Patent Description:
Consumer's demand for minimally processed, high quality products has increased remarkably in the recent years. Market preferences have shifted towards fresh, healthy and ready-to-eat foods that are not only rich in flavour but also have organoleptic properties matching as closely as possible a fresh product. Prolonged shelf life, preferably without the use of preservatives and colourants, is also appreciated.

Fruit and vegetable purees are adequate products for feeding infants due to their consistency and nutritional value. In stores, fruit and vegetable purees for young infants are readily available. These purees are typically stored at ambient temperature and have a shelf life of several months. To enable such an extended shelf life at ambient temperature, the fruit and vegetable purees should be treated with a sterilization process that ensures microbiological safety. However, the use of high temperatures in thermal treatment often negatively affects the nutritional and sensorial proprieties of food products.

Food purees require different types of sterilization depending on the pH of the ingredients. For instance, the pH of fruits is typically more acidic than the pH of vegetables, which also determines the type of microflora present therein. Vegetable purees are typically sterilized at higher temperatures as compared to fruit purees since vegetable purees are less affected by the exposure to high temperatures. For fruit purees, it is important that the temperature during sterilization remains low since thermal processing will negatively affect the appearance, taste and colour thereof.

Currently, retort heating is widely used in the food industry to produce such products. Retort heating includes heating the food that is sealed in a container using steam or other heating methods. The holding times and high temperatures (<NUM>-<NUM>) in retort heating often have a negative impact on the nutritional value and the sensory as well as organoleptic aspects of the food in the container.

The use of high-pressure processing (HPP), also known as pascalization, in food industry was developed as an alternative to common thermal processes, such as pasteurization and sterilization, to produce a microbiologically safe product. In traditional HPP, food products are sealed and placed into a steel compartment containing a liquid (often water), after which pumps are used to create the necessary pressure. Pressure is increased at a certain rate until the target pressure is reached, kept for a specific amount of time and subsequently released at a specific rate. Typical pressures applied to foods range from <NUM> - <NUM> MPa. Although heat may also be applied during HPP, high-pressure processes are normally conducted at refrigerated temperatures. However, due to adiabatic heating, temperatures do naturally increase during a typical HPP treatment. The magnitude of the temperature increase is dependent on the target pressure and food composition. Taking water as an example, the temperature increase is of approximately <NUM> per <NUM> MPa. <NPL>, teaches a high pressure treatment of mango puree at <NUM> MPa at a temperature of <NUM> for <NUM> minutes that could be a feasible treatment to reduce enzymatic activity and preserve fresh-like quality attributes.

Although HPP may be effective for killing microorganisms in food products, spores of some bacteria are pressure-tolerant and normally resist to the HPP treatment. This is not acceptable for products designed for infants.

acidoterrestris is a non-pathogen, spore-forming spoilage bacterium typically present in fruit-containing products. Food spoilage caused by this bacterium is characterized by a bad taste/aroma, which renders food product unsuitable for consumption. acidoterrestris spores show extremely high thermal resistance depending on the type of food matrix, soluble solids content and pH. acidoterrestris D95 values (time in minutes during which the number of living cells decrease by <NUM>%, at <NUM>) typically range from <NUM> to <NUM>, depending on the type of food. The standard pasteurization process using temperatures of <NUM>-<NUM> for a period of time of about <NUM> minutes at ambient pressure, which is aimed at destroying vegetative pathogens, is therefore ineffective against spores of A. acidoterrestris.

HPTS (high pressure thermal sterilization) combines the effects of elevated temperatures (<NUM> to <NUM>) and pressures above or equal to <NUM> MPa to achieve inactivation of bacterial spores. For HPTS treatment, the product needs to be preheated to <NUM> to <NUM> and, through compression heating during the pressure build-up, the process temperature can reach up to <NUM> °<NPL>) inoculated baby food puree with spores of <NUM>. amyloliquefaciens or G. stearothermophilus to a spore count of <NUM><NUM> to <NUM><NUM> and exposed them to a temperature range of <NUM>-<NUM> at <NUM> MPa. The vegetable puree comprised: carrots <NUM>%, peas <NUM>%, zucchini <NUM>%, water <NUM>%, and <NUM>% salt. The Aw-value of the puree was <NUM>, the pH <NUM>, and dry matter content <NUM>%.

<NPL>) inoculated orange juice with spores from A. acidoterrestris to yield an initial count of ca. <NUM><NUM> spores cfu/mL. The packaged orange juice samples (<NUM> °Brix, pH=<NUM>) were submitted to simultaneous high pressure - mild temperature processes with pressures of <NUM> MPa and <NUM> MPa, temperatures of <NUM>, <NUM> and <NUM> and processing times between <NUM> and <NUM>. The high-pressure processing conditions investigated (<NUM> MPa, <NUM> MPa) resulted in a reduction in the microbial numbers, in particular at the higher temperature of <NUM>, where approximately <NUM> log reduction of A. acidoterrestris levels were registered after processing for <NUM>. This reduction is not sufficient from an infant food safety point of view as there is still <NUM><NUM>-<NUM><NUM> spores cfu/ml A. acidoterrestris remaining in the orange juice samples.

<NPL>) describes the preparation of a strawberry mousse. Fresh strawberries were washed twice, cut into small pieces, citric acid and saccharose were added as preservative and for acidity and °Brix correction respectively, and finely homogenized using an electric blender. The samples were stored under refrigerated conditions before the high-pressure experiments. In the high-pressure treatments, the processing conditions were set at <NUM> MPa, <NUM>, <NUM> for the strawberry mousse samples. At the end of the treatment, the samples were stored at <NUM>.

It is an object of the present invention to provide a fruit-containing product for infants that is microbiologically safe, without the use of antimicrobial preservatives, and is suitable for storage at ambient temperatures, while at the same time maintaining a natural appearance, with an appealing colour and taste.

The inventors have surprisingly found that a fruit-containing product, selected from baby foods, fruit purees, fruit spreads, slurries, mousse/mousselines, compotes, creams and jams, which is microbiologically safe, can be obtained by a method including the steps of:.

The method according to the invention allows the preparation of a fruit-containing product that is microbiologically safe even for baby food standards as compared to similar compositions prepared by usual thermal treatment, having extended shelf life at ambient temperatures. Surprisingly, the inventors have found that the method according to the invention is particularly effective to drastically reduce the number of undesired bacterial spores, such as Alicyclobacillus acidoterrestris.

Advantageously, unlike thermal treatments in the art, the fruit containing product prepared by the method of the invention presents a more natural appearance (particularly, an improved colour) than similar products prepared by other thermal treatments of the art. The product prepared by the method of the invention thus more closely resembles a homemade fruit based product in terms of organoleptic characteristics if compared to fruit based products sterilized with standard pasteurization.

The invention further relates to a fruit-containing product that is obtainable by the method according to the invention.

In a first aspect, the present invention relates to by a method of preparing a fruit-containing product containing less than <NUM><NUM> heat-resistant spores per gram of the fruit-containing product, wherein the method comprises the steps of:.

wherein the fruit-containing product is selected from baby foods, fruit purees, fruit spreads, slurries, mousse/mousselines, compotes, creams and jams.

'Fruit-containing product', as used herein, relates to an edible product suitable for human consumption, preferably but not limited to, baby consumption. Examples of fruit-containing products in the context of the invention include 'baby food', fruit purees, fruit spreads, slurries, mousse/mousselines, compotes, creams, jams, and the like. Herein, 'baby food' (interchangeably: 'first diet food') refers to weaning foods having the common meaning in the art as defined, for instance, in the Commission Directive <NUM>/<NUM>/EC. Baby foods are normally used as a supplement to the baby diet (which may be otherwise only comprised of animal milk and/or infant milk formula/growing up formula/young children formula) as infants progress onto a mixed family diet and/or in their progressive adaptation to ordinary food.

'Fruit', in the context of the present invention, has the usual meaning in the art and is thus defined as a product obtained from plants associated with seeds. In the context of the invention, 'fruit material' refers to fruit or a mix of two or more fruits, preferably in the form of pure fruit puree, pure fruit concentrate and/or pure fruit pulp. 'Pure fruit' herein refers to fruits without any additives. Suitable fruits for the preparation of the fruit-containing product according to the method of the invention include, but are not limited to, apples, apricots, bananas, berries, blueberries, cherries, clementines, cress, elderberries, grapes, grapefruit, lemons, mangos, oranges, papaya, peaches, pears (e.g. Williams pear), pineapples, plums, raspberries, rhubarb, sorrel, strawberries, and the like. In the context of the present invention tomatoes are not considered a fruit.

Methods of determining pH and total solids content in the fruit-based composition are usual to the skilled person. For instance, the pH value of the fruit based composition used in the method of the present invention can be determined using e.g. a pH-meter by placing the probe directly in the fruit composition, without dilution in water, at ambient temperature.

As used herein, "total solids" has its usual meaning in the art, e.g., it refers to the carbohydrate and cellulose contents of a fruit based composition, most of which is insoluble. Total solids of the fruit-based composition can be calculated as a sum of total solids provided for each of ingredients of the composition.

As described herein, the term "Brix relative density", "Brix", or "°Bx", refers to a well-known hydrometer scale for measuring the soluble solids content of a liquid as determined by a refractometer calibrated at <NUM>. The Brix scale measures the number of grams of sugar present per <NUM> grams of aqueous sugar solution (the total solubilized solid content). For example, a measurement of <NUM> °Bx refers to <NUM>/ml of sugar in solution. For reference, packaged orange juices normally have around <NUM> °Bx. Soluble solids (°Bx) in the method according to the invention can measured using a refractometer MS REF <NUM> My-Soft, calibrated at <NUM>.

Water activity ('Aw') is measured by common techniques known in the art. For instance, water activity can be measured using AQUALAB 3TE water activity meter (Decagon Device Inc.

Herein, 'viscosity' can be determined at <NUM> using e.g. the RM180 Rheomat Mettler Toledo device. By way of comparison, the viscosity values refer to shear rate of <NUM> sec-<NUM>, unless otherwise indicated. The measurement system, consisting of a tube, filled with the sample (puree or beverage) is coupled with a rotating bob which is immersed in the bath of an F12 refrigerated/heating circulator (Julabo Labortechnik GmbH, Seelbach, Germany) at <NUM>. The inner cylinder or bob starts rotating from <NUM> shear and ramps up to higher shear rates. Preferred fruit-containing products prepared by the method of the invention are fruit purees, which are commonly a non-Newtonian fluid, particularly a shear thinning fluid. A non-Newtonian fluid is not viscosity constant but varies depending on the shear rate. That is to say, these fluids deform or flow in accordance with a shear rate, or the rapidity at which pressure, such as a squeezed tube, is applied. In a shear thinning fluid (such as ketchup), fluid viscosity decreases with increasing shear stress.

According to one embodiment, the pH of the fruit based composition is between <NUM> to <NUM>, preferably <NUM> to <NUM>, most preferably <NUM> to <NUM>. The fruit-containing product prepared by the method of the invention also preferably has a pH between <NUM> and <NUM>, more preferably <NUM> to <NUM>, preferably <NUM> to <NUM>, most preferably <NUM> to <NUM>, since the pressure/temperature treatment of the invention does not substantially alter the pH of the fruit based composition.

The method of the invention is particularly effective in reducing heat-resistant spores at the pH range of <NUM> to <NUM>, which allows other processing parameters, in particular time and temperature, to be applied in the lower ends of the ranges. Accordingly, if necessary, the pH of the fruit based composition of step (a) may be adjusted to the ranges defined for the present method in case the fruit material provides a pH of below <NUM> or above <NUM>. For instance, when the fruit material comprises only or substantially only banana, the pH of the fruit material may be higher than <NUM>, thus requiring pH adjustment to the pH values of the present invention. Methods to adjust the pH are known to the person skilled in the art. According to one embodiment, the method comprises the step of adjusting the pH of the fruit material with a pH adjustment component prior to step (a). The pH adjustment component is acceptable in baby food products, and is preferably selected from fruit juice or fruit juice concentrate, more preferably lemon juice or lemon juice concentrate.

In a preferred embodiment, the total solids content of the fruit based composition starting product and/or the product as obtained is between <NUM>-<NUM> wt. %, preferably between <NUM>-<NUM> wt. %, more preferably between <NUM>-<NUM> wt. %, by weight of the product. According to one embodiment, the fruit-based composition and/or the fruit-containing product according to the invention comprises more than <NUM> wt. % of fruit material, on a total solids basis, more preferably more than <NUM> wt. % of fruit material, even more preferably more than <NUM> wt. % of fruit material, on a total solids basis.

In a preferred embodiment, the 'fruit material' in step (a) comprises one or more fruits selected from the group comprising apple, pear (e.g. Williams pear), berries, banana, cherry, mango, peach, apricot, pineapple and/or combinations thereof. In one embodiment, the 'fruit material' comprises apple and a second fruit selected from the group comprising pear (e.g. Williams pear), berries, banana, cherry, mango, peach, apricot, pineapple in a fruit weight ratio of <NUM>:<NUM>, preferably <NUM>:<NUM>, more preferably <NUM>:<NUM>. In another embodiment, the 'fruit material' comprises banana and a second fruit selected from the group comprising apple, pear (e.g. Williams pear), berries, cherry, mango, peach, apricot, pineapple in a fruit weight ratio of <NUM>:<NUM>, preferably <NUM>:<NUM>, more preferably <NUM>:<NUM>.

Preferably, the product prepared by the method of the invention is free of meat, fish, milk protein, egg protein, legumes (i.e., a plant in the Fabaceae family or a fruit or seed thereof) and/or vegetables, particularly free of meat, legumes and/or vegetables. In the context of the present invention, tomato is defined as a vegetable. Accordingly, in a preferred embodiment, the product is free of tomato, eggplant, potato, zucchini, and the like. More preferably, the product is free of tomato.

In a preferred embodiment, the fruit-based composition used in the method according to the invention has a percent soluble solids (°Bx) of at least <NUM> °Bx, more preferably at least <NUM> °Bx, even more preferably at least <NUM> °Bx. In a preferred embodiment, the method according to the invention does not substantially alter the percent soluble solids of the composition undergoing temperature and high pressure treatment. Thus, the product prepared by the method of the invention has substantially the same percent soluble solids as the initial fruit based composition used in step (a).

The fruit material may optionally be a mix of different fruits and or fruits purees that are blended to achieve the desired percent soluble solids (°Brix) and pH. The °Brix influences the perceived sweetness of the finished product and the pH influences the tartness or acidity of the finished product.

Preferably, the fruit based composition is in the form of a fruit puree (i.e., a spoonable fruit-based composition). As defined herein, 'puree' refers to the pulp of a product that has been crushed or homogenized in a substantially smooth and/or creamy condition, without a substantial amount of conglomerated pulp constituents as fragments or pieces. Transformation of fresh fruit(s) into the desired fruit-based composition may be suitably performed by means of various techniques known to the art (e.g., pulping, cooking, pressing). Accordingly, the present method may optionally include a pre-treatment step (prior to step (a)) that includes, but is not limited to, fruit preparation (e.g. washing, peeling off, etc.), blending or mixing, fruit cell fragmenting (e.g., by means of a homogenizer, sonolator, stone mill, high-shear apparatus, emulsifier or any other device capable of breaking or fragmenting intact fruit cells), removal of skins, stems, seeds, peels and the like. Alternatively, pre-blended puree and/or fruit juice concentrates may also be used. In one embodiment, the fruit puree is not a mousse. Mousses are characterized by having air bubbles incorporated in a smooth composition. Fruit purees are distinct from fruit juices, the water activity of which is too high, thereby providing a baroprotective effect when performing the method according to the invention.

Typically, the fruit-containing products according to the invention have a viscosity of <NUM> to <NUM> Pa. s, preferably <NUM> to <NUM> Pa. s, more preferably <NUM> to <NUM> Pa. s, as determined at a shear rate of <NUM> sec-<NUM> at <NUM>, using e.g. a RM <NUM> Rheomat Mettler Toledo device. The fruit-containing products are typically non-Newtonian, shear thinning fluid. Thus, the viscosity of the product will decrease as the shear rate increases. By means of comparison, when measured at a higher shear rate, the products according to the invention have a viscosity of <NUM> to <NUM> Pa. s, preferably <NUM> to <NUM> Pa. s, as determined at a shear rate of <NUM> sec-<NUM> at <NUM>, using e.g. a RM <NUM> Rheomat Mettler Toledo device. These viscosities are appropriate for a spoonable and/or squeezable fruit-containing product according to the invention. The fruit-based composition in step (a) is preferably homogenized. After homogenization, the fruit-based composition is subsequently pumped to a suitable food processor wherein step (b) is carried out.

Prior to step (b), the fruit based composition may be stored at refrigerated condition (<NUM>-<NUM>). Preferably, prior to step (b), temperature of the fruit based composition is brought to about ambient temperature (<NUM>-<NUM>).

Step (b) is a high pressure processing (HPP). The pressure applied in step (b) is between <NUM> MPa and <NUM> MPa, preferably between <NUM> MPa and <NUM> MPa.

The temperature applied in step (b) of the method according to the invention is between <NUM> and <NUM>, more preferably between <NUM> to <NUM> and even more preferably between <NUM> to <NUM>. In a further preferred embodiment, the fruit-based composition is not exposed to a temperature that exceeds <NUM>, more preferably <NUM>, before and/or after step (b) is executed.

In a preferred embodiment, step (b) is carried out for between <NUM> and <NUM> minutes, more preferably between <NUM> and <NUM> minutes and even more preferably between <NUM> and <NUM> minutes, most preferably between <NUM> and <NUM> minutes.

After step (b), the fruit-containing product is preferably packaged. Suitable packaging includes but is not limited to glass jars or squeezable tubes. The packaging is preferably sterile. Alternatively, packaging can occur prior to step (b), that is prior to temperature and high pressure treatment.

The present inventors have surprisingly found that the method according to the invention advantageously avoids the need for addition of food preservatives for fruit-containing products that are preferably stored at ambient temperature, while achieving excellent microbiological stability including against bacterial spores commonly found in food products. As used herein, "preservative" is described as an agent that is added to the product in order to enhance microbiological stability, preserve/retain flavour, colour, texture, cell wall structure, appearance, moisture, and the like.

The present method advantageously allows preparing fruit-containing compositions with improved microbiological stability (i.e., reduced bacterial spores) as compared to identical products prepared by usual thermal treatments, regardless of the presence of preservatives in the formulations. In one preferred embodiment, the method of the present invention includes the step of adding an edible antimicrobial preservative to the fruit-based composition before, during or after the step (b), preferably after step (b) is applied. Suitable antimicrobial preservative(s) are food grade, and are selected from sorbic acid, a sorbate (e.g. sodium sorbate), benzoic acid, a benzoate (e.g. hydroxybenzoate or sodium benzoate), parabens, nitrate, nitrite, citric acid, lactic acid, acetic acid, propionic acid, a propionate (e.g. sodium propionate), malic acid, tartaric acid, sulphur dioxide and sulphite. Preferably, the preservative is added at an amount of less than <NUM> wt. % based on total weight, more preferably less than <NUM> wt. %, even more preferably less than <NUM> wt.

Fruits may include natural preservatives (e.g., citric acid). However, according to one embodiment, no antimicrobial preservative is added to the fruit-based composition in the method according to the invention, particularly wherein the method is to prepare a baby food fruit containing product. More preferably, no antimicrobial preservative is added after step (b) is applied. Worded differently, in a preferred embodiment, the fruit-containing product according to the invention does not include added preservative(s) (i.e., does not include preservative(s) that is/are added to the composition and/or that is/are not present in the natural fruit ingredient(s) present in the product).

According to one embodiment, the fruit-based composition of step (a) comprises less than <NUM> wt. % of sugars by weight of the product, preferably less than <NUM> wt. % of sugar, even more preferably less than <NUM> wt. % sugar by weight of the product. As defined herein, 'sugars' refer to added sugar that are not naturally present in the fruit material, i.e., in the natural, pure fruit. Thus, 'sugars' as used herein refer to added sugar selected from sucrose, glucose, galactose, saccharose and combinations thereof, more preferably, sucrose. Accordingly, the method according to the invention does not comprise a step of adding sugar to the fruit based composition before, during or after step (b) of applying temperature and high pressure to the fruit based composition.

The composition in step (a) comprises at least <NUM> wt. %, based on total weight, of fruit material, preferably at least <NUM> wt. % fruit material, more preferably, at least <NUM> wt. % fruit material, most preferably at least <NUM> wt. % fruit material.

Typically, the composition and/or product according to the invention has a water content of up to <NUM> wt. % water, based on total weight, more preferably up to <NUM> wt. %, even more preferably up to <NUM> wt. % water, most preferably up to <NUM> wt.

In a preferred embodiment, the fruit-containing composition has a water activity (Aw) of at least <NUM>, more preferably at least <NUM>. Further, it is preferred that the fruit-containing composition does not have a water activity above <NUM>. Preferably, the product as obtained has a water activity between <NUM> to <NUM>, more preferably between <NUM> and <NUM>.

Preferably, the fruit-containing product according to the invention is dairy-free.

In a preferred embodiment, a thickener is present in the fruit-based composition. Preferred thickeners are selected from the group comprising starches, such as starch waxy maize, flours, preferably rice flour, rice, cereals and grains, pasta, and/or crushed/milled biscuits (biscuit flour), and/or gums such as xanthan gum or carrageenan gum. Preferably, thickeners are added in the fruit based composition in the method of the invention in an amount of less than <NUM> wt. % based on total weight, more preferably less than <NUM> wt. % based on total weight, even more preferably less than <NUM> wt. % based on total weight. In one embodiment, the fruit-based composition comprises <NUM> to <NUM> wt. % based on total weight of a thickener, preferably starch such as starch waxy maize. In the context of the invention gelatine is preferably not used as a thickener.

In a preferred embodiment, the method according to the invention includes the step of storing the final fruit containing product at ambient conditions, typically at a temperature in the range of <NUM> - <NUM> or even <NUM> - <NUM>. Preferably, the method comprises the step of storing the product at ambient conditions for at least <NUM> days, more preferably at least <NUM> months, even more preferably, at least <NUM> months at ambient conditions. In a most preferred embodiment, the product prepared by the method of the invention has a shelf-life of at least <NUM> months at ambient storage conditions, or even at least <NUM> months at ambient storage conditions.

According to yet another aspect of the invention, the method is for reducing microbial spoilage of a fruit-containing product, more specifically for reducing the number of heat resistant-spores in a fruit-containing product. Preferably, the method is for achieving at least a <NUM> log reduction, preferably at least a <NUM> log reduction and even more preferably at least a <NUM> log reduction in heat-resistant spores in the product. In a further preferred embodiment, the method achieves at least a <NUM> log reduction, preferably at least a <NUM> log reduction and even more preferably at least a <NUM> log reduction in heat-resistant spores, such as from Alicyclobacillus acidoterrestris, in the product. The method for reducing the number of heat-resistant spores in a fruit-containing product results in a reduction of the number of heat-resistant spores to below <NUM><NUM> spores / g, more preferably to below <NUM><NUM> spores / g or even below <NUM> spores / g. Most preferably, the amount of heat-resistant spores is below the detection limit.

In a second aspect, the present invention relates to a fruit-containing product obtainable by the method of the present invention.

The fruit-containing product prepared by the method of the invention has an energy density of between <NUM> and <NUM> kcal per <NUM>, preferably <NUM>-<NUM> kcal/<NUM> product.

The fruit-containing product according to the invention is preferably pourable, spreadable or spoonable, more preferably spoonable, which is ready-to-consume. The fruit-containing product in the context of the invention is selected from 'baby food', fruit purees, fruit spreads, slurries, mousse/mousselines, compotes, creams, jams, and the like. Alternatively, the spreadable fruit product may be used as a food ingredient or flavouring system in producing other products. For example, the product may be used as a substitute for milk or as an enhancement to such products as yogurt, ice cream or other suitable products employing fruit or fruit flavours.

Preferably, the fruit-containing product is shelf stable at ambient conditions for <NUM>-<NUM> months.

The 'fruit-containing product' according to the present invention is preferably intended to be consumed by a human infant, preferably having an age of between <NUM>-<NUM> months, more preferably between <NUM>-<NUM> months and even more preferably between <NUM>-<NUM> months. Thus, the fruit-containing product may also be referred to as a fruit-containing infant food or even a fruit-containing baby food, preferably an infant fruit puree or a baby fruit puree.

The fruit-containing product obtained by the method of the invention contains less than <NUM><NUM> heat-resistant spores per gram or even below <NUM><NUM> spores per gram. Preferably, the fruit-containing product contains less than <NUM><NUM>, more preferably less than <NUM><NUM> or even less than <NUM> spores of Alicyclobacillus acidoterrestris. Most preferably, spores of Alicyclobacillus acidoterrestris are substantially absent from the fruit-containing product. In a preferred embodiment, the fruit-containing product obtained by the method of the invention comprises at least <NUM> wt. % of a fruit material based on total weight, a pH of <NUM> to <NUM>, an energy density of between <NUM> and <NUM> kcal per <NUM>, and a viscosity of between <NUM> to <NUM> Pa. s at a shear rate of <NUM> sec-<NUM> at <NUM>.

All other limitations and definitions herein provided for the method according to the invention apply mutatis mutandis to the product obtained thereby.

The Saccharomyces cerevisiae strain DSM <NUM> used in this study was obtained from the German Collection of Microorganisms and Cells Cultures.

The Alicyclobacillus acidoterrestris strain DSM <NUM> used in this study was obtained from the German Collection of Microorganisms and Cell Cultures. This strain was isolated from apple juice by Cerny et al. acidoterrestris is a heat-resistant, acidophilic, spore-forming Gram-positive bacterium.

The stock cultures were kept immobilized on sterile beads at - <NUM> in a Cryobank.

A twenty-four-hour culture of S. cerevisiae DSM <NUM> strain grown in YPG-broth (<NUM>% yeast extract - Difco, <NUM>% peptone - Difco, <NUM>% glucose - POCh) was centrifuged at <NUM>°C for <NUM> at <NUM> x g, and the sedimented cells were aseptically re-suspended into phosphate-buffered saline (PBS, pH <NUM>) and again centrifugated. The washing procedure was repeated twice. The model suspensions of S. cerevisiae (<NUM>-<NUM> log cfu/mL) were prepared in PBS and dispensed in <NUM> portions in sterile plastic cryovials (Simport, Canada), in duplicate. Baby food formula was inoculated with S. cerevisiae cells (from the model suspension) to <NUM> log cfu/ g and transferred into bottles (Bionovo) in <NUM> portions, in duplicate.

A fresh culture suspension of A. acidoterrestris was spread-plated onto <NUM> potato dextrose agar plates with a pH of <NUM> (Oxoid). The inoculated plates, packed in sealable plastic bags to prevent drying, were incubated at <NUM>°C for ca. <NUM> days, until over <NUM>% spores had developed. The degree of sporulation was monitored by microscopy after staining with malachite green. The sporulated agar plates were harvested by flooding with <NUM> of sterile, deionized water and scraping with a spreading loop. The spores were washed by centrifugation three times (<NUM>,<NUM> rpm for <NUM> at <NUM>°C). Finally, the spore pellets were re-suspended in <NUM> of sterile deionized water and stored at <NUM>°C until use. The number of spores was evaluated after serial dilutions and subsequent incubation on BAT agar (Merck) for <NUM> days at <NUM>°C. Just before the experiments, spores were suspended in baby food formula at approximately <NUM> log cfu/g.

The baby food formula used in this study (<NUM>,<NUM> °Brix, pH <NUM>,<NUM>) consisted of the following infant-grade ingredients: apple puree <NUM>%, blueberry puree <NUM>%, Williams pear puree <NUM>%, rice flour <NUM>% and <NUM>% vitamin C. Purees as used were manufactured from sources approved for supply of baby-grade food and were free from foreign matter and any additives besides vitamin C (e.g. colourants, aromas, preservatives). The baby food as obtained is a puree or a spoonable composition as typically sold in the supermarkets where it has its own market segment. The water activity (Aw) was measured using AQUALAB 3TE water activity meter (Decagon Device Inc. Aw of the sample, as measured in triplicate, was <NUM>.

Samples were serially diluted in maximum recovery diluents (Merck), and spread on Sabouraud agar (Merck) and Sabouraud agar with <NUM>% NaCl (POCh). This was the maximum concentration of NaCl that caused no reduction in the colony count of unstressed cells of the S. cerevisiae DSM <NUM> strain. The plates were incubated at <NUM>°C for <NUM> days. The difference between the viable (cfu in Sabouraud agar) and non-injured cells (the corresponding cfu in Sabouraud agar with <NUM>% NaCl) was used to estimate injured survivors as based on Somolinos et al. , <NUM> [<NPL>].

The number of the surviving population was determined using the plate method on BAT-agar (Merck) through incubation for <NUM> days at <NUM>. Samples were tested just after the pressure treatment and after storage for the indicated number of days at ambient conditions (<NUM>-<NUM>).

The steps of the HPP procedure were executed using a high-pressure Laboratory-scale Food Processor, Type U4000, Production year <NUM>, manufactured by the Institute of High-Pressure Physics, Unipress Equipment Division.

The technical parameters that can be adjusted include a maximum pressure of <NUM> MPa, working temperature of between -<NUM>° and <NUM>, the sample chamber has a volume of about <NUM> Litre and a sample space of <NUM> diameter and height of <NUM>. The pressure-transmitting fluid used was distilled water and polypropylene glycol (<NUM>:<NUM>). Pressure is controlled by a fully automatized multi-cycle pressure process. A pressure of up to <NUM> MPa was generated in <NUM>-<NUM>; the release time was <NUM>-<NUM>. Data acquisition allows monitoring and recording of all important process parameters, including pressure in the vessel, temperatures in the vessel and in samples and are disclosed herein.

General sample processing scheme: Twenty-five grams samples in HDPE bottles (Bionovo) were exposed to high pressure treatment with <NUM>, <NUM>, or <NUM> MPa at temperatures <NUM>, <NUM> and <NUM> for <NUM>, <NUM> and <NUM>, followed by storage of the treated puree at ambient conditions for several weeks or even months during which at selected time points samples were taken for microbiological analysis to determine spoilage of the treated product. The pressurization times as mentioned do not include the time to ramp-up and the release of pressure. The assays were performed using two independent samples from two independently executed experiments. Unpressurized samples were used as controls. After treatment samples were removed from the chamber and stored in ambient condition at <NUM> - <NUM> for further analysis. Online temperature and pressure measurements showed that the process is well controlled and that no overshoot in the temperature of the sample as subjected occurs. For instance, at a temperature setting of <NUM>, a maximum online temperature was measured of <NUM>.

The puree can be scooped from a jar and fed to an infant using a spoon. Determination of dynamic viscosity at <NUM> was conducted using a RM <NUM> Rheomat Mettler Toledo device using usual protocol for viscosity measurement. Usual protocol for determining viscosity was applied: it includes a measurement system, consisting of a tube, filled with the sample (puree or beverage), coupled with the rotating bob, which is immersed in the bath of an F12 refrigerated/heating circulator (Julabo Labortechnik GmbH, Seelbach, Germany). The temperature applied during measurement was <NUM>. The bob speed was increased taking measurements at different shear rates (s-<NUM>).

Viscosity of the spoonable baby food/puree as prepared in <NUM>, and prepared according to the method of the invention (as described in item <NUM> with the following set-up <NUM>/<NUM> MPa/<NUM>), and stored at <NUM> for <NUM> months) was measured. The values of mean viscosity measured on <NUM> samples each are given below.

The inactivation of S. cerevisiae DSM <NUM> strains after HPP treatment in all types of applied conditions were <NUM> log and remained at this level during at least nine weeks of refrigerated or ambient storage (after <NUM> weeks the shelf-life test was terminated). Inactivation of bacterial A. acidoterrestris spores was also very effective under a variety of different conditions, resulting in a reduction of about <NUM> logs.

Good results were also obtained with testing at <NUM> MPa, a temperature of <NUM> for <NUM>, <NUM> and <NUM> minutes. Under these conditions, samples were stored for a period of between <NUM> and <NUM> days, giving sufficiently low cfu counts for various of the tested conditions.

Good results were also obtained with testing at <NUM> MPa, a temperature of <NUM> for <NUM>, <NUM> or <NUM> minutes, where already after <NUM> days of storage similar low cfu counts (<<NUM> or <NUM>) were recorded. Under these conditions, samples were also stored for up to <NUM> days giving such low cfu counts.

The high-pressure pasteurization process as executed according to example <NUM> allows to obtain a product with an adequate reduction of undesirable heat-resistant spores.

To validate the consumers' acceptance, several sensorial attributes of the fruit products obtained using the HPP process were compared with products obtained by conventional retort pasteurization technology. This conventional treatment is currently practiced to ensure products are obtained that can be stored at ambient conditions for prolonged periods of time while remaining safe for consumption by infants. Thus, the same fruit-based formula as manufactured in a factory obtained using conventional thermal treatment for commercial production was used for comparison purposes.

Sensory attributes as tested included colour, appearance, taste, smell and consistency were assessed according to the six-point scale characteristic by certified sensory food panel experts from Institute of Agriculture and Food Biotechnology, Warsaw, Poland (accreditation no AB <NUM>, granted by PCA). Each time no less than seven experts were independently assessed samples. During all stored periods, the HPP-treated products as obtained were given a higher value for all assessed attributes compared to the conventionally processed recipes.

Colorimetric measurements were performed with the CR-<NUM> Chroma Meter (Minolta, Osaka, Japan) with <NUM>° circumferential illumination and <NUM>° viewing angle geometry measurement, aperture size Ø <NUM> and D65 light source (pulsed xenon lamp). The CIE Lab (CIE L*a*b*) colour scale specified by Commission Internationale de I'Eclairage (CIE) was used. The coordinates in the CIE Lab evaluation system are: L* luminosity (<NUM> for black, <NUM> for white), a* chromaticity on a green (negative values) to red/ magenta (positive values) axis, and b* chromaticity on a blue (negative values) to yellow (positive values) axis. Colour parameters were measured without pre-treatment such as sample extraction or centrifugation. Before analysis, the instrument was calibrated to a white ceramic tile. Standard plastic cuvettes with <NUM> optical path length were filled with ca. <NUM> of the specimens. The cuvettes were covered with cardboard on three sides, excluding the side facing the measuring head of the chromameter, to provide a black background. Measurements were performed in a room without external light sources. Colour coordinates were recorded four times for each sample to ensure accurate measurement repeatability, and the mean values were calculated. The measurement of the background was performed under identical conditions with the use of a blank cuvette and black background cardboard. Since combined coordinates provide better indication of colour than individual ones, results in colour changes are expressed as ΔE as calculated using the CIE L*a*b* colour space-based calculations according to <MAT>.

The higher the delta E (ΔE), the higher the colour difference between the analysed sample (treated) and the reference sample (untreated). Delta E (ΔE) above <NUM> means visible changes (including to the untrained eye); ΔE lower than <NUM> can be visibly detectable by the trained eye.

Total difference of colour (ΔE) was measured in samples prepared as described above in <NUM>. The samples were processed by methods according to the invention (using a pressure of <NUM> MPa and <NUM> MPa, each for <NUM> and <NUM>, at <NUM>) and same recipe of baby food fruits samples processed by standard thermal technique (pasteurization: <NUM>, <NUM> minutes). The total difference of colour was assessed in samples immediately after the treatment, as well as during storage both at ambient and at refrigerated temperature (<NUM> to <NUM>) for <NUM>, <NUM>, <NUM>, <NUM> and <NUM> days.

Significant differences in total colour (i.e., all ΔE><NUM>) were observed between all samples processed by the method according to the invention as compared to the pasteurized samples immediately after processing and throughout storage up to <NUM> days (when analysis stopped) at ambient temperature. At refrigerated storage, ΔE ><NUM> was observed up to <NUM> days and still exhibited visible changes (ΔE > <NUM>) at day <NUM> of storage.

All tested sensorial attributes for smell, taste, appearance, and consistency (data not shown) and colour were improved over the fruit purees obtained by conventional thermal treatment. The acceptance of the fruit purees by the consumer is considered to be improved for all tested attributes, particularly for the colour measurement.

Orange juice from three different brands were obtained from a local convenience store and tested for reduction of Alicyclobacillus acidoterrestris spores according to the procedure mentioned in Example <NUM> -<NUM>.

The soluble solids (°Bx) were measured by standard technique, using a refractometer (MS REF <NUM> My-Soft). The water activity (Aw) of the commercial orange juices was measured using AQUALAB 3TE water activity meter (Decagon Device Inc. Aw of the samples were about <NUM>.

Determination of inactivated spores was performed using the BAT-agar spread plate method as described in Ex.

Whereas the method of the invention achieved a log reduction of <NUM> to <NUM> for fruit puree samples when tested for surviving spores immediately after the processing was executed (see Ex. <NUM>), under these conditions a mere <NUM> to <NUM> log reduction was achieved on orange juice samples. That is, this example confirms the results of <NPL>) wherein a maximum log reduction in the amount of germinating heat-resistant Alicyclobacillus acidoterrestris spores of up to about <NUM> logs was achieved. This value is considered to be inadequate and unsafe for the present purpose which is to obtain fruit-based compositions that are intended to be stored at ambient conditions and intended to be ingested by infants or toddlers. It was surprisingly found that similar processing settings applied on a different food matrix provided improved and satisfactory results.

Analysis of variance and Duncan's multiple-range test using StatSoft® Statistica <NUM> were used to test the significance of the differences (p < <NUM>) between the mean log values of the survival cells after treatment. Microsoft Office Excel <NUM> was used for linear and exponential regression of data plotted on survival curves and calculating the coefficient of determination (R2).

Claim 1:
A method for preparing a fruit-containing product containing less than <NUM><NUM> heat-resistant spores per gram of the fruit-containing product, wherein the method comprises the steps of:
a. providing a fruit-based composition comprising at least <NUM> wt.%, on a total solids basis, of a fruit material, wherein the composition has a pH between <NUM> and <NUM> and a total solids content between <NUM> and <NUM> wt.% by weight of the product, at ambient condition; and
b. subjecting the fruit-based composition to a pressure between <NUM> MPa and <NUM> MPa and a temperature between <NUM> and <NUM>, for <NUM> to <NUM> minutes,
and wherein the fruit-containing product is selected from baby foods, fruit purees, fruit spreads, slurries, mousse/mousselines, compotes, creams and jams.