WO2025099255A1 - Process for the production of whey protein phospholipid concentrate - Google Patents

Abstract

The invention relates to a process for producing a whey protein phospholipid concentrate wherein at least one of the microbial reduction steps comprises a treatment with UV-C radiation, thereby avoiding denaturation of heat-sensitive bioactive ingredients and improving the sustainability of the whey protein phospholipid concentrate production process.

Description

PROCESS FOR THE PRODUCTION OF WHEY PROTEIN PHOSPHOLIPID CONCENTRATE

The present invention relates to a process for the production of whey protein phospholipid concentrate.

The best nutrition supplied to an infant is generally considered to be its own mother's milk; i.e. human milk. However, situations may arise wherein the infant cannot be fed human milk. In such cases, bovine milk-based formula milk is generally used to nourish the infant.

There is an ongoing effort to produce formula milk that approaches the composition of human milk as close as possible; in terms of protein and amino acid content, but also in terms of fat and carbohydrate composition and the perseverance of bioactivity of thermally sensitive ingredients.

Examples of such bioactive thermally sensitive ingredients are bioactive compounds in milkfat globule membrane (MFGM). Milkfat globules are composed of a triglyceride- rich core surrounded by a tri-layer membrane. This membrane, known as MFGM, is a complex mixture of phospholipids, cholesterol, and bio-active proteins in a unique trilayered structure.

Phospholipids are polar lipids with a (charged) head group and apolar tail. Glycerophospholipids contain two fatty acids on a glycerol backbone and a phosphate head group. Phosphosphingolipids have a sphingosine backbone of which the amide group is N-linked to a fatty acid and a phosphate head group. The five types of phospholipids present in milk are phosphatidylcholine (PC), phosphatidyletanolamine (PE), phosphatidylserine (PS), phosphatidylinositol (PI) and sphingomyelin (SPM). Phospholipids are strongly amphiphilic and are practically insoluble, but dispersible, in water and oil. They are highly surface active. The total phospholipid content of raw bovine milk, based on dry matter, is generally around 0.25-0.30 wt%.

MFGM has been associated with various health benefits - e.g. improved brain function and development, maturation of the gut, immune modulation, and antiviral action. When milk is separated in protein-rich, fat-rich (cream), and lactose-rich fractions, MFGM and, hence, phospholipids, mainly end up in the fat-fraction of whey or the whey-fraction of cream.

Examples of phospholipid-enriched milk fractions are cream, sweet buttermilk, alphaserum, beta-serum, cream serum, and whey protein phospholipid concentrate.

Whey protein phospholipid concentrate can be obtained by separating the fat fraction of sweet whey or acid whey using one or more microfiltration steps, optionally followed by drying to obtain a powder. Such a process is disclosed in WO 2022/112552 and WO 2023/001782. S. Sachadeva et al., Kieler Milchwissenschaftlicher Forschungsberichte 49 (1997) 47-68, discloses the recovery of phospholipids from buttermilk.

Conventional preparation processes for whey protein phospholipid concentrate contain heat treatment steps that may cause denaturation of the bioactive ingredients. These process steps are microbial reduction steps, conducted in order to guarantee microbiological safety. An example of such a microbial reduction step is thermal pasteurization (at least 72°C for at least 15 seconds). Denaturation of most bioactive proteins, however, starts above 65°C, meaning that the nutritional value of the product will be negatively affected by such heat treatment. In addition, such heat treatment requires a significant amount of energy.

One such heat-treatment is generally conducted to the whey; before the filtration step(s). Removal of microorganisms at this early stage is important in order to prevent an increase in the bacterial load during processing and to prevent microorganisms from clogging the microfiltration membrane.

Another such heat-treatment is conventionally applied just before spray-drying, where the whey protein phospholipid concentrate is heated in a pasteurizer under thermal pasteurization conditions, transported via a tower feeding line to the top of the spraydrying tower, and atomized in the spray-dryer. During transport to the top of the spraydrier, the product is not actively cooled and thus stays around 72°C, as cooling would negatively impact the capacity and energy input of the spray-drying process and would negatively affect the size of the spray-dried particles. In practice, this means that the whey protein phospholipid concentrate is at a temperature of at least 72°C for a period several minutes. Another microbial reduction step that may be applied to dairy streams, also in the production of whey protein phospholipid concentrate, is ceramic microfiltration (CMF). This ceramic microfiltration step involves the use of a microfiltration membrane with a pore size in the range 0.1 -10 pm and is therefore capable of removing bacteria and spores, but not able to separate casein and whey proteins. In contrast to pasteurization, CMF avoids high temperature (>65°C) treatment and thus does not affect heat-sensitive components such as proteins, flavour and viscosity. In addition, CMF is able to remove heat resistant microorganisms that are more resistant to conventional thermal pasteurization conditions.

However, ceramic microfiltration requires a significant amount of water and energy. Removal of this step, without negatively affecting microbiological quality and nativity of heat-sensitive components, would therefore significantly improve the sustainability of the whey protein phospholipid concentrate production process.

The object of the present invention is the provision of a more sustainable and energy efficient process for producing microbiologically safe whey protein phospholipid concentrate. A further object is the provision of a process that minimizes denaturation of heat-sensitive bioactive ingredients in said concentrate.

It has now been found that these objects can be achieved by applying a treatment with UV-C radiation during at least one of the microbial reduction steps in the production of protein phospholipid concentrates. In a preferred embodiment, at least one thermal pasteurization and/or a CMF step is replaced with a UV-C treatment, thereby avoiding denaturation of heat-sensitive bioactive ingredients and improving the sustainability of the whey protein phospholipid concentrate production process. Alternatively, one or more of the microbial reduction steps involve the simultaneous application of thermal treatment and UV-C treatment. The UV-C treatment is able to inactivate thermo- resistant bacteria, such as Streptococcus thermophilus and Microbacterium spp., while the high temperature during UV-C treatment improves the effectiveness of said treatment.

Ultraviolet (UV) irradiation is subdivided by wavelength into UV-A (320-400 nm), UV- B (280-320 nm), UV-C (200-280 nm), and Vacuum-UV (100-200 nm). UV-C has the highest germicidal effect, specifically between 250 and 270 nm, and is capable of destroying bacteria, viruses, protozoa, yeasts, molds and algae. The penetration depth of UV-C in a liquid depends on the absorbance and scattering of UV light by said liquid.

The use of UV-C irradiation in microbial reduction or disinfection of transparent liquids is well known. Milk and dairy products, however, are not transparent.

M.M. Delorme, Trends in Food Science 102 (2020) 146-154, reviewed the treatment of milk and dairy products with UV-C radiation. Advantages of UV-C treatment of dairy products are an effective deactivation of microorganisms with minimal loss in nutritional and sensory quality and the absence of toxic effects and waste generation. Compared to conventional heat pasteurization, UV-C treatment may require 1000 times less energy. A major disadvantage, however, relates to the fact that milk and dairy products are non-transparent and have a high absorption coefficient at UV-C wavelengths. In other words, the penetration power of the UV-C light is limited, which makes it difficult to ensure that all microorganisms are directly exposed to UV-C light.

In other words, the UV-C treatment of milk or whey - as disclosed in, e.g., WO 2017/027091 , WO 2019/057257, WO 2021/063462, WO 2019/076413, US 2022/0305155, C. Schubert et al., Int. Dairy J. 147 (2023) 105785, L. Christen et al., PLOS ONE (8) 2013 e68120, W. Zhang et al., LWT - Food Sci. Techn., 141 (2021 ) 110945, P. Padademas et al., Animals 11 (2021 ) 42, C. Michel et al., Int. Dairy J. 122 (2021 ) 105149, and J. A. Ansari et al., Innovative Food Science and Emerging Technologies 52 (2019) 387-393 - is already a challenge; the successful microbial reduction in more concentrated dairy streams such as whey protein concentrates, is considered to be an even bigger challenge.

Despite these expected problems, the inventors have found that UV-C treatment is able to reduce the bacterial load in more concentrated dairy streams to such an extent that it can replace at least one of the conventionally applied microbial reduction steps in the production of whey protein phospholipid concentrate.

The present invention therefore relates to a process for producing a whey protein phospholipid concentrate, said process comprising the steps of: a) providing whey, the whey being selected from sweet whey and acid whey, b) subjecting said whey to at least one microbial reduction step to provide a decontaminated whey, c) concentrating the decontaminated whey by ultrafiltration to obtain a whey concentrate as retentate, d) optionally subjecting said whey concentrate to at least one microbial reduction step, e) subjecting the whey concentrate to microfiltration, thereby obtaining a whey protein phospholipid concentrate as the retentate, f) optionally subjecting said whey protein phospholipid concentrate to at least one microbial reduction step, and g) spray-drying the whey protein phospholipid concentrate, wherein at least one of the microbial reduction steps comprises a treatment with UV- C radiation.

The process of the present invention is able to result in whey protein phospholipid concentrates with a plate count at least similar to that obtained with solely thermal pasteurization steps, while at the same time significantly reducing the denaturation of bioactive compounds. It additionally results in whey protein phospholipid concentrates from which thermo-resistant bacteria, such as Streptococcus thermophilus an Microbacterium, are inactivated; bacteria that cannot be inactivated with thermal pasteurization treatments.

The germicidal properties of IIV-C radiation are mainly due to inactivation of bacteria and viruses through DNA mutations induced through absorption of UV light by DNA molecules at very specific germicidal wavelengths, which are normally between 253. 7 and 254.1 nm.

The main commercial UV sources that emit sufficient energy in the germicidal wavelength range are mercury and deuterium lamps. The radiation of mercury lamps is more intense, while deuterium lamps have a broader emission spectrum.

For non-transparent liquids, UV-treatment requires sufficient turbulence. Turbulence can be expressed with the Reynolds number (Re) and is influenced by the diameter of the tube through which the liquid is transported, the flow through the tube, and the viscosity of the liquid. Said viscosity, in turn, depends on the nature of the liquid, its dry matter content, and its temperature.

The UV-C treatments in the process of the present invention are preferably conducted on liquid streams with a Reynolds number (Re) of at least 700, preferably at least 1 ,000, more preferably at least 1 ,500, even more preferably at least 2,300, more preferably at least 3,000, even more preferably at least 4,000, more preferably at least 6,000, and most preferably at least 10,000. In order to prevent protein denaturation, the UV-C treatment is preferably performed at a temperature below 70°C, more preferably below 60°C.

On the other hand, in order to lower the viscosity, it is desired to conduct said UV-C treatment at temperatures above the standard treatment temperature of dairy streams (5°C). Therefore, the UV-C treatment is preferably performed at a temperature in the range 10-70°C, more preferably 10-60°C, even more preferably 20-60°C, most preferably 30-60°C.

Oxidation of any compounds can be minimized by using extra light filters to narrow the bandwidth of the light spectrum (as disclosed in WO 2021/063462), or by degassing the feed stream before the UV-C treatment in order to remove oxygen/air.

In the present specification, milk, acid whey, sweet whey, whey concentrate and whey protein phospholipid concentrate are considered to have been decontaminated if the total plate count has been reduced to a lower value. Total plate count can be determined by ISO 4833-1 :2013, part 1 (pour plate). The total plate count is preferably reduced to less than 1000 CFU/ml, more preferably less than 100 CFU/ml, and most preferably less than 10 CFU/ml.

For thermal pasteurization, various suitable time and temperature combinations can be used. Examples of suitable combinations are: 72-75°C for 15-20 seconds, 63-65°C for 30-40 minutes, and 80-85°C for 1 -5 seconds.

Legal pasteurization requires a composition to be held at >72°C for at least 15 seconds for every part of the product, or equivalent as defined in the Pasteurized Milk Ordinance of the FDA. UV-C treatment can replace one or more of the CMF and/or thermal pasteurization steps usually applied in the production of whey protein phospholipid concentrate.

The process of the present invention starts with the provision of whey. This whey can be either acid whey or sweet whey, with sweet whey being preferred. The term “sweet whey” refers to the serum fraction resulting from the production of cheese from milk. A cheesemaking process generally involves precipitation of caseins by the addition of a coagulant and an acidifier to said milk. Acid whey, on the other hand, is produced by subjecting the milk, preferably after skimming and pasteurization, to a casein precipitation process. This casein precipitation involves the addition of an acid to induce coagulation of casein, resulting in an acid casein fraction (a casein curd) and an acid whey fraction.

The milk that is used for the production of the whey used in the process of the present invention is preferably a ruminant milk. The term “ruminant” includes true ruminants, like cattle, sheep, and goats, and pseudo-ruminants, like camels. The milk is preferably obtained from cattle or goats, meaning that bovine milk and goat milk are the preferred sources. Bovine milk, more in particular cow’s milk, is the most preferred source of milk for making the whey.

The target pH for the acidification of the milk to make cheese and whey is preferably in the range 4.8 to 5.7, more preferably 4.9 to 5.5. Suitable acidifiers include starter cultures (bacterial acidifiers; which convert lactose into lactic acid), acids, acidulants (such as Glucono Delta Lactone or GDL), and combinations of two or more of these. The most common starter cultures include thermophilic starters, typically starters from CSK, Chr. Hansen, or DuPont. Thermophilic starters by Chr. Hansen include frozen cultures STI-02, STI-03, STI-04, STI-06 and freeze-dried cultures STI-12, STI-13 and STI-14. Mesophilic starters may also be used.

Suitable coagulants are known in the art and include, for instance, calf rennet, fermentation-produced rennet and microbial rennet. Examples of calf rennet include Kalase produced by CSK and Naturen produced by Chr. Hansen. Examples of fermentation-produced rennet include Fromase by DSM and Milase by CSK. Examples of microbial rennets are Chy-Max by Chr. Hansen and Maxiren by DSM. Other coagulants include pepsin and various proteolytic enzymes of plant origin. According to step b) of the process of the present invention, whey is subjected to a microbial reduction step, which results in a decontaminated whey.

In one embodiment, IIV-C radiation is used in step b). Performing IIV-C treatment - and thus: inactivating microorganisms, including thermoresistant ones - at this early stage in the process, allows for a very clean process and minimized microbial content at the end of the process. Furthermore, the dry matter content of the liquid to be treated is relatively low at this stage, meaning that absorbance and scattering is also relatively low.

IIV-C treatment can be the sole microbial reduction in step b).

In the alternative, step b) may involve IIV-C treatment simultaneous with a thermal treatment, such as a thermal pasteurization treatment.

In another alternative, IIV-C treatment in step b) can be preceded or followed by thermal treatment, such as a thermal pasteurization treatment.

The dry matter content of the whey that is to be subjected to IIV-C treatment is preferably in the range 5-15 wt%, more preferably 7-12 wt%. This dry matter content may be adjusted to a specific value in this range in order to have a constant effectiveness and to prevent batch-to-batch variations in IIV-C effectiveness. In practice, this adjustment generally involves dilution with water.

The temperature of the whey during the IIV-C treatment is preferably in the range 10- 70°C, preferably 10-60°C, more preferably 20-60°C, most preferably 30-60°C

The decontaminated whey is concentrated by ultrafiltration in order to remove small molecules (e.g. salts and lactose) and water via the permeate and obtaining a whey concentrate as the retentate. This whey concentrate preferably has a dry matter content in the range 5-20 wt%, more preferably 10-20 wt%, and most preferably I Q- 17 wt%.

This ultrafiltration can be performed in conventional ways, well-known to the person skilled in the art. For instance, this ultrafiltration can be performed at a temperature in the range of either 5-20°C or 45-55°C. The temperature is preferably in the range 5- 20°C, more preferably 5-15°C, most preferably 5-12°C in view bacterial growth.

All conventional types of ultrafiltration membranes - spiral wound, ceramic, hollow fibre, etc. - can be used. The membrane can be constructed from various polymer types - such as polysulfone (PS), (modified) polyethersulfone (PES), polyvinylidene difluoride (PVDF), polyacrylonitrile (PAN), cellulose acetate (CA), and polypropylene (PP) - and several ceramic materials - such as aluminum oxide, zinc oxide, and titanium oxide.

The molecular weight cut-off (MWCO) of the membrane is preferably in the range 1- 10 kDa, more preferably in the range 5-10 kDa.

The feed pressure is preferably in the range 0.5-10 bar and the pressure drop per element in the range 0.5-1 .5 bar.

The ultrafiltration may be implemented as a single pass filtration or by using a series of membranes arranged in series. Preferably, the ultrafiltration is arranged as crossflow filtration.

The whey concentrate may subsequently be subjected to a microbial reduction step; step d). This microbial reduction step may comprise a ceramic microfiltration, a treatment with IIV-C radiation, or - most preferably - a combination thereof. If both treatments are used in combination, the IIV-C treatment may be either followed or preceded by ceramic microfiltration. The advantage of performing IIV-C treatment after ceramic microfiltration is that the microbial reduction achieved by ceramic microfiltration improves the effectiveness of the IIV-C treatment. In addition, water is added to the whey concentrate during ceramic microfiltration, thereby reducing the dry matter content and thus lowering the absorbance and scattering so that the IIV-C radiation can penetrate deeper. Furthermore, the IIV-C treatment may kill any bacteria grown during ceramic microfiltration, especially during long runs.

Ceramic microfiltration (CMF) is a well-known technique for the removal particles, such as bacteria and spores. Membranes with a pore size the range 0.1-10 pm, preferably 0.5-2 pm, even more preferably 0.5-1 .8 pm, and most preferably 0.7-1.5 pm, are preferably used. CMF can be performed at a temperature in the range 5-20°C or 40- 60°C. Lower temperatures are preferred from microbiological perspective; higher temperature processing, on the other hand, results in more efficient separation and allows the use of a smaller membrane surface. The temperature is preferably in the range 40-60°C, more preferably 45-55°C.

In order to achieve the required turbulence, the whey concentrate may have to be diluted with water in order to reduce its dry matter content. The extent of this dilution will depend on, e.g., the viscosity of the whey concentrate at the applied temperature, the tube diameter through which the whey concentrate flows during the IIV-C treatment, and the flow rate through the tube.

The advantage of performing IIV-C early in the process is that the stream to be treated has a lower microbial content than later in the process, meaning that there are less microbes to inactivate at this stage. The disadvantage of performing IIV-C at this stage is that it allows for recontamination or microbial growth during the remainder of the process.

The whey concentrate is subsequently subjected to at least one microfiltration step in order to obtain a phospholipid-enriched fraction - i.e. the whey protein phospholipid concentrate - as retentate. A whey protein enriched fraction is obtained as the permeate.

This microfiltration can be performed in conventional ways, well-known to the person skilled in the art. For instance, this microfiltration can be performed at a temperature in the range of either 10-20°C or 50-55°C. The temperature is preferably in the range 10-20°C, more preferably 10-15°C.

All conventional types of microfiltration membranes - spiral wound, ceramic, hollow fibre, etc. - can be used. The membrane can be constructed from various polymer types - such as polysulfone (PS), (modified) polyethersulfone (PES), polyvinylidene difluoride (PVDF), polyacrylonitrile (PAN), cellulose acetate (CA), and polypropylene (PP) - and several ceramic materials - such as aluminum oxide, zinc oxide, and titanium oxide.

The molecular weight cut-off (MWCO) and pores size of the membrane is preferably in the range 50-1000 kDa and/or 0.01-1.0 micrometer, more preferably in the range 100-800 and/or 0.02-0.5 micrometer, most preferably in the range 100-500 kDa and/or 0.05-0.2 micrometer.

The microfiltration is preferably operated with a trans-membrane pressure of 0.1-5 bar, more preferably 0.2-3 bar, most preferably 0.2-1 bar.

The resulting retentate (i.e. the whey protein phospholipid concentrate) preferably has been concentrated to a dry matter content in the range 5-22 wt%, more preferably I Q- 20 wt%, and most preferably 12-18 wt%.

The microfiltration may be implemented as a single pass filtration or by using a series of membranes arranged in series. Preferably, the microfiltration is arranged as crossflow filtration. The feed flow rate is preferably in the range 14 to 16 m³/hr.

The flux over the membranes, i.e. the ratio between product flow and membrane surface, is preferably relatively low, more preferably in the range 2-30, preferably 2-12 l/m²/hr.

The microfiltration is preferably combined with diafiltration, more preferably with a ratio between diafiltration flow and membrane surface in the range 0.5-30, more preferably 0.5-15, and most preferably 1 -8 l/m²/hr.

The cross flow over the membranes is preferably in the range of 0.2-3 m³/hr.

The whey protein phospholipid concentrate may optionally be further concentrated, preferably to a dry matter content of at least 25 wt%, more preferably at least 30 wt%. This can be achieved with ultrafiltration and/or nanofiltration.

The whey protein phospholipid concentrate is finally spray-dried.

Before such spray-drying, the whey protein phospholipid concentrate may be subjected to a microbial reduction - step f).

In one embodiment, said microbial reduction comprising a treatment with IIV-C radiation. Performing this treatment at this late stage in the process ensures microbiological quality, including the inactivation of thermo-resistant microorganisms. This IIV-C treatment may be the sole microbial reduction in step f). In the alternative, step f) may involve IIV-C treatment simultaneous with a thermal treatment. In another alternative, the IIV-C treatment can be preceded or followed by thermal treatment.

The big advantage of treating the product just before the drying tower is that there is almost no risk of recontamination or microbial growth after the microbial reduction step.

In order to achieve the required turbulence for the IIV-C treatment, the whey protein phospholipid concentrate may have to be diluted with water in order to reduce its dry matter content. The extent of this dilution will depend on, e.g., the viscosity of the whey protein phospholipid concentrate at the applied temperature, the tube diameter through which the whey protein phospholipid concentrate flows during the IIV-C treatment, and the flow rate through the tube. An appropriate dry matter content is generally in the range 6-12 wt%, preferably 6-10 wt%. Furthermore, the temperature of the whey protein phospholipid concentrate during the UV-C treatment is preferably in the range 10-70°C, preferably 10-60°C, more preferably 20-60°C, most preferably 30-60°C

Prior to, during, or after such UV-C treatment, the whey protein phospholipid concentrate may be pre-heated to a temperature in the range 40-65°C, preferably 50- 60°C, which is below the denaturation temperature of most (health promoting) bioactive molecules. Pre-heating should not result in temperatures exceeding 65°C. This pre-heating serves to lower the electric energy input during the subsequent spraydrying step and thus to make the process more energy-efficient. This pre-heating can be performed batch-wise or continuously in any suitable equipment. Batch-wise preheating can be performed in a vessel; continuous pre-heating can be performed using a heat exchanger (e.g a plate heat exchanger). The pre-heating is preferably performed continuously.

After reaching the desired pre-heating temperature, the pre-heated liquid composition is transported to the top of a spray drying tower. There is no need for holding the composition at the desired temperature for a certain time period before conducting said transport. Therefore, the entire process can be performed in a continuous manner. If not pre-heated, the whey protein phospholipid concentrate is preferably held in step e) below 20°C, more preferably below 15°C, and most preferably below 10°C until reaching the top of the spay-drying tower.

The resulting whey protein phospholipid concentrate is spray-dried. Spray-drying is preferably performed using hot air with a temperature in the range 140-300°C, preferably 150-260°C, and most preferably 170-210°C.

Any type of spray-dryer can be used, such as Single Stage, 2-Stage, Multi Stage and Filtermat® type spray dryers.

The whey protein phospholipid concentrate that results from the process of the present invention preferably has a total phospholipid content, based on dry matter, of at least 1 .0 wt%, preferably at least 1 .5 wt%, more preferably 2.5 wt%, even more preferably at least 4 wt%, more preferably at least 6 wt%, even more preferably at least 7 wt%. even more preferably in the range 7-20 wt%, and most preferably in the range 7-15 wt%. This total phospholipid content can be determined by lipid extraction using a Rose- Gotlieb method and the subsequent determination of phosphorus in the lipid extract using ICP (Induced Coupled Plasma).

Sphingomyelin is a phospholipid present in milk, but not present in vegetable phospholipid sources. The sphingomyelin content in the whey protein phospholipid concentrate resulting from the process of the present invention, based on total phospholipid content, is preferably at least 20 wt%, more preferably in the range 20- 35 wt%, and most preferably in the range 20-30 wt%. This sphingomyelin content can be determined with ¹³P-NMR.

The whey protein phospholipid concentrate prepared according to the process of the present invention can be used to prepare formula milk by combining the whey protein phospholipid concentrate with at least a further protein source, a further lipid source, a carbohydrate source, vitamins, and minerals.

The further protein source generally includes milk, whey protein concentrate, serum protein concentrate, and/or hydrolyzed (whey) proteins.

The further lipid source may be any lipid or fat suitable for use in formula milk. Preferred fat sources include milk fat, safflower oil, egg yolk lipid, canola oil, olive oil, coconut oil, palm kernel oil, soybean oil, fish oil, palm oleic, high oleic sunflower oil and high oleic safflower oil, and microbial fermentation oil containing long-chain, polyunsaturated fatty acids. In one embodiment, anhydrous milk fat is used. The lipid source may also be in the form of fractions derived from these oils such as palm olein, medium chain triglycerides, and esters of fatty acids such as arachidonic acid, linoleic acid, palmitic acid, stearic acid, docosahexaeonic acid, linolenic acid, oleic acid, lauric acid, capric acid, caprylic acid, caproic acid, and the like. It may also be added small amounts of oils containing high quantities of preformed arachidonic acid and docosahexaenoic acid such as fish oils or microbial oils. The fat source preferably has a ratio of n-6 to n-3 fatty acids of about 5:1 to about 15:1 ; for example about 8:1 to about 10:1. In a specific aspect, the infant formula comprises an oil mix comprising palmitic acid esterified to triacylglycerols, for example wherein the palmitic acid esterified in the sn-2 position of triacylglycerol is in the amount of from 20% to 60% by weight of total palmitic acid and palmitic acid esterified in the sn-1/sn-3 position of triacylglycerol is in the amount of from 40% to 80% by weight of total palmitic acid. Examples of vitamins and minerals that are preferably present in formula milk are vitamin A, vitamin B1 , vitamin B2, vitamin B6, vitamin B12, vitamin E, vitamin K, vitamin C, vitamin D, folic acid, inositol, niacin, biotin, pantothenic acid, choline, calcium, phosphorous, iodine, iron, magnesium, copper, zinc, manganese, chloride, potassium, sodium, selenium, chromium, molybdenum, taurine, and L-carnitine. Minerals are usually added in salt form.

Examples of carbohydrates that are preferably present in formula milk are lactose, non-digestible oligosaccharides such as galacto-oligosaccharides (GOS), fructooligosaccharides (FOS), inulin, xylo-oligosaccharides, and human milk oligosaccharides (HMOs). Suitable HMOs include 2’-FL, 3-FL, 3’-GL, 3’-SL, 6’-SL, LNT, LNnT, and combinations thereof. HMO’s are commercially available or can be isolated from milk in particular from human breast milk.

If necessary, the nutritional composition may contain emulsifiers and stabilisers such as soy lecithin, citric acid esters of mono- and di-glycerides, and the like. It may also contain other substances which may have a beneficial effect such as lactoferrin, nucleotides, nucleosides, probiotics, and the like.

Suitable probiotics include Lactobacteria, Bifidobacterium lactis such as Bifidobacterium lactis Bb12, Streptococcus thermophilus, Lactobacillus johnsonii La1, Bifidobacterium longum BL999, Lactobacillus rhamnosus LPR, L rhamnosus GG, Lactobacillus reuteri, Lactobacillus salivarius. Such prebiotics are commercially available.

Formula milk is usually prepared for bottle-feeding or cup-feeding from powder (mixed with water) or liquid (with or without additional water).

Formula milk is generally available as a spray-dried powder. Spray-drying involves an additional heating step. In order to preserve as much of the native proteins as possible, it is desired to keep the heating conditions during spray-drying as mild as possible.

In order to reduce the number of processing steps that may denature any proteins, it is preferred to dry blend the whey protein phospholipid concentrate with the other ingredients of the formula milk.

The formula milk according to the invention can be in the form of a dry, semi-dry or liquid composition. For example, it can be a powdered composition that is suitable for making a liquid composition after reconstitution with an aqueous solution, preferably with water.

In another embodiment, it is a liquid composition, for instance a ready-to-consume drinkable or spoonable composition.

EXAMPLES

Comparative Example 1

A whey protein phospholipid concentrate (WPPC) was prepared by subjecting cheese whey to thermal pasteurization. The pasteurized whey was concentrated by ultrafiltration, the retentate was subjected to ceramic microfiltration and the resulting permeate was subjected to microfiltration.

The resulting retentate - i.e. the WPPC - had a dry matter content of 16 wt%.

This WPPC was subsequently pasteurized at different temperatures, ranging from 65°C to 73°C, using a continuous flow micro pasteurizer equipped with tubular heat exchangers and holders in order to mimic industrial pasteurization. The holding time in all experiments was 180 seconds, thereby representing an industrial 18 seconds pasteurization followed by 162 seconds of transport to the top of a spray-drying tower.

Total plate count, also known as aerobic mesophilic count, was determined according to ISO 4833-1 :2013. 1 ml of product was poured in plate count milk agar and plates were incubated for 72 hours aerobically at 30°C. Plates were subsequently counted and the number of colony forming units (CFU) per ml of product was reported for a dilution where the number of observed colonies on the plate was between 10 and 300. For the low thermoresistant plate count, the sample was treated at 63.5°C for 30 minutes prior to plating. This method is equivalent to NEN 6807.

For the high thermoresistant plate count, the sample was treated at 80°C for 5 minutes prior to plating.

The total plate count in all experiments was reduced from 2.2- 10⁶ cfu/ml to T10³ cfu/ml after pasteurization at 65-71 °C and 10 cfu/ml after pasteurization at 73°C. The number of high and low thermoresistant bacteria and spores started to decrease after pasteurization above 70°C. As measure of the bioactivity of the proteins, the native IgG and lactoferrin contents were determined using the bovine IgG and lactoferrin ELISA quantitation set as described by R.L. Valk-Weeber, T. Eshuis-de Ruiter, L. Dijkhuizen, and S.S. van Leeuwen, International Dairy Journal, Volume 110, November 2020, 104814.

The thermal treatment resulted denaturation of bioactive compounds. After pasteurization at 73°C, the native IgG and IgA contents were reduced to 60% of their original value; the native IgM content was reduced to 20% of its original value, and the native lactoferrin content was reduced to 40% of its original value.

In addition, the vitamin B5, B6, and B8 contents were significantly reduced.

The resulting WPPC was spray-dried on a small scale pilot dryer.

Example 2

Comparative Example 1 was repeated, except that, instead of thermal pasteurization of the WPPC, said WPPC was submitted to IIV-C treatment using Lyras® pilot UV-C equipment with a capacity of 100-800 L/h. In view of the high turbidity of the 16 wt% dry matter suspension, the suspension was first diluted to a dry matter content of 8 wt%.

Cold (10°C) diluted WPPC was pumped through spiral wound transparent tubes (7 mm diameter) with maximum possible flow (406 L/h) and a Reynolds number of 3959. This flow was limited by the pump capacity and the allowed pressure drop of the UV- C equipment.

The WPPC in the tubes was exposed to light using lamps placed at the outside of the spiral. Between the lamps and the tube, filters were placed in order to ensure a small wavelength peak around 254 nm, thereby minimizing chemical side reactions like oxidation reactions.

The UV-C power used - i.e. the percentage of the maximum power of the equipment - was 80%.

As a result of this UV-C treatment, the total plate count was reduced from 2.2- 10⁶ cfu/ml to 20 cfu/ml; and the thermoresistant microorganisms - that could not be inactivated by pasteurisation (LTR and HTR) - were reduced from 2000 cfu/ml to 20 cfu/ml. It was further observed that the native IgG and lactoferrin contents were not affected by UV-C treatment. They remained stable at a value of 327 mg/100 g and 33.5 mg/100 g, respectively. Also the native IgA and IgM contents remained above 90% of their initial values. In addition, the vitamin B5, B6, and B8 remained the same.

Subsequent spray-drying slightly reduced the native IgG and lactoferrin contents; to, respectively, 95% and 90% of their original value.

Example 3

Example 2 was repeated, except that the suspension was first diluted to a dry matter content of 12.5 wt% and heated to 30°C before being pumped through the spiral wound transparent tubes and exposed to UV-C light.

Total plate count, thermoresistant microorganism content and the extent of denaturation of the bioactive compounds was similar to that obtained in Example 2.

Example 4

Example 2 was repeated, except that the suspension was first diluted to a dry matter content of 14.7 wt% and heated to 52°C before being pumped through the spiral wound transparent tubes and exposed to UV-C light.

Different flow rates, in the range 57-448 L/h, were used. With increasing flow rate, not only Reynolds numbers increase, but also residence time and, hence, UV-C energy input. When correcting for the difference in energy input, it was observed that inactivation of microorganisms (total plate count), including thermoresistant microorganisms, significantly increased at Reynolds numbers of 1500 and above.

Claims

1 . Process for producing a whey protein phospholipid concentrate, said process comprising the steps of: a) providing whey, selected from acid whey and sweet whey, b) subjecting said whey to at least one microbial reduction step to provide a decontaminated whey, c) concentrating the decontaminated whey by ultrafiltration to obtain a whey concentrate as retentate, d) optionally subjecting said whey concentrate to at least one microbial reduction step, e) subjecting the whey concentrate to microfiltration, thereby obtaining a whey protein phospholipid concentrate as the retentate, f) optionally subjecting said whey protein phospholipid concentrate to at least one microbial reduction step, and g) spray-drying the whey protein phospholipid concentrate, wherein at least one of the microbial reduction steps comprises a treatment with UV-C radiation.

2. Process according to claim 1 wherein said treatment with UV-C radiation is conducted on a liquid stream with a Reynolds number (Re) of at least 700, preferably at least 1 ,000, more preferably at least 1 ,500, even more preferably at least 2,300, more preferably at least 3,000, even more preferably at least 4,000, more preferably at least 6,000, and most preferably at least 10,000.

3. Process according to claim 1 or 2 wherein said treatment with UV-C radiation is performed at a temperature in the range 10-70°C, preferably 10-60°C, more preferably 20-60°C, most preferably 30-60°C.

4. Process according to any one of the preceding claims wherein step b) comprises a treatment of the whey with UV-C radiation.

5. Process according to claim 4 wherein the treatment with IIV-C in step b) is the sole microbial reduction after step a) and before step e).

6. Process according to claim 4 wherein step b) involves simultaneous thermal pasteurization and IIV-C radiation treatment.

7. Process according to any one of claims 4-6 wherein the dry matter content of the whey that is subjected to IIV-C treatment is in the range 5-15 wt%, preferably 7- 12 wt%.

8. Process according to any one of claims 4-7 wherein the whey that is subjected to IIV-C radiation has a temperature in the range 10-70°C, preferably 10-60°C, more preferably 20-60°C, most preferably 30-60°C.

9. Process according to any one of the preceding claims wherein said whey concentrate is subjected to at least one microbial reduction step in step d), said microbial reduction step comprising a treatment with IIV-C radiation, optionally followed or preceded with ceramic microfiltration.

10. Process according to claim 9 wherein the microbial reduction step of step d) involves ceramic microfiltration followed by a treatment with IIV-C radiation.

11 . Process according to any one of the preceding claims wherein the whey protein phospholipid concentrate is subjected to at least one microbial reduction in step f), said microbial reduction comprising a treatment with IIV-C radiation, optionally followed or preceded by a thermal pasteurization step.

12. Process according to any one of claims 1 -11 wherein step f) involves simultaneous thermal pasteurization and IIV-C radiation treatment.

13. Process according to claim 11 or 12 wherein the whey protein phospholipid concentrate that is subjected to IIV-C radiation has a dry matter content of in the range 6-12 wt%, preferably 6-10 wt%.

14. Process according to any one of claims 11-13 wherein the whey protein phospholipid concentrate that is subjected to IIV-C radiation has a temperature in the range 10-70°C, preferably 10-60°C, more preferably 20-60°C, most preferably 30-60°C.