WO2024115523A1

WO2024115523A1 - Process for preparing a frozen or freeze-dried solid additive composition

Info

Publication number: WO2024115523A1
Application number: PCT/EP2023/083437
Authority: WO
Inventors: Bernadette Theresia MOREL; Pieter Vonk
Original assignee: Dsm Ip Assets B.V.
Priority date: 2022-11-29
Filing date: 2023-11-28
Publication date: 2024-06-06

Abstract

The invention provides a process for preparing a frozen or freeze-dried solid additive composition, comprising or consisting of the following steps: a) preparing an aqueous mixture comprising or consisting of (i) formic acid or a salt or ester thereof and (ii) a crystallization inhibitor; b) preparing a frozen solid additive composition by freezing the aqueous mixture of a); and c) optionally preparing a freeze-dried solid additive composition by freeze-drying the frozen solid additive composition of b). In addition the invention provides a solid additive composition so prepared.

Description

PROCESS FOR PREPARING A FROZEN OR FREEZE-DRIED SOLID ADDITIVE COMPOSITION

Field of the invention

[001] The invention relates to a process for preparing a frozen or freeze-dried solid additive composition, a solid additive composition obtained or obtainable therewith, a kit of parts comprising such solid additive composition, a process of producing a cultured food product using such solid additive composition or such kit of parts, and the use of such solid additive composition or such kit of parts in a process for making a food or feed product.

Background of the invention

[002] Lactic acid bacteria are used world-wide in the dairy industry to produce a variety of fermented dairy products such as cheese, yoghurts, sour cream, kefir, butter and koumiss. Selected strains of lactic acid bacteria initiating and carrying out the desired fermentations are essential in the manufacture of the above products. They are often referred to as starter cultures.

[003] Milk can be inoculated without intermediate transfer and/or propagation with so-called direct vat set (DVS) or direct-to-vat inoculation (DVI) starter cultures.

[004] Such starter cultures are generally available from commercial manufacturers in frozen or lyophilized/freeze-dried form. They can comprise only a single lactic acid bacterium species, but can also be mixed cultures comprising two or more different lactic acid bacterium species.

[005] In order to maintain high cell counts and/or improved stability of liquid, frozen or lyophilized/freeze- dried starter cultures, cryoprotectants can be employed.

[006] W02009/056979 describes a starter composition made by adding appropriate amounts of powdered forms of the sources of formate and purine to a lyophilized (i.e., freeze dried) cell culture. It also describes a starter composition made by blending a frozen bacterial pellet with a frozen solution containing the sources of formate and purine.

[007] US 2005/0069862 describes that measures that have to be provided by an effective cryo protective agent are very much different depending on whether the cryoprotective agent is designed to protect liquid or frozen cultures, and consequently, additives, which are effective cryoprotective agents of liquid cultures, may not be effective for frozen cultures. US 2005/0069862 states that one example of such an additive is Na-formate. US 2005/0069862 refers to WO 00/39281 and mentions that 3% Na-formate is effective to increase the storage stability of liquid lactic acid bacterial starter culture concentrates. However, as illustrated in Example 15 of US 2005/0069862, 3% Na-formate decrease the storage stability of frozen lactic acid bacterial starter culture.

[008] WO2012/076665 describes a process for making a starter culture composition comprising a microorganism, a cryoprotectant and at least one stimulating additive, the process comprising the steps of: a) culturing a microorganism in a culture medium; and b) collecting the microorganism from the culture medium; and c) adding a cryoprotectant to the microorganism obtained in step b); and d) freezing the microorganisms obtained in step c); and e) adding at least one stimulating additive to microorganisms obtained in step d). In the examples combinations of frozen culture pellets and frozen sodium formate pellets are described and also combinations of freeze-dried culture pellets and freeze-dried sodium formate pellets are described. In some examples a cryoprotectant solution containing (in wt%): maltodextrin (28,6%), sorbitol (10,3%), calcium ascorbate (11 ,4%), glutamate (2,9%) and water (46,8%) is added to 65.84 g of the culture concentrate directly after centrifugation and before freezing or freeze-drying to create the culture pellets. In other examples the sodium formate pellets contain 40% (w/w) sodium formate, 2% (w/w) Arabic gum with the remainder being water.

[009] Although the above process of WO2012/076665 has been proven to be advantageous, further improvement is still desired.

Summary of the invention

[010] Inventors have noted that when applying freeze-dried pellets, granules, tablets or a powder comprising formic acid or a salt or ester thereof as described in WO2012/076665, the water activity (Aw) inside these pellets, granules, tablets or powder may increase during storage. Such an increase in water activity (Aw) is an indication of physical instability and undesired. Without wishing to be bound by any kind of theory it is believed that this increase in water activity (Aw) may have a negative effect on any freeze- dried pellets, granules, tablets or powder comprising cultures that are to be stimulated by the formic acid or the salt or ester thereof.

[011] After extensive study, it has now surprisingly been found that the above increase in water activity (Aw) can be significantly decreased by using (at least partly) amorphous formic acid or the salt or ester thereof and mixing such with a component that stabilizes this amorphous state. Such a component that stabilizes the amorphous state is herein also referred to as a crystallization inhibitor.

[012] The invention accordingly provides a process for preparing a frozen or freeze-dried solid additive composition, comprising or consisting of the following steps: a) preparing an aqueous mixture comprising or consisting of (I) formic acid or a salt or ester thereof and (ii) a crystallization inhibitor; b) preparing a frozen solid additive composition by freezing the aqueous mixture of a); and c) optionally preparing a freeze-dried solid additive composition by freeze-drying the frozen solid additive composition of b).

[013] The above process can preferably produce the formic acid or a salt or ester thereof in the frozen or freeze-dried solid additive composition existing partly or wholly in an amorphous state.

[014] Without wishing to be bound by any kind of theory, it is believed that the prepared frozen or freeze- dried solid additive composition can be helpful to prepare a stable starter culture as illustrated in detail below. In addition it has surprisingly been found that the process according to the invention can allow for a faster freeze-drying process, which is economically attractive. [015] Accordingly, the invention further provides a solid additive composition, preferably frozen or freeze- dried, preferably in the form of a pellet, a granule, a tablet, a powder or another solid particle, comprising or consisting of:

- formic acid or a salt or ester thereof, wherein this formic acid or salt or ester thereof is partly or wholly present in an amorphous state; and

- a crystallization inhibitor, which solid additive composition preferably does not comprise any type of lactic acid bacteria, present, individually, in a viable cell count density of equal to or more than 1 x 10⁶ cfu/gram; and which solid additive composition preferably is obtained or obtainable by a process as described above.

[016] The invention also provides a kit of parts comprising:

- a first solid composition comprising or consisting of the solid additive composition as described above; and

- a second solid composition comprising or consisting of a solid culture composition, preferably frozen or freeze-dried, preferably in the form of a pellet, a granule, a tablet, a powder or another solid particle, wherein the solid culture composition comprises at least one type of lactic acid bacteria in an individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram.

[017] Preferably the above kit of parts is a starter culture. As illustrated in the examples, such starter culture may advantageously have an improved stability.

[018] In addition, the invention provides a method for producing a cultured food product, the method comprising the steps of:

- inoculating a precursor material, preferably a milk base, with a kit of parts as described above;

- culturing the precursor material with the kit of parts, and

- producing the cultured food product.

[019] Preferably the cultured food product is a fermented milk product.

[020] Lastly, the invention provides a use of the solid additive composition described above or the kit of parts described above in a process for making a food or feed product. This food or feed product is preferably a fermented milk product.

Brief description of the drawings

[021] The invention is illustrated by figures 1 and 2.

[022] Figure 1 illustrates a comparison of freeze drying (I) pellets with a composition, based on dry matter, of 100% w/w sodium-formate (“100% NAF”); (II) pellets with a composition, based on dry matter, of 98 % w/w sodium formate and 2 % w/w maltodextrin (“NAF-MD (2%)”); and (ill) pellets with a composition of 96 % w/w sodium formate and 4 % w/w maltodextrin (“ NAF-MD (4%)”). It is illustrated that the NAF-MD (2%) composition and the NAF-MD (4%) composition allow for a faster freeze-drying process, which is economically attractive. The NAF-MD (4%) composition allows for the fastest freeze-drying. [023] Figure 2 illustrates a comparison of freeze drying (I) pellets with a composition, based on dry matter, of 100% w/w sodium-formate (“NaF 100%”, the line at the top); (II) pellets with a composition, based on dry matter, of 98 % w/w sodium formate and 2 % w/w sucrose (“NaF 98% + Sucrose 2%”, the second line from the top); (ill) pellets with a composition, based on dry matter, of 98 % w/w sodium formate and 2 % w/w maltodextrin (“NaF 98% + Maltodextrin 2%”, the third line from the top); and (iv) pellets with a composition, based on dry matter, of 80 % w/w sodium formate and 20 % w/w maltodextrin (“ NaF 80% + Maltodextrin 20%”, the bottom dots). It illustrates the increase in water activity (Aw) during storage at 30°C forthe various pellets and shows that the negative effect of this increase in water activity (Aw) can be reduced by using the crystallization inhibitor maltodextrin or sucrose. It is shown that already after 8 days of storage, the water activity (Aw) was lower when use was made of pellets with compositions “NaF 98% + Sucrose 2%”, “NaF 98% + Maltodextrin 2%” and “NaF 80% + Maltodextrin 20%”. The effect continues to exist, even after 40 hours.

Detailed description of the invention

Definitions

[024] Unless defined otherwise or clearly indicated by context, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art.

[025] Throughout the present specification and the accompanying claims, the words "comprise" and "include" and variations such as "comprises", "comprising", "includes" and "including" are to be interpreted inclusively. That is, these words are intended to convey the possible inclusion of other elements or integers not specifically recited, where the context allows.

[026] The articles “a” and “an” are used herein to refer to one or to more than one (i.e. to one or at least one) of the grammatical object of the article. By way of example, “an element” may mean one element or more than one element. When referring to a noun (e.g. a compound, an additive, etc.) in the singular, the plural is meant to be included. Thus, when referring to a specific moiety, e.g. a "strain", this means "at least one" of that strain, e.g. "at least one strain", unless specified otherwise.

[027] When referring to a compound of which several isomers exist (e.g. a D and an L enantiomer), the compound in principle includes all enantiomers, diastereomers and cis/trans isomers of that compound that may be used in the particular aspect of the invention; in particular when referring to such as compound, it includes the natural isomer(s).

[028] Unless explicitly indicated otherwise, the various embodiments of the invention described herein can be cross-combined.

[029] The term "milk" is intended to encompass milks from mammals and plant sources or mixtures thereof. Preferably, the milk is from a mammal source. Mammals sources of milk include, but are not limited to cow, sheep, goat, buffalo, camel, llama, horse or reindeer. In an embodiment, the milk is from a mammal selected from the group consisting of cow, sheep, goat, buffalo, camel, llama, horse and deer, and combinations thereof. Plant sources of milk include, but are not limited to, milk extracted from soy bean, pea, peanut, barley, rice, oat, quinoa, almond, cashew, coconut, hazelnut, hemp, sesame seed and sunflower seed. Bovine milk is preferred. In addition, the term "milk" refers to not only whole milk, but also skim milk or any liquid component derived thereof or reconstituted milk.

[030] The term "milk base" refers to a base composition, comprising milk or milk ingredients, or derived from milk or milk ingredients. As indicated above, the milk can be from an animal, preferably mammal, source, from a plant source, or a mixture thereof. The milk base can be used as a raw material for the fermentation to produce a fermented milk product. The milk base may for example comprise or consist of skimmed or non-skimmed milk, or reconstituted milk. Optionally the milk base may be concentrated or in the form of powder, or may be reconstituted from such. By reconstituted milk is herein understood liquid milk obtained by adding liquid, such as water, to a skim milk powder, skim milk concentrate, whole milk powder or whole milk concentrate. Furthermore, the milk base may or may not have been subjected to a thermal processing operation which is at least as efficient as pasteurization. Preferably the milk base is from a bovine source.

[031] As used in this specification, the terms "fermented milk product", "fermented dairy product" and "acidified milk product" are used interchangeably and are intended to refer to products which are obtained by the multiplication of lactic acid bacteria in a milk base leading to a milk coagulum. The particular characteristics of the various fermented milk products depend upon various factors, such as the composition of milk base, the incubation temperature, the composition of the lactic acid bacteria and/or presence of further non-lactic acid microorganisms. Thus, fermented milk products manufactured herein include, for instance, various types of yoghurt (including for example set yoghurt, low fat yoghurt, non-fat yoghurt), kefir, dahi, ymer, buttermilk, butterfat, sour cream and sour whipped cream as well as fresh cheeses such as quark and cottage cheese. Petit Suisse or Mozarella is yet another example of a fermented dairy product. Preferably the fermented milk product is a yoghurt.

[032] Two basic types of yoghurt exist, according to their physical state in the retail container: set yoghurt and stirred yoghurt. Set yoghurt is fermented after being packed in a retail container, and stirred yoghurt is almost fully fermented in a fermentation tank before it is packed, the yoghurt gel being broken up during the stirring. The fermented milk product produced in the current invention can be a stirred yoghurt or a set yoghurt. Preferably the fermented milk product is a set yoghurt.

[033] The terms "yoghurt" and "yogurt" are used interchangeably herein. The term "yoghurt" refers to products comprising or obtained by means of lactic acid bacteria that include at least Streptococcus salivarius thermophilus and Lactobacillus delbruekii subsp. bulgaricus, but may also, optionally, include further microorganisms such as Lactobacillus delbruekii subsp. lactis, Bifidobacterium animalis subsp. lactis, Lactococcus lactis, Lactobacillus acidophilus and Lactobacillus casei, or any microorganism derived therefrom. Such lactic acid strains other than Streptococcus salivarius thermophilus and Lactobacillus delbruekii subsp. bulgaricus, can give the finished product various properties, such as the property of promoting the equilibrium of the gut microbiota. As used herein, the term "yoghurt" encompasses set yoghurt, stirred yoghurt, drinking yoghurt, heat treated yoghurt and yoghurt-like products. More preferably, the term "yoghurt" encompasses, but is not limited to, yoghurt as defined according to French and European regulations, e.g. coagulated dairy products obtained by lactic acid fermentation by means of specific thermophilic lactic acid bacteria only (i.e. Lactobacillus delbruekii subsp. bulgaricus and Streptococcus salivarius thermophilus) which are cultured simultaneously and are found to be live in the final product in an amount of at least 10 million CFU (colony-forming unit) / g. Preferably, the yoghurt is not heat-treated after fermentation. Yoghurts may optionally contain added dairy raw materials (e.g. cream) or other ingredients such as sugar or sweetening agents, one or more flavouring(s), fruit, cereals, or nutritional substances, especially vitamins, minerals and fibers. Such yoghurt advantageously meets the specifications for fermented milks and yoghurts of the AFNOR NF 04-600 standard and/orthe codex StanA- lla-1975 standard. In order to satisfy the AFNOR NF 04-600 standard, the product must not have been heated after fermentation and the dairy raw materials must represent a minimum of 70% (m/m) of the finished product.

[034] In the present context, the terms "fresh cheese", "unripened cheese", "curd cheese" and "curdstyle cheese" are used interchangeably herein to refer to any kind of cheese such as natural cheese, cheese analogues and processed cheese in which the protein/ casein ratio does not exceed that of milk. [035] The term "starter" or "starter culture" as used herein refers to a culture of one or more food-grade micro-organisms, more preferably a culture comprising lactic acid bacteria, which are responsible for the acidification of the milk base. Starter cultures may be fresh (liquid), frozen or freeze-dried. Freeze dried cultures need to be regenerated before use. For the production of a yoghurt, the starter culture (i.e. the total weight of all lactic acid bacterial combined) can for example be added in an amount from 0.001 to 10% by weight, suitably in an amount of 0.01 to 3% by weight, of the total amount of milk base. For the production of cheese, dosages in the lower part of the range can be used such as from 0.006% by weight of the total amount of milk base.

[036] As used herein, the term "lactic acid bacteria", "LAB", "lactic acid bacterial strains" and "lactic bacteria" are used interchangeably and refer to food-grade bacteria producing lactic acid as the major metabolic end-product of carbohydrate fermentation. These bacteria are related by their common metabolic and physiological characteristics and are usually Gram positive, low-GC, acid tolerant, non- sporulating, non-respiring, rod-shaped bacilli or cocci. During the fermentation stage, the consumption of lactose by these bacteria causes the formation of lactic acid, reducing the pH and leading to the formation of a protein coagulum. These bacteria are thus responsible for the acidification of milk and for the texture of the dairy product. As used herein, the term "lactic acid bacteria" or "lactic bacteria" encompasses, but is not limited to, bacteria belonging to the genus of Lactobacillus spp., Bifidobacterium spp., Streptococcus spp., Lactococcus spp., such as Lactobacillus delbruekii subsp. bulgaricus, Streptococcus salivarius thermophilus, Lactobacillus lactis, Bifidobacterium animalis, Lactococcus lactis, Lactobacillus casei, Lactobacillus plantarum, Lactobacillus helveticus, Lactobacillus acidophilus and Bifidobacterium breve.

[037] A strain is a genetic variant or subtype of a microorganism, in this case a subtype or variant of a lactic acid bacteria. [038] By a “crystallization inhibitor” is herein preferably understood a compound that can stabilize an amorphous state, preferably the amorphous state of formic acid or a salt or ester thereof. Such a crystallization inhibitor can suitably prevent crystallization or re-crystallization. Most preferably the crystallization inhibitor is maltodextrin.

The process for preparing a frozen or freeze-dried solid additive composition

[039] As indicated above the invention provides a process for preparing a frozen or freeze-dried solid additive composition, comprising or consisting of the following steps: a) preparing an aqueous mixture comprising or consisting of (i) formic acid or a salt or ester thereof and

(II) a crystallization inhibitor; b) preparing a frozen solid additive composition by freezing the aqueous mixture of a); and c) optionally preparing a freeze-dried solid additive composition by freeze-drying the frozen solid additive composition of b).

[040] Preferably the aqueous mixture of step a) does not comprise any type of lactic acid bacteria, individually, in any individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram. More preferably the aqueous mixture of step a) does not comprise any type of lactic acid bacteria, individually, in any individual viable cell count density of equal to or more than 1 x 10⁴ cfu/gram, before the freezing and/or freeze-drying of the mixture.

[041] More preferably the aqueous mixture of step a) comprises a total amount of lactic acid bacteria of equal to or less than 1 x 10¹⁰ cfu/gram, more preferably of equal to or less than 1 x 10⁸ cfu/gram, still more preferably of equal to or less than 1 x 10⁶ cfu/gram, even more preferably of equal to or less than 1 x 10⁵ cfu/gram, yet more preferably of equal to or less than 1 x 10⁴ cfu/gram, and yet even more preferably of equal to or less than 1 x 10³ cfu/gram. Most preferably the aqueous mixture of step a) comprises a total amount of lactic acid bacteria of equal to or less than 1 x 10² cfu/gram, preferably equal to or less than 1 x 10¹ cfu/gram. Ideally the aqueous mixture of step a) comprises no lactic acid bacteria.

[042] As explained below, preferably the frozen or freeze-dried solid additive composition does not comprise any type of lactic acid bacteria in any individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram, preferably of equal to or more than 1 x 10⁴ cfu/gram.

[043] As further explained below, preferably the formic acid or a salt or ester thereof in the frozen or freeze-dried solid additive composition exists partly or wholly in an amorphous state.

[044] Preferably the aqueous mixture of step a) comprises or consists of:

- in the range from equal to or more than 10 % w/w to equal to or less than 60 % w/w, more preferably equal to or more than 20 % w/w to equal to or less than 50 % w/w, and most preferably equal to or more than 30 % w/w to equal to or less than 45 % w/w of formic acid or a salt or ester thereof; and

- in the range from equal to or more than 0.05 % w/w to equal to or less than 20 % w/w, more preferably in the range from equal to or more than 0.5 % w/w to equal to or less than 10 % w/w, and most preferably in the range from equal to or more than 1 % w/w to equal to or less than 7 % w/w of crystallization inhibitor, wherein the balance in the aqueous mixture preferably consists of water.

[045] The freezing in step b) can be carried out in any manner known by the person skilled in the art to be suitable therefore. Preferably the frozen solid additive composition is prepared by freezing the aqueous mixture to a temperature below -1 °C, more preferably to a temperature equal to or less than -10°C, even more preferably to a temperature equal to or less than -20°C. Still more preferably the frozen solid additive composition is prepared by freezing the aqueous mixture to a temperature equal to or less than -50°C, even more preferably to a temperature equal to or less than -80°C. Yet more preferably the frozen solid additive composition is prepared by freezing the aqueous mixture to a temperature equal to or less than -100°C, even more preferably to a temperature equal to or less than -180°C. Most preferably the freezing is carried out with the help of liquid nitrogen (also referred to as cryogenic liquid nitrogen). More preferably step b) comprises dripping or spraying of the aqueous mixture into liquid nitrogen. If so desired the frozen solid additive prepared in step b) can subsequently be milled. That is, step b) may suitably further comprise milling of the frozen solid additive composition.

[046] Step b) may comprise preparing a frozen solid additive composition by freezing the aqueous mixture of a), wherein the frozen solid additive composition may have the form of a pellet, a granule, a tablet, a powder or another solid particle. More preferably step b) comprises preparing a frozen solid additive composition by freezing the aqueous mixture of a), into the form of a pellet or a granule.

[047] If freeze-dried, the frozen solid additive composition is preferably freeze-dried until an Aw of equal to or less than 0.2, preferably an Aw of equal to or less than 0.1 is reached.

[048] Preferably the process comprises step c), comprising or consisting of preparing a freeze-dried solid additive composition by freeze-drying the frozen solid additive composition of b).

[049] If freeze-dried, the frozen solid additive composition is preferably freeze-dried until an Aw of equal to or less than 0.2, preferably an Aw of equal to or less than 0.1 is reached. That is, more preferably the process is a process wherein the mixture is freeze-dried until an Aw of less than 0.2, preferably an Aw of less than 0.1 , is reached.

[050] If freeze-dried, such freeze drying can comprise one or more steps and the freeze-drying pressure and temperature can vary. Freeze-drying may further be carried out by for example plate freeze-drying or radiation freeze-drying. The pressure during freeze-drying preferably lies in the range from equal to or more than 0.10 mbar, more preferably equal to or more than 0.20 mbar to equal to or less than 3.0 mbar, more preferably equal to or less than 2.0 mbar. The temperature during the freeze-drying preferably lies in the range from equal to or more than 5°C, more preferably equal to or more than 10°C and still more preferably equal to or more than 20°C to equal to or less than 90°C, more preferably equal to or less than 80°C and most preferably equal to or less than 70°C. For example, after freezing by dripping in cryogenic liquid nitrogen, the frozen particles can be submitted to plate freeze-drying at 0.50 mbar during primary drying and 0.18 mbar during secondary drying while applying a shelf heating temperature up to 50°C. [051] In the process according to the invention the time needed for freeze-drying can advantageously be reduced. The drying process preferably takes in the range from equal to or more than 25 to equal to or less than 60 hours. In comparison with conventional processes, drying time can be reduced by using the process according to the invention.

[052] Advantageously the invention also provides a preferred improved freeze-drying process comprising radiation freeze-drying using a pressure in the range from 0.35 mbar to 0.45 mbar, preferably 0.40 mbar, during primary drying and a pressure in the range from 0.05 mbar to 0.15 mbar, preferably 0.10 mbar, during secondary drying, in combination with a shelf heating temperature up to 50°C, preferably in the range from 5°C to 50°C.

[053] Preferably the frozen and/or freeze-dried solid additive composition is produced in the form of solid particles, preferably in the form of pellets or granules, preferably having a particle size distribution with a mean particle size in the range from preferably equal to or more than 0.5 mm, more preferably equal to or more than 1 mm, still more preferably equal to or more than 2 mm, and most preferably equal to or more than 3 mm to preferably equal to or less than 15 mm, more preferably equal to or less than 10 mm, still more preferably equal to or less than 8 mm and most preferably equal to or less than 6 mm. Most preferably the pellets or granules have a particle size distribution wherein the mean particle size ranges from 1 mm to 8 mm, preferably from 2 to 6 mm. Particle size distributions and their mean particle size can be determined by sieving (such as for example described in ASTM D6913-04(2009)), size selective particle filtering and/or by particle size analyzers (PSAs) based on laser diffraction (LD).

The crystallization inhibitor

[054] Preferably the crystallization inhibitor is chosen from the group consisting of mono-saccharides, disaccharides and polysaccharides. Examples of possible crystallization inhibitors include glucose, fructose, galactose, mannose, maltose, sucrose, lactose, starch, cyclodextrins and maltodextrin. More preferably the crystallization inhibitor is sucrose or maltodextrin.

[055] More preferably the crystallization inhibitor comprises or consists of sucrose, carboxymethyl cellulose, maltodextrin or a combination of two or more of these.

[056] In one embodiment the crystallization inhibitor preferably does not comprise or consist of maltodextrin.

[057] In an alternative preferred embodiment the crystallization inhibitor comprises or consists of maltodextrin.

[058] In one embodiment the crystallization inhibitor preferably does not comprise or consist of sucrose. [059] In an alternative preferred embodiment the crystallization inhibitor comprises or consists of sucrose. [060] The crystallization inhibitor is preferably present in an amount in the range from equal to or more than 0.05 % w/w to equal to or less than 40 % w/w, more preferably in the range from equal to or more than 0.5 % w/w to equal to or less than 20 % w/w, and most preferably in the range from equal to or more than 1% w/w to equal to or less than 14 % w/w of crystallization inhibitor, based on the total weight of the crystallization inhibitor and the formic acid or salt or ester thereof together.

Other components

[061] Preferably the solid additive composition does not comprise purine or inosine.

[062] Preferably the solid additive composition does not comprise yeast extract.

[063] Preferably the solid additive does not comprise Arabic gum or any other type of gum.

[064] In addition to, or as an alternative for, the crystallization inhibitor, the pellets further may or may not comprise hydrocolloid molecules such as for example sodium alginate, Cellulose gum, Gellan gum, Guar gum, Locust bean gum, Pectin, Xanthan gum. In one embodiment the pellets do not comprise hydrocolloid molecules such as for example sodium alginate, Cellulose gum, Gellan gum, Guar gum, Locust bean gum, Pectin, Xanthan gum. In an alternative embodiment the pellets do comprise hydrocolloid molecules such as for example sodium alginate, Cellulose gum, Gellan gum, Guar gum, Locust bean gum, Pectin, Xanthan gum.

The formic acid or a salt or ester thereof

[065] In this invention the additive is the formic acid or a salt or ester thereof. The formic acid or a salt or ester thereof can be present as the formic acid itself. Preferably the formic acid or a salt or ester thereof is present is a salt. Preferred examples of a formic acid or a salt or ester thereof include sodium formate, potassium formate, ammonium formate, calcium formate, ethyl formate and methyl formate. More preferably the formic acid or a salt or ester thereof is a formate salt. Most preferably the formic acid or a salt or ester thereof is present in the form of ammonium formate, potassium formate or sodium formate. Sodium formate is especially preferred.

[066] The formic acid or salt or ester thereof is preferably at least partly present in an amorphous state. Preferably at least 10% w/w, more preferably at least 20% w/w, even more preferably at least 30% w/w, still more preferably at least 40% w/w and even still more preferably at least 50% w/w of the formic acid or salt or ester thereof is present in an amorphous state. More preferably at least 60% w/w, even more preferably at least 70% w/w, still even more preferably at least 80% w/w, and yet more preferably at least 90% w/w of the formic acid or salt or ester thereof is present in an amorphous state. Most preferably at least 95% w/w of the formic acid or salt or ester thereof is present in an amorphous state. Ideally at least 99% w/w of the formic acid or salt or ester thereof is present in an amorphous state.

[067] Amorphous content (amorphicity) in materials can be determined for example by Differential Scanning Calorimetry (DSC), Modulated DSC® (MDSC®), Continuous Relative Humidity Perfusion Microcalorimetry (cRHp), Dynamic Vapor Sorption (DVS), and Solution Calorimetry (SolCal). For the aspects as described here, a determination of amorphous content by Differential Scanning Calorimetry (DSC) is preferred. The solid additive composition

[068] By an additive composition is herein understood a composition comprising at least one compound that can suitably function as an additive in a starter culture comprising one or more selected strains of lactic acid bacteria. In this invention the additive is the formic acid or a salt or ester thereof. Formic acid or a salt thereof can be suitable as an additive for starter cultures, as described for example in WO2012/076665.

[069] By a solid additive composition is herein understood a composition in a solid form, in contrast to a composition in a liquid form. For the purposes of this invention, a frozen form is herewith also understood to be a solid form. Preferably the solid additive composition is a solid additive composition in a frozen or freeze-dried form. Most preferably the solid additive composition is a solid additive composition in a freeze- dried form.

[070] Preferably the, preferably frozen or freeze-dried, solid additive composition comprises a water activity (Aw) of less than 0.2, more preferably an Aw of less than 0.1 .

[071] Preferably the solid additive composition does not comprise any type of lactic acid bacteria present, individually, in a viable cell count density of equal to or more than 1 x 10⁶ cfu/gram. More preferably the solid additive composition does not comprise any micro-organisms in a viable cell count density of equal to or more than 1 x 10⁶ cfu/gram.

[072] More preferably the solid additive composition is a lactic acid bacteria (LAB) depleted or LAB free composition, comprising, of each potentially present type of LAB individually, a viable cell count density of equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram. Without wishing to be bound by any kind of theory, it is believed that at a presence of equal to or less than 1 x 10⁶ cfu/gram or less, and certainly at a presence of equal to or less than 1 x 10³ cfu/gram, any lactic acid bacteria (LAB) will have very limited activity or even be inactive in any process for preparing a cultured food product.

[073] Even more preferably the solid additive composition is a microorganism depleted or microorganism free composition, comprising, of each potentially present type of microorganism individually, a viable cell count density of equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram. Without wishing to be bound by any kind of theory, it is believed that at a presence of equal to or less than 1 x 10⁶ cfu/gram or less, and certainly at a presence of equal to or less than 1 x 10³ cfu/gram, any microorganism will have very limited activity or even be inactive in any process for preparing a cultured food product.

[074] For example, the solid additive composition preferably comprises less than 1 ,10⁶ cfu/g, preferably less than 10³ cfu/g, of each of Streptococcus spp, Leuconostoc spp, Lactococcus spp and/or Lactobacillus Leuconostoc spp. [075] Thus, the invention also provides a solid additive composition, in the form of a pellet, a granule, a tablet or a powder, comprising a mixture of:

- formic acid or a salt or ester thereof at least partly present in an amorphous state; and

- a crystallization inhibitor, which composition comprises of any type of LAB present, individually, a viable cell count density of equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram.

[076] Even more preferably the solid additive composition is a composition comprising a total amount of lactic acid bacteria (LAB) that is equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram.

[077] Still even more preferably the solid additive composition is a composition comprising a total amount of microorganisms that is equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram.

[078] Herein, the expression cfu/g means "colony forming units per gram". This is also referred to as the viable cell count density. The determination of viable cell count density of the lactic acid bacterial strains described or claimed herein is known to the skilled person. The viable cell count density (expressed in cfu/g or in cfu/ml) of Streptococcus thermophilus is preferably determined using Streptococcus Thermophilus Isolation Agar or STA. This agar has the following composition: Casein enzymic hydrolysate 10.0 g/l; Yeast extract 5.0 g/l; Sucrose 10.0 g/l; Dipotassium phosphate 2.0 g/l; Agar 15.0 g/l; Final pH 6.8 +/- 0.2 at 25°C. Cultural characteristics after anaerobic incubation for 48-72 hours at 35-37°C. STA is conveniently obtained as Fluka cat.# 17257 Streptococcus Thermophilus Isolation Agar from Sigma-Aldrich Chemie GmbH, Industriestrasse 25, Postfach CH-9471 Buchs, Switzerland. In a mixed culture or in a frozen pellet comprising a mixture of Streptococcus thermophilus and lactobacilli, streptococci and lactobacilli can be together enumerated on the same STA plate; streptococci are counted as smooth, round colonies and lactobacilli are counted as fluffy colonies. In a single strain culture or in a frozen pellet comprising a single strain, the viable cell count density (expressed in cfu/g or in cfu/ml) of lactobacilli, especially of Lactobacillus helveticus is preferably determined using tryptone glucose meat extract agar for example as detailed by Galesloot, T. E., F. Hassing, and J. Stadhouders. 1961. Neth. Milk Dairy J. 15:127-150. Cultural characteristics are conveniently identifiable after incubation for 2-3 days at 37°C. [079] The solid additive composition is suitably present in the form of a pellet, a granule, a tablet, a powder or another solid particle. More preferably the solid additive composition is present as a pellet or a granule.

[080] The pellets are preferably pellets manufactured by:

- dripping or spraying an aqueous solution or slurry comprising the additive and the crystallization inhibitor into a cryogenic liquid, preferably nitrogen liquid, to produce frozen droplets; and

- freeze drying the frozen droplets to produce solid additive composition pellets or solid additive composition granules.

[081] Preferably the pellets or granules have a particle size distribution wherein the mean particle size ranges from preferably equal to or more than 0.5 mm, more preferably equal to or more than 1 mm, still more preferably equal to or more than 2 mm, and most preferably equal to or more than 3 mm to preferably equal to or less than 15 mm, more preferably equal to or less than 10 mm, still more preferably equal to or less than 8 mm and most preferably equal to or less than 6 mm. Most preferably the pellets or granules have a particle size distribution wherein the mean particle size ranges from 1 mm to 8 mm, preferably from 2 to 6 mm. Particle size distributions and their mean particle size can be determined by sieving (such as for example described in ASTM D6913-04(2009)), size selective particle filtering and/or by particle size analyzers (PSAs) based on laser diffraction.

[082] The, preferably freeze-dried, solid additive composition is preferably uncoated. That is, the, preferably freeze-dried, solid additive composition is preferably not coated. WO2012/076665 describes that in one embodiment of the therein claimed process, the freeze- dried microorganism and/or the stimulating additive added to the freeze-dried microorganism and/or the freeze-dried starter culture composition is coated. Without wishing to be bound by any kind of theory, it is believed that such a coating could disadvantageously reduce the working of the additive, i.e. the working of the formic acid or a salt or ester thereof. It is therefore preferred that the solid additive composition is uncoated. To create optimum contact, most preferably, neither the first solid composition nor the second solid composition is coated.

Starter culture composition

[083] As illustrated above, the present invention therefore also provides a kit of parts comprising:

I) a solid additive composition as described above; and

II) a solid culture composition, preferably in the form of a pellet, a granule, a tablet, a powder or another solid particle, wherein the solid culture composition comprises at least one type of lactic acid bacteria in an individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram.

[084] Preferably the kit of parts, or starter culture composition, is a kit of parts, respectively starter culture composition, wherein the solid additive composition and/or the solid culture composition is in a frozen or freeze-dried form.

[085] Preferably the kit of parts, or starter culture composition, is a kit of parts, respectively starter culture composition, wherein: - the solid additive composition is present in the form of solid additive composition pellets, granules or tablets; and

- the solid culture composition is present in the form of solid culture composition pellets, granules or tablets; and

- the solid additive composition pellets, granules or tablets are distinctive, different and/or separate from the solid culture composition pellets, granules or tablets.

That is, preferably the solid additive composition pellets, granules or tablets are not the same as the solid culture composition pellets, granules or tablets.

[086] Preferably the solid additive composition comprises of any type of LAB present, individually, a viable cell count density of equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram. More preferably the solid additive composition comprises a total amount of lactic acid bacteria (LAB) that is equal to or less than 1 x 10⁶ cfu/gram, more preferably equal to or less than 1 x 10⁵ cfu/gram, still more preferably equal to or less than 1 x 10⁴ cfu/gram, even more preferably equal to or less than 1 x 10³ cfu/gram, still even more preferably equal to or less than 1 x 10² cfu/gram and most preferably equal to or less than 1 x 10¹ cfu/gram.

[087] The solid culture composition preferably comprises a total amount of lactic acid bacteria (LAB) that is equal to or more than 1 x 10⁶ cfu/gram, more preferably equal to or more than 1 x 10⁵ cfu/gram, still more preferably equal to or more than 1 x 10⁷ cfu/gram, even more preferably equal to or more than 1 x 10⁸ cfu/gram, still even more preferably equal to or more than 1 x 10⁹ cfu/gram and most preferably equal to or more than 1 x 1O¹⁰ cfu/gram. More preferably the solid culture composition comprises of any type of LAB present, individually, a viable cell count density that is equal to or more than 1 x 10⁶ cfu/gram, more preferably equal to or more than 1 x 10⁵ cfu/gram, still more preferably equal to or more than 1 x 10⁷ cfu/gram, even more preferably equal to or more than 1 x 10⁸ cfu/gram, still even more preferably equal to or more than 1 x 10⁹ cfu/gram and most preferably equal to or more than 1 x 1O¹⁰ cfu/gram. Still more preferably the solid culture composition comprises more than 1 ,10⁶ cfu/g, preferably more than 10⁹ cfu/g, of one or more of Streptococcus spp, Leuconostoc spp, Lactococcus spp and/or Lactobacillus Leuconostoc spp., bifidobacteria, propioni bacteria, Enterococci. Most preferably the solid culture composition comprises more than 1.10⁶ cfu/g, preferably more than 10⁹ cfu/g, of one or more of bacteria belonging to the genus of Lactobacillus spp., Bifidobacterium spp., Streptococcus spp., Lactococcus spp., more preferably one or more of as Lactobacillus delbruekii subsp. bulgaricus, Streptococcus salivarius thermophilus, Lactobacillus lactis, Bifidobacterium animalis, Lactococcus lactis, Lactobacillus casei, Lactobacillus plantarum, Lactobacillus helveticus, Lactobacillus acidophilus and Bifidobacterium breve.

[088] The solid culture composition may in addition comprise one or more cryoprotectants. Suitable cryoprotectants include those described in WO2012/076665, herein incorporated by reference. However, the cryoprotectant(s) and the crystallization inhibitors have different functions. Hence, preferably, within one kit of parts or within one starter culture composition, any potential cryoprotectant(s) in the solid culture composition, and the crystallization inhibitor in the solid additive composition, are different compounds. That is, preferably any potential cryoprotectant(s) in the solid culture composition are not the same compound as the crystallization inhibitor in the solid additive composition.

[089] Preferably the solid culture composition and/or the solid additive composition is frozen or freeze- dried. Most preferably both the solid culture composition and the solid additive composition are present in the form of frozen or freeze-dried pellets, granules or tablets.

[090] The kit of parts can suitably be used as a starter culture composition.

[091] “Starter culture” is defined herein as a preparation containing microbial cells that is intended for inoculating a medium to be fermented. “Microorganism” as used herein includes, but is not limited to, algae, protozoa, viruses, bacteria (such as lactic acid bacteria, Brevibacterium and Propionibacterium) and fungi (such as the genera Penicillium, Geotrichum, Saccharomyces and Kluyveromyces). The microorganisms may be non-modified (i.e. wild type) or genetically modified. Genetically modified microorganisms may be provided with and without the use of recombinant DNA-technology.

[092] The invention advantageously also provides for the use of such a starter culture composition in a process for making a food or feed product.

Process of producing a cultured food product

[093] The invention further provides a process of producing a cultured food product, the process comprising the steps of: a) inoculating a precursor material with a starter culture composition prepared according to the process of the invention or a kit of parts as described above. b) culturing the precursor material with the starter culture composition or kit of parts, and c) producing the cultured food product.

[094] Preferably the cultured food product is a fermented milk product. As indicated above, milks from mammals and plant sources or mixtures thereof are preferred. More preferably, the milk is from a mammal source. Preferred fermented milk products include yoghurt (including for example set yoghurt, low fat yoghurt, non-fat yoghurt), kefir, dahi, ymer, buttermilk, butterfat, sour cream and sour whipped cream as well as fresh cheeses such as quark and cottage cheese. Petit Suisse or Mozarella is yet another example of a fermented milk product. Preferably the fermented milk product is a yoghurt.

In view of the above, the invention also provides a use of the solid additive composition as described above, or a kit of parts as described above, in a process for making a food or feed product.

Experiments

Example 1, production of freeze-dried solid additive composition pellets

[095] Freeze-dried NaF pellets were produced with the compositions as illustrated in Table 1 Table 1

[096] An aqueous mixture was prepared comprising the respective amounts of sodium formate and maltodextrin to arrive at the above compositions of table 1 (using 100 grams water and grams of the components as mentioned above).

[097] The respective aqueous mixtures were subsequently frozen by dripping in liquid nitrogen. The dripping needles selected in combination with the applied flow, produced frozen pellets of 3 to 5 mm.

[098] The frozen pellets were subsequently freeze dried. Freeze drying was carried out in a Christ Epsilon 2-4 LCSplus plate freeze-dryer at 0.50 mbar during primary drying and 0.175 mbar during secondary drying while applying a shelf heating temperature up to 50°C. The time needed to freeze dry until an Aw of 0.05 was measured and the values are listed in table 2 below.

Table 2

As illustrated by table 2 and figure 1 the solid additive compositions (according to the invention) comprising sodium formate formulated with maltodextrin (2 and 4 w/w% on dm) is freeze drying faster than the NaF as such (100%). The overall drying time could therefore advantageously be reduced by more than 15-20%.

Example 2, storage stability of the freeze-dried solid additive composition pellets

[099] The freeze-dried NaF pellets as prepared in example 1 and as listed in table 1 were tested during storage at 30°C for slightly more than 40 days. The increase in Aw was measured and the trendline is given in figure 2. The results are reflected in table 3 below. As illustrated by table 3 and figure 2 advantageously the solid additive compositions (according to the invention) allow for a reduction in Aw increase. That is, advantageously the increase in Aw is less compared to a composition not containing a crystallization inhibitor, such as sucrose or maltodextrin.

Table 3

Claims

1 . A process for preparing a frozen or freeze-dried solid additive composition, comprising or consisting of the following steps: a) preparing an aqueous mixture comprising or consisting of (i) formic acid or a salt or ester thereof and (ii) a crystallization inhibitor; b) preparing a frozen solid additive composition by freezing the aqueous mixture of a); and c) optionally preparing a freeze-dried solid additive composition by freeze-drying the frozen solid additive composition of b).

2. The process according to claim 1 , wherein the aqueous mixture of step a) does not comprise any type of lactic acid bacteria in any individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram, preferably of equal to or more than 1 x 10⁴ cfu/gram, before the freezing and/or freeze-drying of the mixture.

3. The process according to claim 1 or 2, wherein the frozen or freeze-dried solid additive composition does not comprise any type of lactic acid bacteria in any individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram, preferably of equal to or more than 1 x 10⁴ cfu/gram.

4. The process according to any one of claims 1 to 3, wherein the formic acid or a salt or ester thereof in the frozen or freeze-dried solid additive composition exists partly or wholly in an amorphous state.

5. The process according to any one of claims 1 to 4, wherein the aqueous mixture comprises or consists of:

- in the range from equal to or more than 0.05 % w/w to equal to or less than 20 % w/w, more preferably equal to or more than 0.5 % w/w to equal to or less than 10 % w/w, and most preferably equal to or more than 1% w/w to equal to or less than 7 % w/w of crystallization inhibitor, wherein the balance in the aqueous mixture preferably consists of water.

6. The process according to any one of claims 1 to 5, wherein step b) comprises dripping of the aqueous mixture into liquid nitrogen.

7. The process according to any one of claims 1 to 6, wherein the mixture is frozen or freeze-dried into the form of a pellet, a granule, a tablet, a powder or another solid particle.

8. The process according to any one of claims 1 to 7, wherein the frozen and/or freeze-dried solid additive composition is produced in the form of solid particles having an average diameter in the range from equal to or more than 1 mm to equal to or less than 10 mm, preferably in the range from equal to or more than 3 mm to equal to or less than 5 mm.

9. The process according to any one of claims 1 to 8, wherein a frozen solid additive composition is freeze-dried until an Aw of equal to or less than 0.2, preferably an Aw of equal to or less than 0.1 , is reached and a freeze-dried solid additive composition is produced.

10. The process according to any one of claims 1 to 9, wherein the crystallization inhibitor comprises or consists of sucrose, carboxymethyl cellulose, maltodextrin or a combination of two or more of these.

11 . The process according to any one of claims 1 to 10, wherein the crystallization inhibitor does not comprise or consist of maltodextrin.

12. The process according to any one of claims 1 to 10, wherein the crystallization inhibitor comprises or consists of maltodextrin.

13. The process according to any one of claims 1 to 12, wherein formic acid or a salt or ester thereof is sodium formate.

14. A solid additive composition, preferably frozen or freeze-dried, preferably in the form of a pellet, a granule, a tablet, a powder or another solid particle, comprising or consisting of:

- a crystallization inhibitor, which solid additive composition preferably does not comprise any type of lactic acid bacteria present, individually, in a viable cell count density of equal to or more than 1 x 10⁶ cfu/gram; and which solid additive composition preferably is obtained or obtainable according to any one of claims 1 to 13.

15. A kit of parts, or a starter culture composition, comprising:

- a first solid composition comprising or consisting of the solid additive composition according to claim 14; and - a second solid composition comprising or consisting of a solid culture composition, preferably frozen or freeze-dried, preferably in the form of a pellet, a granule, a tablet, a powder or another solid particle, wherein the solid culture composition comprises at least one type of lactic acid bacteria in an individual viable cell count density of equal to or more than 1 x 10⁶ cfu/gram.

16. A method for producing a cultured food product, the method comprising the steps of:

- inoculating a precursor material, preferably a milk base, with a kit of parts according to claim 15;

- culturing the precursor material with the kit of parts, and

- producing the cultured food product.

17. Use of the solid additive composition according to claim 14 or the kit of parts according to claim

15 in a process for making a food or feed product.