collapse can occur in close vicinity or at the surface of the plant membranes causing microfractures (Vinatoru, 2001). The occurrence of microfracture by ultrasound was demonstrated in soybean flakes (Haizhou et al., 2004). Cavitation collapse can occur on the plant surfaces, resulting in a micro-jet directed into the solid surface. Cavitation at cell surfaces has the ability to punch holes through cell wall as recently demonstrated with studies of bacterial cell sonication (Ugarte-Romero, Feng, Martin, Cadwallader, & Robinson, 2006). Preferentially micro-jetting will occur onto hydrophilic particle surfaces (Arora, Claus-Dieter, & Knud, 2004). Variation in the extraction yield from different plant varieties may result from structure, rheology (hardness of the seed structure) or the compositional differences resulting in varying degrees of susceptibility to ultrasound shock waves and likelihood that cavitation bubble will contact with the plant surface causing micro-jetting (Haizhou et al., 2004). Factors such as plant tissue turgor and the mobility of particles such as starch granules within the cell cytoplasm can be expected to influence ultrasound energy dispersion and extraction effectiveness (Zhang, Niu, Eckhoff, & Feng, 2005). In the study on supercritical fluid extraction enhancement by ultrasound Balachandran et al. (2006) they were able to demonstrate that the effectiveness of ultrasound was gained by the increase in the superficial mass transfer and that effectiveness declined sharply after the readily accessible surface solute had been removed. However, by reducing the substrate particle size major gains in extraction efficiency and extraction time reduction could be achieved. Solvent selection is usually based on achieving high molecular affinity between the solvent and solute. When ultrasound is also applied the cavitation will be affected by the physical properties of the solvent. Cavitation intensity decreases as vapour pressure and surface tension are increased. Haizhou et al. (2004) demonstrated this phenomenon in soybean oil extraction where greater UAE was achieved by isopropanol compared with hexane, the later having approximately 5-fold higher vapour pressure. 4. Adjunct processes During extraction, ultrasound may also achieve adjunct processes, whereby the food extract, ingredient or product

functionality may be modified by physical and sonochemical mechanisms. One such modification has been reported by Cravotto, Binello, Merizzi, and Avogadro (2004) in rice bran wax conversion to policosanol (common name for a mixture of C24C34 linear saturated fatty alcohols), a rich source of nutrients and pharmacologically active compounds. Both the first bran fraction from rice polishing and the discarded wax from the manufacture of rice oil were convenient and profitable starting materials for the production of policosanol. In the date syrup industry, ultrasound was applied for improving the quantity and quality of the syrup extraction. Entezari, Nazary, and Khodaparast (2004) successfully optimized ultrasonic processing conditions in laboratory trials which lead to a higher extraction in a shorter time with improved physical quality of the date syrup extract. Most importantly, the sonication significantly decreased the microbial count in comparison to the conventional method. This study also confirmed the presence of anti-microbial substances in date fruit, and that ultrasonic waves can accelerate their effects. The anti-oxidative activity provided by phenolic compounds has been shown to inhibit the oxidation of low-density proteins (Frankel, Waterhouse, & Teissedre, 1995). Resveratrol (trans-3, 5, 4-trihydroxystilbene), a stilbene phyto-alexin, is a phenolic compound possessing anti-oxidant activity. Resveratrol has been shown to provide health-promoting activities such as lowering the incidence of coronary heart disease and provide cancer chemo-preventive activity (Frankel, Waterhouse, & Kinsella, 1993; Jang et al., 1997). The combined use of ultraviolet light and ultrasound treatments on peanut kernels was reported (Rudolf & Resurreccion, 2005) for the elicitation of trans-resveratrol, total phenolic compounds, and anti-oxidant activity. A short exposure of ultrasound (4 min) to sliced peanuts and further incubation for 36 h at ambient temperature resulted in an 8-fold increase of trans-resveratrol as compared to untreated control samples. It was also reported that the anti-oxidative activity in stressed peanuts was negatively correlated with transresveratrol concentration, indicating that as anti-oxidant activity decreased trans-resveratrol concentration increased. To potentially replace the conventional destructive extraction process of menthol extraction from peppermint plants Shotipruk et al. (2001) studied the feasibility of using ultrasound to extract menthol from biologically viable peppermint plants (Mentha xpiperita). The results showed that plants ultrasonicated for 1 h at 22 C in a standard 40 kHz ultrasonic bath released approximately 17.8 g of menthol per gram of leaf tissue (2% of total product). The amount of menthol release increased with the time of treatment and was greatly affected by the temperature of the ultrasonic bath water. An increase from 2% to 12% of total product was observed when the temperature was increased from 22 C to 39 C. When the temperature effects were isolated, the mechanism of the product release was found to be that of cavitation. The treated plants remained viable and were ready for the subsequent ultrasound extraction after approximately 4 days of recuperation. However, the amount of product released was reduced in subsequent extractions. This study has shown the possibility of using an online ultrasonic, non-destructive extraction method to continuously release intracellular

plant metabolites from the plants while maintaining the plant's viability. The application of ultrasound treatment to yellow dent corn at different points in the conventional wet milling process enhanced starch separation, providing an increase in final starch levels of 6.35 to 7.02% (Zhang et al., 2005). The starches produced by ultrasonic treatments showed a significant increase in whiteness and decrease in yellowness that were comparable to starches produced by conventional wet milling. The ultrasound-treated starches exhibited higher paste viscosities. These viscosity changes during ultrasound treatments to starch granules in the slurry were attributed to the phenomenon of cavitation. The intense ultrasound treatment which generated localized spots of very high temperature and pressure might lead to configurational modifications of the granular structure, which could be in the forms of diffuse erosion or pitting of the starch granules as earlier observed by Degrois, Gallant, Baldo, and Guilbot (1974). The cork taint is one the major problem in wine corks. Trichloroanisoles (TCA) a natural contaminant chemical during processing of corks is responsible for wine spoilage. There is a limited efficacy of conventional washing processes for removal of TCA. The Ultracork process involving UAE of TCA, followed by application of a silicone barrier coating has provided an improved approach to overcome cork taint (Rowe, 2003). 5. Industrial extraction application design The use of ultrasound in food processing has been reviewed by Mason, Paniwynyk and Lorimer (1996). Recently, the design of ultrasound processing equipment has advanced to provide industrially robust processing capability. Enabling design and operational features have included; (a) automated frequency scanning to enable maximum power delivery during fluctuation of processing conditions, (b) non-vibrational flanges on sonotrodes for construction of high-intensity inline flow-cells and (c) construction of radial and hybrid sonotrodes to provide greater range in application design and product opportunities. Presently, 16 kW is the largest available single ultrasound flowcell, which can be configured in-series or in parallel modules. Industrial ultrasound manufactures within the last 2 years have promoted industrial processing capability for food extraction applications (Hielscher, 2006). Several ultrasound reactor designs have been described by Chisti (2003) and Vinatoru (2001), the latter specifically for industrial extraction of plant tissue. These included (a) stirred ultrasound horn (sonotrode) directly immersed into stirred bath or reactor, (b) stirred reactor with ultrasound coupled to the vessels walls and (c) recycling of product from stirred reactor through an external ultrasonic flow-cell. These configurations may provide both intermittent and continuous ultrasound exposure, from low intensity in a large volume reactor (0.01 to 0.1 W/cm3) to high intensity (1 to 10 W/cm3) in an external flow-cell. Mixed frequency reactors have been shown to offer advantages with respect to process efficiency and energy distribution (Moholkar, Rekveld, & Warmoeskerken, 2000; Swamy & Narayana, 2001; Tatake & Pandit, 2002; Feng, Zhao, Zhu, & Mason, 2002; Delgadino, Bonetto, & Lahey Jr., 2002). Reactor

geometries that are asymmetrical and polygons preferably with odd numbered sides using swept frequencies are also reported to be more effective (Gogate, Mujumdar, Thampi, Wilhelm, & Pandit, 2004; Puskas, personal communication). Modern ultrasonic systems include automated frequency scanning which adjusts operation of the system to the optimal frequency to ensure that maximum power is transmitted to the extraction vessel. The benefit of automated frequency scanning as opposed to a fixed frequency was demonstrated by Romdhane and Gourdan (2002) where the former achieved a 32% increase in pyrethrine extraction and a 30% increase in power delivered to the product. The presence of a dispersed phase contributes to the ultrasound wave attenuation. The active sonication region in a reactor is restricted to a zone located at the surface of the probe. Where it is not a disadvantage to extract oily materials as stable emulsions, ultrasound can be used to carry out aqueous extraction of oily materials with yields of the order of 50% (Food Science Australia, unpublished results). To improve effectiveness the material to be extracted should be reduced to as smaller particle size as practical without denaturing the material to be extracted and commensurate with separation from the solvent post extraction. If this is done very high yields and extraction rates are possible with ultrasonic augmentation of the extraction process (Balachandran et al., 2006). The proposed benefits of UAE for the food industry include, (a) overall, enhancement of extraction yield or rate, (b) enhancement of aqueous extraction processes where solvents cannot be used (juice concentrate processing), (c) providing the opportunity to use alternative (GRAS) solvents by improvement of their extraction performance, (d) enable sourcing/substitution of cheaper raw product sources (variety) while maintaining bioactive levels and (e) enhancing extraction of heat sensitive components under conditions which would otherwise have low or unacceptable yields. 6. New opportunities for UAE in the food industry There is an opportunity to capture new intellectual property in the area of ultrasound processing particularly where the technology can provide commercially attractive advantages and outcomes unique to ultrasound processing. Ultrasound has the unique capacity to both enhance extraction from substrates while simultaneously encapsulating the extracted substance with an encapsulate material in the extraction fluid by hydroxyl radical initiated covalent bonding and microsphere formation. To successfully accomplish this, the encapsulating material should have a higher reductive potential than the material being extracted and be relatively more hydrophobic. Preferably a mixed frequency ultrasound field is used, a relatively low frequency to facilitate extraction and a higher frequency under independent amplitude control to facilitate hydroxyl radical production for cross linking and microsphere formation. Proteins are suggested encapsulants as the sonochemistry and conditions favouring sphere development have been established. Vessel geometries, frequency combinations and frequency modulation to achieve the desired outcomes on a large scale suitable for scale up to industrial application would need to be explored and optimized.

7. Conclusion State of the art in UAE can achieve worthwhile gains in extraction efficiency and extraction rate, which if realised on industrial scale would represent worthwhile economic gains. Ultrasonic equipment engineering is such that it is commercially viable and scaleable to consider industrial-scale ultrasonic aided extraction. Potential exists for applying UAE for enhancement of aqueous extraction and also where organic solvents can be replaced with generally recognised as safe (GRAS) solvents. UAE can also provide the opportunity for enhanced extraction of heat sensitive bioactive and food components at lower processing temperatures. There is also a potential for achieving simultaneous extraction and encapsulation of extracted components to provide protection through the use of ultrasonics. Acknowledgement This work was supported by CSIRO - Food Science Australia, Food Futures Flagship. This work was partially presented at Food innovation: Emerging Science, Technologies & Application (FIESTA), 3rd Innovative Foods Centre Conference held at Melbourne, Australia on 1617 October, 2006. References
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