Manufacture of a novel cultured micellar casein concentrate ingredient for emulsifying salt–free process cheese products applications

Micellar casein concentrate (MCC) is a high protein ingredient that is typically produced using 3 stages of microfiltration with a 3× concentration factor and diafiltration. Acid curd is an acid protein concentrate, which can be obtained by precipitating the casein at pH 4.6 (isoelectric point) using starter cultures or direct acids without the use of rennet. Process cheese product (PCP) is a dairy food prepared by blending dairy ingredients with nondairy ingredients and then heating the mixture to get a product with an extended shelf-life. Emulsifying salts are critical for the desired functional characteristics of PCP because of their role in calcium sequestration and pH adjustment. The objectives of this study were to develop a process to produce a novel cultured micellar casein concentrate ingredient (cMCC; culture-based acid curd) and to produce PCP without emulsifying salts using different combinations of protein from cMCC and MCC in the formulations (2.0:1.0, 1.9:1.1, and 1.8:1.2). Skim milk was pasteurized at 76°C for 16 s and then microfiltered in 3 microfiltration stages using graded permeability ceramic membranes to produce liquid MCC (11.15% total protein; TPr and 14.06% total solids; TS). Part of the liquid MCC was spray dried to produce MCC powder (75.77% TPr and 97.84% TS). The rest of the MCC was used to produce cMCC (86.9% TPr and 96.4% TS). Three PCP treatments were formulated with different ratios of cMCC: MCC, including 2.0:1.0, 1.9:1.1, and 1.8:1.2 on the protein basis. The composition of PCP was targeted to 19.0% protein, 45.0% moisture, 30.0% fat, and 2.4% salt. This trial was repeated 3 times using different batches of cMCC and MCC powders. All PCP were evaluated for their final functional properties. No significant differences were detected in the composition of PCP made with different ratios of cMCC and MCC except for the pH. The pH was expected to increase slightly with elevating the MCC amount in the PCP formulations. The end apparent viscosity was significantly higher in 2.0:1.0 formulation (4,305 cP) compared with 1.9:1.1 (2,408 cP) and 1.8:1.2 (2,499 cP). The hardness ranged from 407 to 512 g with no significant differences within the formulations. However, the melting temperature showed significant differences with 2.0:1.0 having the highest melting temperature (54.0°C), whereas 1.9:1.1 and 1.8:1.2 showed 43.0 and 42.0°C melting temperature, respectively. The melting diameter (38.8 to 43.9 mm) and melt area (1,183.9 to 1,538.6 mm 2 ) did not show any differences in different PCP formulations. The PCP made with a 2.0:1.0 ratio of protein from cMCC and MCC showed better functional properties compared with other formulations


INTRODUCTION
Microfiltration (MF) is a membrane process that is used to fractionate casein and serum protein (SP) from skim milk using a 0.1-µm semipermeable membrane.The skim milk is driven force through the membrane to separate casein (retentate side) and SP (permeate side) based on their sizes (0.1-0.4 µm vs. 0.003-0.01µm, respectively).The product on the retentate side is called micellar casein concentrate (MCC), which is mostly native casein.The MCC is a high protein ingredient that is typically manufactured in 3 MF stages using a 3× concentration factor (CF) with diafiltration (DF).Several MF membranes have been studied to produce MCC (Hammam et al., 2021), such as spiral-wound membranes (Govindasamy- Lucey et al., 2007;Lawrence et al., 2008;Zulewska et al., 2009;Beckman et al., 2010;Beckman and Barbano, 2013;Marella et al., 2021), isoflux ceramic membranes (Adams and Barbano, 2013), uniform transmembrane pressure (UTP) ceramic membranes (Vadi and Rizvi, 2001;Hurt et al., 2010), and graded permeability (GP) ceramic membranes (Zulewska and Barbano, 2014;Tremblay-Marchand et al., 2016;Hammam et al., 2022a).The spiral-wound membranes are cheaper and have lower operating costs, but they have a shorter shelf-life, low chemical stability, limited viscosity range, and less efficiency to remove SP as compared with ceramic membranes.The UTP and GP membranes are commonly used to produce MCC due to their high SP removal.The GP membranes have low operating costs (do not require permeate recirculation pump), although they are expensive compared with UTP membranes (Hammam et al., 2021).
The MCC has promising applications in some dairy and nondairy products due to its unique physicochemical and functional characteristics (e.g., foaming, emulsifying, wetting, dispersibility, heat stability, bland flavor, and water-binding ability).The high casein content in MCC makes it heat stable and thereby it can be used in beverages that require sterilization (Beliciu et al., 2012;Sauer and Moraru, 2012).The nondairy applications for MCC are pasta, confectionery, meat products, special dietary preparations, textured products, convenience foods, toothpaste, cosmetics, and wound treating preparations (Hammam et al., 2021).The dairy applications for MCC include cheddar cheese (Amelia et al., 2013;Li et al., 2020;Xia et al., 2021), Feta cheese (Hammam et al., 2022c), Greek-style yogurt (Bong and Moraru, 2014), imitation mozzarella cheese (Hammam, 2022;Salunke et al., 2022), recombined MCC (Lu et al., 2016(Lu et al., , 2017)), process cheese (PC) and process cheese products (PCP; Hammam et al., 2022b;Salunke and Metzger, 2022a,b), and acid curd (Hammam, 2022).We recently developed a process to produce soft acid curd (basically cottage cheese curd) from MCC using direct acidification (lactic acid; Hammam et al., 2023) and starter cultures (Hammam, 2022) to set the curd.Although direct acids such as lactic acid take less time to produce the acid curd, those acids are costly and produce acid curd with a bland flavor compared with the varieties of starter cultures that cost less and can be utilized to develop flavors in the curd.
Acid curd is a protein concentrate, which can be obtained by precipitating the casein at pH 4.6 (isoelectric point) using starter cultures or direct acids without the use of rennet.The colloidal calcium phosphate that exists in the micelles is dissolved at the isoelectric point, which leads to a low mineral (calcium) content in the acid curd.In contrast to the low mineral content of acid curd, MCC has a high level of casein-bound calcium due to its pH of 6.5 to 6.7.Acid curd could be produced from skim milk in a process similar to cottage cheese manufacture (Klei et al., 1998).There is a possibility of using MCC instead of skim milk to produce acid curd.Making acid curd from MCC has advantages as compared with skim milk because manufacturing MCC using MF results in milk-derived whey protein as a byproduct, which can be used in many applications, particularly making whey protein isolate.In contrast, acid curd produced from skim milk results in acid whey as a byproduct, which is more difficult to use.The typical composition of MCC (3 stages using a 3× CF with a DF) is > 9% true protein (TP) and > 13% TS (Zulewska et al., 2009).This MCC could be used immediately in making acid curd or diluted to lower protein levels before making acid curd if required.For the rest of this paper, we will refer to acid curd made from micellar casein using cultures as cultured MCC ingredient (cMCC).In our previous studies, we tried to produce acid curd from MCC with different protein content (3, 6, and 9% protein).We found that MCC with 9% protein is the optimum product to produce acid curd (Hammam et al., 2023).
Process cheese and PCP are dairy foods prepared by blending dairy ingredients (e.g., natural cheese, protein concentrates, butter, nonfat dry milk, whey powder, permeate) with nondairy ingredients [e.g., sodium chloride, water, emulsifying salts (ES), color, and flavors] and then heating the mixture with continuous agitation to produce a homogeneous product with an extended shelf-life (Meyer, 1973;Thomas, 1973;Caric et al., 1985;Schmid, 1992;Guinee et al., 2004;Mcsweeney, 2007;Kapoor and Metzger, 2008;Kammerlehner, 2009;Hammam et al., 2022b).Process cheese and PCP are remarkably similar products; however, PC is defined by the Code of Federal Regulations, whereas PCP does not have a Code of Federal Regulations definition (FDA, 2006).The major ingredient in PC is natural cheese, whereas PCP may or may not contain cheese.In that case, they typically rely on other ingredients, such as MCC and whey protein concentrate for structure building (Lu et al., 2007;Hammam et al., 2021).Process cheese has been made since the late nineteenth and early twentieth century to extend the shelf-life of natural cheeses.Approximately one-third of all-natural cheese produced in the United States is used in making PC (Kommineni et al., 2012), which is considered one of the leading varieties of cheese in the world and has several applications (Sorensen, 2001;Kapoor and Metzger, 2008).
A critical reaction that occurs during PC and PCP manufacture is calcium sequestration using ES (sodium citrate, disodium phosphate, and so on).Emulsifying salts are critical for the functional characteristics of PC and PCP due to their role in improving the emulsification characteristics of casein by sequestrating a portion of the calcium from the calcium-casein-phosphate network in natural cheese or casein containing ingredients (Figure 1).As shown in Figure 1, ES such as disodium phosphate sequester the calcium from the calcium-casein-phosphate network by donating their sodium ions.As a result, the major molecular forces that cross-link the various monomers of casein are partially disrupted.This disruption leads to hydration and dispersion of caseins.The partially dispersed monomers of casein, such as any protein, have hydrophilic and hydrophobic regions.These amphiphilic dispersed caseins, in the presence of heat and mixing, get hydrated via hydrophobic interactions and at the same time emulsify fat phase via hydrophobic interactions to produce an emulsified PC (Guinee et al., 2004).
If cMCC is mixed with MCC (Figure 2), it may be possible to create a partially de-aggregated casein network without the use of ES.The ratio of cMCC to MCC will have an effect on the level of de-aggregation and the pH of the final PC product.We recently patented a method of producing PCP with no ES using cMCC and MCC (Metzger and Hammam, 2020).We hypothesized that a ratio of 2 parts of protein from cMCC to 1 part of protein from MCC will create a partially de-aggregated casein network similar to a typical PC that uses ES (Figure 2).
No studies have used MCC in making cMCC powder as well as using this ingredient in making PCP with no ES.Therefore, the objectives of this study were to develop a process to produce a novel cMCC powder ingredient and to produce PCP without ES using different combinations of protein from cMCC and MCC in the formulations (2.0:1.0,1.9:1.1, and 1.8:1.2).

Experimental Design
Manufacture of MCC was completed in approximately 10 h in 1 d at Davis Dairy Plant at South Dakota State University (Brooking, SD).The experiment was repeated 3 times with different lots of skim milk.Part of the final MCC was dried using a spray dryer to produce MCC powder, whereas the rest of the MCC was fermented using starter culture to produce cMCC, which was eventually dried in the fluid bed.The MCC and cMCC powders were mixed in different ratios to produce PCP with no ES.
No human or animal subjects were used, so this analysis did not require approval by an Institutional Animal Care and Use Committee or Institutional Review Board.

Preparation of Skim Milk
The MCC was manufactured as described in our previous study with some modifications (Hammam et al., 2022a).Approximately 750 kg of whole bovine milk was separated (Model MSE 140-48-177 AirTight centrifuge; GEA Co.) at 4°C at the South Dakota State University Davis Dairy Plant and then pasteurized in a plate heat exchanger (model PR02-SH, AGC Engineering) at 76°C for 16 s.The pasteurized skim milk was then kept at ≤4°C until MF was conducted.

Microfiltration Operation
To fractionate skim milk into casein and SP to produce MCC, a pilot-scale ceramic GP MF system (TIA, Rond-point des, Portes de Provence, Rue Robert Schumann 84500) was used.The GP MF system was equipped with 7 ceramic tubes (19 channels with a diameter of 3.3 mm) mounted in the system vertically.The ceramic GP MF membranes had a 0.1-µm pore size, 1.68 m 2 surface areas, and a 1.02-m membrane length.The GP MF system was also equipped with a feed pump and a retentate recirculation pump (TIA).The MF of skim milk (approximately 670 kg) was performed in 3 stages to produce MCC.

Manufacture of MCC
First Stage.The GP MF system was started with soft water at 50°C using 3× CF (1 kg of retentate:2 kg of permeate) in a feed and bleed mode (1-way pass) with 400 kPa retentate pressure inlet (Rpi), 200 kPa retentate pressure outlet (Rpo), and 200 kPa permeate pressure outlet (Ppo).The skim milk (~10.3%Brix) was heated to 50°C with a heat plate exchanger (SABCO Plate-pro Sanitary Chiller; NP925-41) before processing.When the processing conditions were stable while running with water, the system was transitioned to milk.The skim milk was MF with the GP MF system at a constant flux (71.42 L/m 2 per h) using a 3× CF in a feed and bleed mode at 50°C (Figure 3).The water at the beginning of the process was flushed out with skim milk by collecting about 36.0 kg of permeate and 17.0 kg of retentate.The permeate flow rate was set at 120 L/h (flux of 71.42 L/m 2 per h) and the retentate flow rate was 60 L/H to produce a 3× retentate.After this startup, retentate and permeate were collected and weighed continuously.During MF of skim milk, Rpi, Rpo, and Ppo were targeted to maintain 400, 200, and 200 kPa, respectively.The CF was calculated every 15 min by collecting permeate and retentate samples.The composition of retentate and permeate during MF was monitored using an infrared spectrophotometer (FT-IR spectroscopy, Model Dairyspec FT, Bentley Instrument Inc).A composite permeate sample was collected during the process.The collected retentate was kept in tanks during the MF process.The processing time of the first stage was approximately 4 h.
Second Stage.The retentate (~17.0%Brix) from the first stage was diluted with soft water (approximately 212.0 kg of retentate was mixed with 424.0 kg of water) to obtain a DF of 3× to get back the original volume of skim milk.After mixing, the diluted retentate (~5.5% Brix) was heated to 50°C and processed with the GP MF system using a 3× CF, as described in the first stage.The water at the beginning of the process was flushed out of the system with the diluted retentate by collecting about 36.0 kg of permeate and 18.0 kg of retentate.The processing conditions in the second stage were similar to the first stage.The Rpi, Rpo, and Ppo were set at 400, 200, and 200 kPa, respectively.The permeate flow rate was 120 L/h (flux of 71.42 L/m 2 per h) and the retentate flow rate was 60 L/h.Permeate and retentate were weighed continuously, as described in the first stage.A composite permeate sample was collected during the process.The retentate was collected in sanitized cans.The processing time of the second stage was approximately 3.5 h.
Third Stage.Approximately 175.0 kg of the retentate (~13.0%Brix) was MF in a recirculation mode.The retentate of the second stage was placed in the tank of MF unit and then proceed to the third stage using a 3× CF at 50°C.The processing conditions in the third stage were similar to the first and second stages.The following conditions were applied: Rpi, Rpo, and Ppo were 400, 200, and 200 kPa, respectively, whereas the permeate flow rate was 120 L/h (flux of 71.42 L/ m 2 per h) and the retentate flow rate was 60 L/h.The retentate was recirculated while permeate was collected until the TS reached approximately 14 to 15% (CEM Smart System5 SL7199) or ~16.0 to 17.0% Brix.Increasing the solids content of MCC during MF led to decreasing the Ppo.The decrease of Ppo is related to the concentration polarization and membrane fouling that accumulated on the membrane during recirculation.The final MCC resulting from the third stage (approximately 125.0 kg) was collected.A composite sample of the permeate was sampled for compositional analysis.The processing time for the third stage was around 1 h.The MCC was then pasteurized at 63°C/30 min.Approximately 50.0 kg of the liquid MCC was dried using a spray dryer to produce MCC powder, whereas the rest of the liquid MCC (75.0 kg) was used in making cMCC.This trial was replicated 3 times using 3 separate lots of raw milk.

Cleaning the Membrane
After processing, the GP MF system was flushed with soft water to remove all retentate residues from the system.The initial flux was measured with approximately 60 kg of soft water at 27°C.During the flux measurement, the retentate valves were closed and the permeate valves were completely opened with the feed pump running.Subsequently, 30 kg of soft water was added to the system and heated to 74°C, then 900 mL of Ultrasil 110 Alkaline cleaner (Ecolab Inc.) and 200 mL of XY 12 (Ecolab Inc.) were added to get pH 11 (Accumet, Fisher Scientific).This solution was recirculated for 30 min at a 350 L/h permeate flow rate (flux of 208 L/m 2 per h).After cleaning the MF system with the alkaline solution, the membrane was cooled to 50°C (less than 10°C per min).The alkaline solution was flushed out of the MF system with soft water until the pH of outlet water ranged from 8.3 to 8.5.The flux was measured again, as described previously.The system was cleaned with an acid solution (Ultrasil 78 acid cleaner) by adding 30 kg of soft water and heated to 52°C; subsequently, 400 mL of Ultrasil 78 (Ecolab Inc.) was added to get a pH of 2. The recirculation of the acid solution was applied for 20 min at a flux of 208 L/m 2 per h.Subsequently, the machine was turned off, and the acid was retained inside the system.Before using the system again, the acid solution was flushed out with soft water until the pH reached 8.3 to 8.5.The flux was measured again after flushing the acid solution.Within each MF stage, membrane was flushed using water, and flux was measured.The membrane was cleaned within stages using the abovementioned procedures when the flux did not show the original flux.

Drying of MCC
A pilot-scale spray dryer at Davis Dairy Plant at South Dakota State University was used to dry the MCC.The nozzle used to dry the MCC had a core size of 21 and an orifice size of 66.The inlet pressure was set at 2,250.0 psi using a high-pressure pump speed, and it was adjusted manually through the fan (30.0%).The supply fan and exhaust fan were set at 80.0 and 90.0%, respectively.The inlet temperature was 175°C, whereas the outlet temperature was 82°C.The dryer was connected to a fluid bed (Dahmes Stainless INC: DSI, Model no 10011-11) that was equipped with a sieve.The fluid bed was attached to 3 fans (hot = 71°C and 40.0%speed; warm = 50°C and 50.0%speed; cool = 21°C and 40.0%speed).Approximately 50.0 kg of the liquid MCC was heated to 50°C in a water bath before feeding into the dryer.The powder was collected in an airtight container and stored at room temperature until further analysis.The weight of the final powder was about 7.0 kg.

Manufacture of cMCC
The rest of the liquid MCC (75.0 kg) was fortified with lactose by adding 4.7% milk permeate powder (IdaPro milk permeate, Idaho Milk Products) to add approximately extra 3% lactose to ensure there was enough lactose for fermentation using starter cultures.The milk permeate was hydrated in the MCC and then the mix was pasteurized at 65°C for 30 min and cooled to 43°C to be ready for fermentation.The fortified MCC was then inoculated with 0.5% starter cultures (i455, Batch No 3489654, Chr.Hansen) at 43°C to get the pH of 4.6 in 12 to 14 h.A small amount (100 mL) of lactic acid (88% Lactic Acid FCC, product code: 175820, lot number: 1501277028) was added before cutting if the pH did not drop to the desired pH (4.6).Once the pH of 4.6 was reached, the curd was completely cut and left to heal for 30 min with gentle mixing, and then it was heated gradually to 50°C in 1 h.The whey was subsequently drained from the curd and sampled.The water at 50°C was used to wash the curd for 5 min at a ratio of 1:1 and then the water was drained.When the curd was washed with water, the pH of the curd increased slightly due to the buffering capacity.As a result, approximately 50 mL of lactic acid was added to the water (~100 lb) before washing the curd to keep the pH at 4.6.The washing step was repeated 3 times.The water was drained, and the curd was pressed for 6 h at 80 psi.The pressed curd was kept at ≤4°C until the next day.

Drying of cMCC
After pressing, the curd was milled into small pieces using a grinder (Humboldt Model 5DPJ3, split phase motor from Dayton) equipped with a 4.5-mm diameter stainless sieve.
A pilot-scale spray dryer at Davis Dairy Plant at South Dakota State University was used to dry the cMCC.The dryer was not turned on only the exhaust fan and 3 fans connected to the fluid bed were running manually.The exhaust fan was set at 20%.The 3 fans attached to the fluid bed were set as follows: hot air fan = 71.0°Cand 40.0%speed, warm air fan = 71.0°Cand 50.0%speed, and cool air fan = 21.0°C and 15.0% speed.
The curd was then placed on the sieve of the fluid bed, which is part of the spray dryer.The curd was evenly distributed on the surface of the fluid bed.The temperature of fans was elevated or decreased by controlling the speed of fans manually.The curd was moved and mixed every 30 min upside down.The curd was left in the fluid bed for around 4 h to ensure the curd was completely dried.The dried curd was collected in an airtight container and stored at room temperature until grinding.The weight of the dried curd was approximately 14.0 kg.The dried curd was then turned into powder using a high-speed multifunctional grinder (item no.HC-700Y).The powder was then separated using a metal sieve (item no.8RTG5, Sieve #70, Grainger) with a 212-µm pore size to have the typical particle size of powders.Powder particles with over 212 µm were collected and milled again in the Retsch Ultracentrifugal mill (Brinkmann Retsch ZM1) equipped with a 0.2-mm stainless steel ring sieve to have a fine powder.The cMCC powder was collected in bags and stored at room temperature until further analysis.

Manufacture of PCP
Techwizard (Excel-based-formulation software program provided by Owl Software) was used to develop PCP formulations (Metzger, 2010) with different ratios of cMCC and MCC (2.0:1.0 formulation = FR-2.0:1.0,1.9:1.1 formulation = FR-1.9:1.1, and 1.8:1.2formulation = FR-1.8:1.2) to produce PCP with 45% moisture, 30% fat, 19% protein, and 2.4% salt.The percentage of ingredients used in PCP formulations is shown in Table 1.In each formulation, the amount of protein from cMCC and MCC was adjusted based on the formulation.As a result, the amount of cMCC was decreased, whereas the amount of MCC was increased from 2.0:1.0 to 1.8:1.2formulations.The ingredients included aged natural cheddar cheese (Kraft Heinz Cheddar), salted butter (Land O'Lakes Inc.), cMCC, MCC, milk permeate (product lot: 19113D40, Idaho Milk Products), and salt (Morton Salt Inc.).Approximately 15% of aged cheddar was added into PCP formulations to get mild cheddar flavor in the final PCP.Milk permeate powder was used to standardize the solids content in all formulations.Process cheese products formulations were prepared by mixing cMCC powder, MCC powder, and water for 10 min in a kitchenaid at room temperature.Then other ingredients were added and mixed for 30 min to produce a homogeneous paste.A 20-g sample of the paste was weighed in a canister, warmed in a 40°C water bath for 30 min, and then cooked in the rapid visco analyzer (RVA; Perten RVA 4500) for 3 min at 95°C.We added 0.5 g of water into each canister to compensate for the water that evaporated during mixing and cooking in the RVA.The stirring speed was 1,000 rpm for the first 2 min and 160 rpm for the last min.Each batch was divided into 10 canisters.The cooked PCP was then poured into copper molds (diameter = 20 mm; height = 30 mm) to measure the hardness by using texture profile analysis.The cooked PCP was poured into plastic molds (diameter = 28.3mm; height = 25 mm), which was used to measure the melt temperature by using dynamic stress rheometry (DSR) and melt diameter by using the Schreiber melt test.This experiment was repeated 3 times using different batches of cMCC and MCC powders.

Functional Analyses
End Apparent Viscosity.The end apparent viscosity of the PCP was set to be measured by the end of the cooking time in the RVA at 95°C after 3 min by calculating the mean of the last 5 apparent viscosity readings from the RVA curve.However, due to the high viscosity of the treatments in this study, the RVA automatically stopped after 1 min of cooking, so this cooking time was standardized in all treatments and the last point referred to the end apparent viscosity.The end apparent viscosity was measured in all canisters of the batch.
Texture Profile Analysis.The hardness of the PCP was measured by using the texture profile analysis.The PCP was prepared by pulling the cheese out from the copper cylinders and then cutting the cheese into 200-mm height.The PCP was analyzed for hardness using a TA.XT-Plus Texture Analyzer (TA.XT-Plus, 6 Patton Drive) equipped with a 38-mm diameter cylindrical flat probe (TA-4) and using uniaxial double bite 10% compression with 1 mm/s crosshead speed.The maximum force of the first compression was referred to as the hardness of the cheese.This test was repeated 6 times for each batch.
Dynamic Stress Rheometry.The PCP was prepared by cutting the cheese into slices (2 mm thick and a 28.3-mm diameter) using a wire cutter (Hammam et al., 2022b).A stress sweep test of the PCP was performed at a frequency of 1.5 Hz, and stress ranged from 1 to 1,000 Pa at 20°C using a rheometer with parallel plate geometry (MSR 92, Anton Paar).The stress sweep experiment determined that the maximum stress limit for the linear viscoelastic region was 500 Pa.The dynamic rheological properties of the PCP were then analyzed with a dynamic temperature ramp test that ranged from 20 to 90°C with a ramp rate of 1°C/ min using a frequency of 1.5 Hz and constant stress of 500 Pa.The temperature at which tan δ = 1 [loss modulus (G′′)/storage modulus (G′)] was referred to as the cheese melting temperature.A duplicate was performed on each batch.
Schreiber Melt Test.The PCP samples were cut into cylinders (diameter = 28.3mm and height = 7 mm) and placed on 0.75 mm thick aluminum plates (100 mm × 90 mm).The plates were transferred to a forced draft oven at 90°C for 7 min (Hammam et al., 2023).After cooling the plates, the diameter of the melted PCP samples was measured in 4 different places using a vernier caliper and reported in millimeters.This test was repeated 4 times for each batch.

Statistical Analysis
Statistical analysis was performed to study the effect of the ratio of protein from MCC and cMCC on the functional properties of PCP.Each replicate was considered as a block for the design, which was manufactured on a different day.Analysis of variance was done to obtain the mean squares and P-values using the GLM procedure available in R software (R x64 3.3.3using R studio).Differences were tested using the least significant difference comparison test when a significant difference was detected at P < 0.05.

Microfiltration to Produce MCC
Composition of Skim Milk.The mean composition of skim milk used in this study is shown in Table 2.The skim milk showed an average of 3.37, 0.29, 0.87, 0.58, 9.01, 4.41, 0.20, and 0.21% for TPr, NPN, NCN, SP, TS, lactose, lactic acid, and fat, respectively.The TP in skim milk was 3.08% with 2.50% casein.The casein as a percentage of TP was approximately 81.08%, whereas the ash content in skim milk was 0.72% including 0.12% Ca and 0.09% P. No noticeable differences were detected in the composition of skim milk within the 3 replicates.The mean composition of skim milk did not differ from the skim milk reported in other MF studies (Beckman et al., 2010;Hurt and Barbano, 2010;Adams and Barbano, 2013;Hammam et al., 2022a).The high pasteurization temperature can result in denaturation of β-LG on κ-CN through disulfide bonding (Singh, 1995), which can increase the percent of casein to TP (CN%TP) and thereby decrease the SP removal during MF (Hurt and Barbano, 2010).The CN%TP in this study was typically similar to other studies (Hurt and Barbano, 2010;Adams and Barbano, 2013;Beckman and Barbano, 2013) because the pasteurization temperature did not exceed the recommended value.The extensive pasteurization can be noticed in the NCN values, which is increasing with higher temperature be- cause SP is denaturated on casein.It has been found that the Ca and P in milk were around 0.11 and 0.09%, respectively (Wong et al., 1976), and those values are similar to the Ca and P values reported in Table 2.
Composition of Permeate.The composite permeate composition collected from the 3 stages of MF is shown in Table 2.The composite permeate had 0.44, 0.16, 0.43, 0.26, 4.01, 3.11, and 0.17% for TPr, NPN, NCN, SP, TS, lactose, and lactic acid, respectively.We found 0.01% casein in the permeate, whereas TP was 0.28%, therefore the CN%TP was 4.28%.The ash content in the permeate was 0.32% with 0.01% Ca and 0.02% P. The composition of permeate was similar within replicates.We have produced MCC previously using the same procedures to have >25% TS and >20% protein (Hammam et al., 2022a).The composite permeate of the first and second stages in our previous study can be used for comparison reason to this study because we did not go further to produce highly concentrated MCC.No differences were detected in the composition of permeate of our previous study relative to the current study.It showed 4.01% TS, 0.43% TPr, 0.14% NPN, 0.29% SP, and 0.32% ash, which are similar to the current study.The composition of permeate in this study was also similar to the permeate reported in other studies (Hurt and Barbano, 2010;Hurt et al., 2010;Zulewska and Barbano, 2014;Tremblay-Marchand et al., 2016).The slight differences in the composition of permeate within studies can be due to the differences in membrane type, DF steps, as well as the composition of skim milk.
Composition of MCC.The composition of MCC made from MF of skim milk is illustrated in Table 2.The average composition of MCC made from the 3 replicates was 11.15% TPr, 0.16% NPN, 1.26% NCN, 1.10% SP, 14.06% TS, 1.32% lactose, and 0.18% lactic acid.The ash content was 1.18% with 0.34% Ca and 0.20% P. The CN%TP was approximately 90% with an average of 9.89% casein and 10.99% TP.We previously produced MCC with similar composition using the GP MF ceramic membranes following the same procedures (Hammam, 2022).We found that the TS, TPr, and ash were 13.79, 9.54, and 1.03%, respectively.Similar results were also found by Tremblay-Marchand et al. (2016).The latter study reported that the retentate of 3× MF GP membranes had 89.59%CN%TP with 0.92% SP and 15.31% TS.However, the CN%TP of MCC produced in other studies was high relative to our study due to the extra DF stage that those studies used between the second and third stages (Hurt and Barbano, 2010;Hurt et al., 2010;Zulewska and Barbano, 2014).This led to increasing the CN%TP in MCC due to the high SP amount that was removed during MF.

Culture-Based Acid Curd
Composition of Fortified MCC.The composition of MCC used in making cMCC is exemplified in Table 2.The same MCC product produced from MF was used in making cMCC after fortification with milk permeate powder.The TPr, NPN, NCN, SP, TS, lactose, and lactic acid in fortified MCC were 10. 95, 0.25, 1.45, 1.20, 17.37, 4.54, and 0.24%, respectively.The ash content was 1.39% including 0.35% Ca and 0.23% P. The CN%TP was 88.78% with 9.50 and 10.70% casein and TP, respectively.No differences were detected between the MCC solutions before and after fortification with milk permeate powder except for TS and lactose.It was expected that lactose will be elevated by >3% due to the addition of milk permeate powder as a source of lactose to ensure there is enough lactose for fermentation using starter cultures.Additionally, the TS increased by around 3.3% for the same reason.
Composition of Acid Whey.The composition of acid whey produced as a byproduct of making cMCC from MCC is presented in Table 2.The whey showed approximately 3.82, 0.37, 1.52, 1.14, 10.61, 2.49, and 2.30% for TPr, NPN, NCN, SP, TS, lactose, and lactic acid, respectively.The whey showed around 1.37, 0.33, and 0.16% ash, Ca, and P. The TPr presented in whey includes 3.44% TP and 2.30% casein to show 65.55% CN%TP.We previously produced cMCC from MCC (13.02% TS, 1.96% lactose, 1.01%NCN, and 9.18% TPr) using starter cultures at a laboratory scale (Hammam, 2022).The composition of acid whey produced as a byproduct of making cMCC at a laboratory scale did not differ much from that produced on the pilot scale.The percentage of TPr to TS in acid whey produced on a laboratory scale was 28.1%, which is close to the value in this study (36.0%).Approximately 1.4 to 1.8% lactose was required to reach a pH of 4.6 in the acid curd/whey.As a result, around 2.5% lactose from a total of 4.23% was left in the whey.Using 2.0% lactose MCC left around 0.6% of lactose in the laboratory scale acid whey (Hammam, 2022).The loss of components in whey especially protein depends on the composition of initial material as well as handling the curd in the cheese vat and this loss gets higher with increasing the scale.The composition of acid whey produced as a byproduct of making cMCC was different from the acid whey produced from milk in previous studies (Peri and Dunkley, 1971;Durham and Hourigan, 2007;Saffari and Langrish, 2014;Chandrapala et al., 2015;Lievore et al., 2015;Chen et al., 2016;Hammam, 2022) due to the differences in the composition of starting material (MCC), which was expected.As a result, TS, ash, TPr, lactose, and lactic acid contents in acid whey can be changed based on the composition of MCC.Those studies found that the TS of acid whey can range from 5.0 to 7.0%, whereas TPr and ash can be 0.5 to 1.0 and 0.5 to 1.0%, respectively.The TS, TPr, and ash in acid whey produced from MCC were high because they were high in MCC compared with acid whey produced from using milk in curd manufacture.
Composition of Soft Acid Curd.The composition of soft cMCC produced from MCC is shown in Table 2.The TPr, NPN, NCN, SP, TS, lactose, and lactic acid content in produced cMCC was 36.45, 0.74, 1.33, 0.58, 40.37, 0.80, and 1.34%, respectively.The ash content was 0.96% including 0.11% Ca and 0.18% P. The casein in cMCC was 35.13%, whereas TP was 35.71% to present 98.36% CN%TP.The average pH of cMCC was targeted to have 4.60.The composition of cMCC depends on the composition of initial materials, final pH, and process conditions (e.g., cooking temperature, washing curds, pressing) (Wong et al., 1976).The step of washing the curd has a significant role in increasing the ratio of casein to TP and decreasing the ash, Ca, P, lactose, and lactic acid content in the final cMCC.Because the lactose is converted to lactic acid using starter cultures as a result of fermentation, lactose decreases and lactic acid increases as the pH reaches 4.6.It is also expected that the ash content decreases when the MCC is turned into cMCC (McDowall and Dolby, 1935;Czulak et al., 1969;Wong et al., 1976;Hill et al., 1985;Lucey and Fox, 1993).The Ca is converted from insoluble (colloidal form) to soluble form and released in the whey as the pH decreases (Davies and White, 1960;Wong et al., 1976;Dalgleish and Law, 1989;Guinee et al., 1993).As a result, the Ca and P drop as the pH decreases with having a low ratio of Ca to P (Kindstedt and Kosikowski, 1988;Lucey and Fox, 1993).Wong et al. (1976) have reported that the Ca in cottage curd is reduced by 68.2% (washed 3 times) relative to the Ca content in skim milk, which is similar to our results (68.6%).Another review reported that the Ca and P in cottage cheese were 0.08 and 0.16%, respectively (Lucey and Fox, 1993).

Composition of MCC powder, cMCC powder, and PCP
Composition of MCC Powder.The liquid MCC was spray dried to produce MCC powder.Table 2 presents the composition of MCC powder.The average composition of MCC powder showed 75.77%TPr, 2.07% NPN, 9.66% NCN, 7.60% SP, 97.84% TS, 8.69% lactose, 1.73% lactic acid, and 2.98% fat.The ash content was 8.03% with 2.42% Ca and 1.29% P. The CN%TP was approximately 89.70% with an average  (Hammam et al., 2021).The MCC 80 has approximately 80.0% TPr, 95.0%TS, 4.5% lactose, 3.0% fat, and 8.0% ash.Similar results were reported in several studies (Nasser et al., 2017(Nasser et al., , 2018;;Carter et al., 2018).Those studies found that TPr, TS, lactose, and ash ranged from 80.0 to 83.0, 94.0 to 95.0, 1.0 to 2.5, and 7.0 to 8.0%, respectively.The lactose content in the commercial MCC and other studies is low due to the multiple DF stages that are used to deplete more lactose and low molecular weight components in the permeate during MF.The slight differences can be related to the composition of initial material, MF process conditions (e.g., temperatures, pressures, CF, DF), and types of membrane (e.g., ceramic membranes, spiral-wound membranes) (Hammam et al., 2021).
The composition of cMCC powder was relative to the composition of soft cMCC on a dry basis and similar to this range reported in previous studies (Huffman and James Harper, 1999;Hammam et al., 2021).The acid curd made from milk could have less ash content as well as Ca and P due to the low content of those components in milk as compared with MCC, which has higher ash, Ca, and P contents.The slight differences in the composition of cMCC are due to the differences in the composition of the initial material and process used to make the curd.
Composition of PCP.The composition of PCP made from cMCC and MCC is shown in Table 3.The mean squares and the P-values for the composition of PCP are indicated in Table 4.The moisture content was 44.19, 44.21, and 44.15% in PCP made from FR-2.0:1.0,FR-1.9:1.1, and FR-1.8:1.2, respectively.The ash content of PCP made from the 2.0:1.0 ratio of protein from cMCC to MCC was 3.28%, whereas it was 3.42 and 3.40% in the PCP made from 1.9:1.1 and 1.8:1.2,respectively.No differences (P > 0.05) were detected in the moisture and ash contents of PCP because all formulations were standardized before cooking in the RVA to have the same composition.The PCP exhibited 0.31, 0.33, and 0.41% Ca in formulations of FR-2.0:1.0,FR-1.9:1.1, and FR-1.8:1.2, respectively.Although a slight increase in the Ca content was noticed with decreasing the cMCC and increasing the MCC, no significant difference (P > 0.05) was detected in the Ca content.It was reported that the Ca of PC made using ES and natural cheeses ranged from 0.3 to 0.4% (Kapoor et al., 2007), which is similar to our values.However, increasing the amount of MCC led to differences (P < 0.05) in the pH of final PCP, which was 5.25 in 2.0:1.0 formulation, 5.32 in 1.9:1.1 formulation, and 5.37 in 1.8:1.2formulation.This was expected as the MCC had higher pH when compared with cMCC.The ratio of cMCC to MCC has an effect on the level of de-aggregation and the pH of the final PCP.It was indicated that the Ca and P, residual lactose, and salt-to-moisture ratio in natural cheeses affect the pH of PC (Kapoor et al., 2007).It has been reported that the typical pH of PC Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.
The pH of PCP in this study was close to the typical values.However, it has been found that pH of PC can range from 5.1 to 5.2 and still shows good functional properties (Kommineni et al., 2012).We previously have found that the pH of PCP or imitation mozzarella cheese produced with no ES using cMCC and MCC ranged from 5.3 to 5.5 (Hammam, 2022;Hammam et al., 2023), which is similar to this study.The differences in pH could affect the structure and quality of final PCP and thereby its functional properties due to its effects on the protein interactions (Palmer and Sly, 1943;Meyer, 1973;Marchesseau et al., 1997).Palmer and Sly (1943) stated that the emulsion of PC is low when the pH is lower than 5.4 or higher than 5.8.It was found that as the pH of PC drops to 5.2, the protein-protein interaction increases (Marchesseau et al., 1997) because the pH is close to the isoelectric point of caseins (4.6).This induces the aggregation of the protein, which in turn, results in a poor emulsion of fat in PCP.On the other hand, the PC had an open structure when the pH elevated to 6.1, which eventually led to weaker emulsification (Marchesseau et al., 1997).Marchesseau et al. (1997) also found that as the pH increased up to 5.7 it resulted in PC with better uniform fat emulsion with a closely knit protein network.

Functional Characteristics of PCP
End Apparent Viscosity of PCP.The end apparent viscosity of PCP made from cMCC and MCC is illustrated in Table 5.The mean squares and the P-values for the end apparent viscosity and hardness of PCP are presented in Table 6.The end apparent viscosity of PCP made from FR-2.0:1.0,FR-1.9:1.1, and FR-1.8:1.2 was approximately 4,305.0,2,408.8, and 2,499.3cP, respectively.The end apparent viscosity was different (P < 0.05) within treatments showing the highest viscosity in the FR-2.0:1.0PCP.The highest viscosity in FR-2.0:1.0 could be due to the differences in pH.The viscosity of PC increased as the pH decreased.As the pH dropped to the isoelectric point (4.6), the protein-protein interactions increased which led to high viscosity (Kapoor, 2007).
Hardness of PCP.The hardness of PCP made from cMCC and MCC is shown in Table 5.The mean squares and the P-values for the hardness of PCP are exemplified in Table 6.The hardness was approximately 424.0 g for the 2.0:1.0 ratio, 512.0 g for the 1.9:1.1 ratio, and 407.0 g for the 1.8:1.2ratio.The hardness of PCP did not show any significant differences (P > 0.05) between different formulations made from different ratios of protein from cMCC and MCC.Although the hardness should be correlated with viscosity (Kapoor and Metzger, 2008) and pH (Kapoor, 2007), we did not find differences in the hardness of the PCP made from different ratios of protein from cMCC and MCC.The hardness of our PCP looks better compared Means in the same column not sharing a common superscript are different (P < 0.05).
1 Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.Means in the same column not sharing a common superscript are different (P < 0.05).
with those PCP made using ES.In our previous study, we found that the hardness of PCP made using MCC and ES ranged from 100.0 to 212.0 g (Hammam et al., 2022b).Another study reported that the hardness of PCP made from MCC using ES was 110.0 to 135.0 g (Salunke, 2013).
Melting Characteristics of PCP.The melting characteristics of PCP made from cMCC and MCC are shown in Table 7.The mean squares and the P-values for the melting characteristics of PCP are illustrated in Table 8.We detected a significant difference (P < 0.05) in the melting temperature of PCP made from 2.0:1.0,1.9:1.1, and 1.8:1.2formulations.The highest melting temperature was 54.0°C in the 2.0:1.0 ratio, whereas it was 43.0 and 42.0°C in the 1.9:1.1 and 1.8:1.2ratios, respectively.It was expected that 2.0:1.0 formulation will have the highest melting temperature because the PCP showed higher end apparent viscosity than other formulations.We found a correlation between viscosity and melt temperature.It was mentioned that the melt temperature elevated as the end apparent viscosity increased (Kapoor and Metzger, 2008), which our work pointed out in Table 5.Additionally, the melting of PC increased as the pH decreased.As the pH decreased to the isoelectric point (4.6), the protein-protein interactions increased, which required more temperature to be melted (Kapoor, 2007).
It has been reported that the melt diameter or melt area should increase as the melt temperature decreases (Kapoor and Metzger, 2008).Although we detected a significant difference in the melting temperature of PCP, no differences (P > 0.05) were detected in the melt diameter or area.We found an increase in the melt diameter of PCP, but this increase was not significant.The melt diameter and melt area increased with elevating the protein ratio from MCC compared with cMCC.The melt diameter was 38.8, 43.9, and 42.4 mm for PCP made with 2.0:1.0,1.9:1.1, and 1.8:1.2 of protein from cMCC to MCC, respectively.The melt area was approximately 1,184.0,1,539.0,and 1,415.0mm 2 for PCP made with 2.0:1.0,1.9:1.1, and 1.8:1.2 of protein from cMCC to MCC, respectively.The melting temperature is an indicator of the melt diameter.As a result, a similar trend was found in the melt diameter compared with the melt temperature.Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.Means in the same row not sharing a common superscript are different (P < 0.05). 1 Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.Means in the same row not sharing a common superscript are different (P < 0.05). 1 Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC.
The differences in the onset of melting can be explained by the differences in pH of those cheeses.As the pH drops to the isoelectric point, the net negative charges on caseins reduce which increase the proteinprotein interactions, and this leads to aggregation of protein and thereby low meltability.This led to a higher melting temperature with less melted diameter and area.
The Rheological Properties of PCP.The G′ (elastic) and G′′ (viscous) moduli of PCP measured during heating from 20 to 90°C at 10°C increments are shown in Tables 9 and 10, respectively.The ANOVA table with mean squares and P-values at 20, 70, and 90°C for both elastic (G′: Pa) and viscous (G″: Pa) moduli of the PCP made from cMCC and MCC are shown in Table 11.The effect of using different ratios of protein cMCC and MCC (2.0:1.0,1.9:1.1, and 1.8:1.2) in manufacture of PCP with no ES on G′ and G″ during heating from 20 to 90°C is presented in Figures 4  and 5, respectively.
The G′ values of PCP made using different ratios of protein from cMCC and MCC (2.0:1.0,1.9:1.1, and 1.8:1.2) were not different (P > 0.05) at a temperature range of 20 to 30°C (Figure 4).However, we found differences (P < 0.05) in G′ values of PCP made using different ratios of protein from cMCC and MCC from 1 Treatment: FR-2.0:1.0 = PCP formulation made with 2.0:1.0 ratio of protein from cMCC to MCC; FR-1.9:1.1 = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC. 1.0 ratio of protein from cultured micellar casein concentrate (cMCC) to micellar casein concentrate (MCC); FR-1.9:1.1 (▼) = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 (■) = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC during heating from 20 to 90°C using dynamic rheological analysis.40 to 80°C (Table 9).These differences in G′ values of PCP made using different ratios of protein from cMCC and MCC were not found (Table 11) at 90°C (Table 9).Process cheese product made from 2.0:1.0 (protein from cMCC to MCC) gave the highest G′ values at temperatures of 40 to 80°C, followed by 1.9:1.1 and 1.8:1.2formulations.
The G′′ of PCP made using different ratios of protein from cMCC and MCC (2.0:1.0,1.9:1.1, and 1.8:1.2) followed the same trend of G′ at a temperature range of 20 to 90°C (Figure 5).The G′′ values of PCP made using different ratios of protein from cMCC and MCC (2.0:1.0,1.9:1.1, and 1.8:1.2) were not different (P > 0.05) at a temperature range of 20 to 30°C (Table 10).However, we detected differences (P < 0.05) in G′′ values made using different ratios of protein from cMCC and MCC from 40 to 80°C (Table 10).These differences in G′′ values of PCP made using different ratios of protein from cMCC and MCC were not found at 90°C (Table 11).Process cheese product made from 2.0:1.0 (protein from cMCC to MCC) gave the highest G′ values at temperatures of 40 to 80°C, followed by 1.9:1.1 and 1.8:1.2formulations.
The G′ of PCP made using different ratios of protein from cMCC and MCC (2.0:1.0,1.9:1.1, and 1.8:1.2) before melting was higher than G′′.This indicates that the PCP has elastic behavior (gel) more than the viscous behavior (liquid).The G′ and G′′ are decreased during measuring the melting using the DSR.Both moduli are decreased until the cross point, which is the cheese melting point.The same trend was found in other studies (Hennelly et al., 2005;Subramanian et al., 2006;Zhong et al., 2007;Guinee and O'Kennedy, 2009;Kommineni et al., 2012;Hosseini-Parvar et al., 2015).The improvement in the functional characteristics of PCP with no ES made using different ratios of protein from cMCC and MCC (2.0:1.0,1.9:1.1, and 1.8:1.2) may be a result of a better emulsion of cMCC and MCC during making the cheese.

CONCLUSIONS
The GP MF membranes can be effectively used to produce MCC with >9% TP and >13% TS using 3 stages MF with a 3× CF and DF.Additionally, cMCC was produced from the liquid MCC.A unique method was developed to produce cMCC powder by drying and grinding the curd.Different ratios of protein from cMCC powder to MCC can be used in making PCP with no ES.The cMCC and MCC protein of different 1.0 ratio of protein from cultured micellar casein concentrate (cMCC) to micellar casein concentrate (MCC); FR-1.9:1.1 (▼) = PCP formulation made with 1.9:1.1 ratio of protein from cMCC to MCC; FR-1.8:1.2 (■) = PCP formulation made with 1.8:1.2ratio of protein from cMCC to MCC during heating from 20 to 90°C using dynamic rheological analysis.ratios create a partially de-aggregated casein network that results in a PC with functionality similar to PC produced with ES.We also found that PCP made from the 2.0:1.0 formulation showed better functional properties as compared with other formulations.The novel ingredients (cMCC and MCC powders) produced in this study can be mixed in a specific ratio and market to be ready for the manufacture of clean label imitation mozzarella cheese and PCP with no ES.

Figure 1 .
Figure 1.Emulsifying salts interaction during making process cheese or process cheese products.

Figure 2 .
Figure 2. Cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC) interaction in making clean label process cheese products.
Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE

Table 1 .
, total proteinHammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE Mean (n = 3) composition of process cheese products (PCP) formulations made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC) Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE

Table 2 .
Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE Mean (n = 3) composition (% by weight) of ingredients used in manufacture of micellar casein concentrate (MCC) and cultured micellar casein concentrate (cMCC) Ingredient Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE of 66.10% casein and 73.70% TP.The composition of MCC powder is relatively similar to the liquid MCC on a dry basis.The composition of MCC powder in this study is similar to the composition of commercial MCC 80 reported previously

Table 3 .
Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE Mean (n = 3) composition of process cheese products (PCP) made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC) 1

Table 4 .
Mean squares and P-values (in parentheses)for the composition of the process cheese products (PCP)

Table 5 .
Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE Mean values (n = 3) of end apparent viscosity (cP) and hardness (g) of the process cheese products (PCP) made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC)

Table 6 .
Mean squares and P-values (in parentheses)for the end apparent viscosity (cP) and hardness (g) of the process cheese products (PCP) made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC)

Table 8 .
Hammam et al.: NOVEL CULTURED MICELLAR CASEIN CONCENTRATE Mean squares and P-values (in parentheses) for the melting properties of the process cheese products (PCP) made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC)

Table 10 .
Mean viscous modulus (G′′: Pa) of process cheese products (PCP) made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC) during heating from 20 to 90°C using dynamic rheological analysis (DSR)

Table 11 .
Hammam et al.:NOVEL CULTURED MICELLAR CASEIN CONCENTRATE Mean squares and P-values (in parentheses) for elastic modulus (G′: Pa) and viscous modulus (G′′: Pa) of process cheese products (PCP) made with different ratios of cultured micellar casein concentrate (cMCC) and micellar casein concentrate (MCC) during heating from 20 to 90°C using dynamic rheological analysis (DSR)