The effect of passion fruit juice on quality of mozzarella cheese

Thesis assignment: Effect of passion fruit juice on qualities of mozzarella includes the following contents: + Analysis of the chemical composition of materials ash, moisture, protein, l

MINISTRY OF EDUCATION AND TRAINING

HO CHI MINH CITY UNIVERSITY OF TECHNOLOGY AND EDUCATION

FACULTY FOR HIGH QUALITY TRAINING

Ho Chi Minh City, August, 2022

ADVISOR: PHAM THI HOAN STUDENT: TRAN THI THUY DUONG NGUYEN THI MINH THU

SKL 0 0 8 9 3 8

GRADUATION THESIS FOOD TECHNOLOGY

THE EFFECT OF PASSION FRUIT JUICE ON

QUALITY OF MOZZARELLA CHEESE

THE EFFECT OF PASSION FRUIT JUICE ON QUALITY

OF MOZZARELLA CHEESE

Major: FOOD TECHNOLOGY

HO CHI MINH CITY – AUGUST 2022

THE EFFECT OF PASSION FRUIT JUICE ON QUALITY

OF MOZZARELLA CHEESE

Major: FOOD TECHNOLOGY

HO CHI MINH CITY – AUGUST 2022

68

GRADUATION THESIS ASSIGNMENT

Student name: Trần Thị Thùy Dương Student ID: 18116008

Student name: Nguyễn Thị Minh Thư Student ID: 18116038

Supervisor: Phạm Thị Hoàn, Ph.D Email: hoanpt@hcmute.edu.vn

Date of assignment: 14/02/2022 Date of submission: 08/08/2022

1 Thesis title:

Effect of Passion fruit juice on quality of Mozzarella cheese

2 Thesis assignment:

Effect of passion fruit juice on qualities of mozzarella includes the following contents:

+ Analysis of the chemical composition of materials (ash, moisture, protein, lipid, pH, titrable acidity and microbiological indicators, fruit weight and diameter)

+ Investigate the influence of passion fruit juice volume on properties of passion fruit mozzarella cheese (%H, %TSwhey, %TScheese, CSY (soild cheese yield), colorspace measurement and texture analysis)

+ Investigate the influence of coagulation temperature on passion fruit mozzarella qualities

+ Investigate the influence of rennet concentration on the qualities of mozzarella cheese qualities + Evaluate the qualities of passion fruit mozzarella (Chemical composition: total dry matter content, proteins, lipids, acidity, pH, microstructure of products, sensory test, microbiological criteria)

+ Evaluate the qualities of passion fruit mozzarella during 21 days of storage

The content and requirements of the graduation thesis have been approved by the Chair of the Food

Technology program

CHAIR OF THE PROGRAM

(Sign with full name)

Ho Chi Minh City, August 9, 2022

SUPERVISOR

(Sign with full name)

THE SOCIALIST REPUBLIC OF VIETNAM

Independence – Freedom– Happiness

-

69

ACKNOWLEDGEMENTS

Thanks to the devotion of the teachers and the knowledge accumulated after more than three years sitting on the lecture chair of Ho Chi Minh City University of Technical Education Besides my efforts to learn and research, I also receive a lot of support and encouragement fromthe school, teachers, friends and family First of all, I would like to express my sincere thanks to Ph.D, Phạm Thị Hoàn for teaching, guiding and helping us throughout the project implementation process

We would like to thank the teachers and students of the Department of Chemical and Food Technology, Ho Chi Minh City University of Technical Education for their dedication, teaching and building of the basic knowledge needed in during the four years of study and practice at the school, create the premises and conditions for the successful implementation of this graduation thesis Lastly, we would like to sincerely thank all those who have helped and assisted us in completing this project in this food technology

Although we have tried a lot to complete the project, with limited time and ability, this is the first time we have entered into researching, calculating and perfecting a technology product,

so it is unavoidable errors We look forward to receiving the contributions of teachers

Sincerely, Trần Thị Thùy Dương & Nguyễn Thị Minh Thư

STATEMENT OF ORGINALITY

I hereby declare that the Graduation Thesis “Effect of Passion fruit juice on quality of Mozzarella cheese is the results of my own research and has never been published in any work of others During the implementation process of this project, I have seriously taken research ethics; all findings of this projects are results of my own research and surveys; all references in this

project are clearly cited according to regulations

I bear full responsibility for the fidelity of the number and data and other contents of my graduation project

Date:

Student

Trần Thị Thùy Dương Nguyễn Thị Minh Thư

TABLE OF CONTENTS

ACKNOWLEDGEMENTS i

STATEMENT OF ORGINALITY ii

ASSESSEMENT FORM GRADUATION THESIS OF FOOD TECHNOLOGY(FOR COMMITTEE MEMBER) iii

ASSESSEMENT FORM GRADUATION THESIS OF FOOD TECHNOLOGY(FOR REVIEWER) ix

ASSESSEMENT FORM GRADUATION THESIS OF FOOD TECHNOLOGY(SUPERVISOR) ix

TABLE OF CONTENTS xv

LIST OF FIGURES xviii

LIST OF TABLES xix

LIST OF ABBREVIATIONS xx

ABSTRACT xxi

Chapter 1: INTRODUCTION 1

1.1 Pose the problem 1

1.2 Topic goal 2

1.3 Subject and scope of research 2

1.4 Research content 2

1.5 The scientific and practical values of the topic 3

Chapter 2: LITERATURE REVIEW 4

2.1 Cheese 4

2.1.1 Overview of cheese 4

2.1.2 Classification 5

2.2 Mozzarella Cheese 6

2.2.1 Classification 7

2.2.2 Casein coagulation 8

2.3 Raw material 11

2.3.1 Raw milk 12

2.3.2 Passion fruit 14

2.5 Research situations of the topic 17

2.5.1 Domestic situation 17

2.5.2 Overseas situation 18

2.6 Reasons for choosing the topic 18

Chapter 3: MATERIALS AND RESEARCH METHODS 20

3.1 Locations, chemicals, laboratory equipment 20

3.1.1 Locations 20

3.1.2 Chemicals 20

3.1.3 Laboratory tools and equipments 20

3.2 Materials 20

3.2.1 Fresh cow's milk 20

3.2.2 Passion fruit 21

3.2.3 Citric acid 21

3.2.4 Rennet 21

3.3 Research schema 23

3.4 Manufacturing of passion fruit mozzarella 24

3.5 Experimental arrangements 28

3.5.1 Experiment 1: The effect of the passion fruit juice volume on the properties mozzarella 28

3.5.2 Experiment 2: The effect of temperature in coagulation process on the properties of mozzarella 28

3.5.3 Experiment 3: The effect of rennet concentration on the propertiesof mozzarella 29 3.5.4 Evaluation of the qualities of passion fruit mozzarella during storage period 30

3.6 Research methods 30

3.6.1 Method for determining chemical composition 30

3.6.2 Determining the yield of cheese efficiency 30

3.6.3 Determination of cheese solid yield (CSY) 30

3.6.4 Determining titratable acidity 31

3.6.5 Colorspace measurement 31

3.6.6 Microstructure analysis using scanning electron microscope (SEM) 32

3.6.7 Texture profile analysis (TPA) 32

3.6.8 Sensory evaluation 33

3.6.9 Microbiological analysis 34

3.6.10 Data analysis 34

CHAPTER 4: RESULTS AND DISCUSSION 35

4.1 The chemical composition and physicochemical properties of raw materials 35

4.1.1 Chemical composition of raw milk 35

4.1.2 Physicochemical properties of passion fruit juice 35

4.2 The effect of the passion fruit juice volume on the quality properties of mozzarella 36

4.2.1 Effect of passion fruit juice volume 36

4.2.2 Effect of passion fruit juice proportion on mozzarella texture properties 38

4.2.3 Effect of passion fruit proportion on color change of mozzarella 39 4.3 The effect of temperature in coagulation process on the properties of mozzarela 41 4.3.1 Effect of temperature in coagulation process 41 4.3.2 Effect of temperature in coagulation process on mozzarella texture properties 42 4.3.3 Effect of temperature in coagulation process on color change of mozzarella 43 4.4 The effect of rennet concentration on the quality of mozzarella 45 4.4.1 Effect of rennet concentration 45 4.4.2 Effect of rennet concentration on mozzarella texture properties 46 4.4.3 Effect of rennet concentration on color change of mozzarella 47 4.5 Conclusion on the production process of passion fruit mozzarella 48 4.6 Evaluation of the quality properties of passion fruit mozzarella 49 4.6.1 Chemical composition and microbiological criteria 49 4.6.2 Evaluation of the microstructure of mozzarella 51 4.6.3 Sensory evaluation of passsion fruit mozzarella 51 4.7.1 Color change of passion fruit mozzarella during storage 53 Chapter 5: CONCLUSION AND RECOMMENDATIONS 55 5.1 Conclusion 55 5.2 Recommendations 55

REFERENCES Error! Bookmark not defined

APPENDIX 59

LIST OF FIGURES

Figure 1 1 U.S Consumption of Cheese (1975-2018) 1Figure 2 1 Illustrations of cheese products 4Figure 2 2 Mozzarella cheese 7Figure 2 3 Purple passion fruit 15Figure 2 4 Yellow Passion Fruit 16Figure 2 5 Illustrate the mechanism of coagulation by rennet 10Figure 3 1 Nong Lam Milk 20 Figure 3 2 Passion fruit (Passiflora edulis) 21 Figure 3 3 Naturen Extra 220 rennet 21 Figure 3 4 Research schema 23 Figure 3.5 Flowchart of technological process of PM 24 Figure 3.6 Protein coagulation 25

Figure 3.7 Curds 26

Figure 3 8 Curd after aging 26 Figure 3 9 Draining & kneading 27 Figure 3 10 Moulding 27 Figure 3 11 Shema of experimental arrangement 1 28 Figure 3 12 Shema of experimental arrangement 2 29 Figure 3 13 Colorspace measurement 32 Figure 3 14 Texture profile analysis 33 Figure 4 1 The surface of milk curd at different PJ volume in the coagulation process 38 Figure 4 2 Mozzarella samples at different volume of Passion fruit juice 39 Figure 4 3 Milk curds at different coagulation temperatures 42 Figure 4 4 Milk curds surface at different coagulation temperatures 42 Figure 4 5 Milk curdling surface at different rennet concentration 46 Figure 4 6 Mozzarella samples at different rennet concentration 48 Figure 4 7 Flowchart of technological process of passion fruit mozzarella (PM) 49 Figure 4 8 Microstructure of the first reference sample (RS1), the second reference sample (RS2) passion fruit mozzarella (PM) 51 Figure 4.9 The result of sensory evaluation of mozzarella samples 52 Figure 4 11 Changes in Lightness value (L*) of RS1, RS2, PM samples during storage 53 Figure 4 12 Changes in a* value of RS1, RS2, PM samples during storage 53 Figure 4 13 Changes in b* value of RS1, RS2, PM samples during storage 54

LIST OF TABLES

Table 2 1 Classification of cheese by water content 5 Table 2 2 Classification of cheese by fat content 5 Table 2 3 Classification of cheese by processing 6 Table 2 4 Classification of cheese by coagulation method 6 Table 2 5 Mozzarella identity standards from CFR (Code of Federal Regulations 8 Table 2 6 Chemical composition ( Mean  SD) of raw camel and cow milk 12 Table 2 7 The nutritional value of passion fruit juice 16 Table 3 1 The technical parameters of rennet 22Table 3 2 Parameter and setting value of structural measuring device 33Table 4 1 Chemical composition of fresh cow's milk 35 Table 4 2 Physicochemical properties of PJ 35 Table 4 3 The effect of passion fruit juice (PJ) volume on mozzarella quality properties 37 Table 4 4 Effect of PJ on texture properties of mozzarella 38 Table 4 5 Colorspace results of mozzarella changed with casein coagulation temperature 39 Table 4 6 The effect of coagulation temperature on mozzarella quality properties 41 Table 4.7 Effect of coagulation temperature on texture properties of Mozzarella 43 Table 4 8 Color change results of PM and the reference samples 43 Table 4 9 The effect of rennet concentration on the mozzarella quality properties 45 Table 4 10 Effect of rennet concentration on texture properties of Mozzarella 46 Table 4.11 Color change results of PMs and the reference samples 47 Table 4 12 Quality criteria of PM and RS2 50 Table 4 14 The result of sensory evaluations of mozzarella samples 52

LIST OF ABBREVIATIONS

ABSTRACT

The research topic "Effect of Passion fruit juice on quality of Mozzarella cheese" was

carried out: a) to evaluate the effect of passion fruit juice concentration on the mozzarella properties; b) to determine the technological parameters (temperature, rennet concentration) of casein coagulation by rennet; c) to evaluate some of cheese qualities during 21 days storage

The analytical results showed that the passion fruit mozzarella (PM) made with passion fruit juice (PJ) volume of 9 (mL) had appropriate yield efficiency at 8.23% and reached the pH

at 5.9 Next to cheese made with rennet concentration of 75L/300 mL of milk and coagulation temperature at 37  1 ; as a result, the cheese yield efficiency (%H), total solids cheese (%TScheese), cheese solids yields (%CSY) was 9.47%; 55.86%; 45.94%, respectively Furthermore, the finished cheese had proteins, lipids, dry matter content: 24.10%, 21.30%, 52.20%, respectively Besides that, microbiological analysis also was conducted and gave the

result that no detected of harmful pathogens such as: Escherichia coli, Salmonella spp, Listeria

monocytogenes, Coagulase-positive Staphylococci The microstructure of PM had many sharp

edges, the shape of the structure resembles a cluster of particles Moreover, in descriptive sensory test, the PM samples had the difference in term of texture and smell compared with 2 other reference samples (RS1 and RS2) In addition, the average acceptability of PM samples were the highest, at 6.77/9, and RS1, RS2 samples were 5.87; 5.95 in grade 9 respectively

Through all the above results, we found that passion fruit juice had the influence on mozzarella making-process such as: chemical composition, texture, microstructure characteristics, sensory and nutritional value of the product Therefore, passion fruit juice is believed that it will have many potential applications in the food industry as whole and dairy product industry in particular

Chapter 1: INTRODUCTION 1.1 Pose the problem

Cheese is a high nutritional value food with a wide variety of flavors and textures Among them, Mozzarella is one of the most consumed cheeses worldwid They play a large role in several world markets, especially the North American marke Cheese factories typically produce more than 100,000 kg of pizza cheese per day (Fox et al., 2017c)

Source: USDA ERS

Figure 1.1 U.S Consumption of cheese (1975-2018)

With the development of the economy and the cultural exchange between the West and the East, cheese has become popular all over the world, including in Vietnam Cheese in general and mozzarella, in particular, are products rich in essential nutrients: especially proteins, amino acids, fats, fatty acids, vitamins, and minerals (Walther et al., 2008) Currently, the demand for Mozzarella cheese is increasing not only because of its health benefits but also can be used to combine with other dishes such as pizza, hotdogs, and hamburgers Therefore, the demand for cheese from customers is constantly increasing, requiring cheese manufacturers to make appropriate changes to meet the demand for that increase With hundreds of cheeses in different flavors, consumers will find the product that meets their needs During the information collection process, we found that the collection of documents on the production process of mozzarella and the influence of passion fruit juice on mozzarella was minimal There are few domestic studies

on the production process of mozzarella from raw milk in Vietnam with the conditions in Vietnam In addition, there have not been any direct studies in Vietnam on adding passion fruit juice to improve the quality and structure of mozzarella Therefore, it is necessary to perfect a mozzarella production process in the actual conditions of our country In mozzarella making process, the micelle casein will coagulate when the pH is reduced to a critical point by adding an acidic agent, typically citric acid - a flavor enhancer to food products, also a natural preservative, inhibiting the growth of bacteria and preventing oxidation in food products In Vietnam, citric acid is abundant in passion fruit, a popular fruit with a characteristic flavor and aroma Passion fruit juice contains citric acid, malate acid, lactic acid, ascorbic acid, and other organic acids (Casimir et al., 1981) In addition, passion fruit juice is a source of vitamin C ascorbic acid and vitamin A carotenoids (Knight & Sauls, 1994) This is also the second purpose of this research,

with the desire to create mozzarella cheese products that not only achieve improvements in performance and texture but also bring high nutritional values and new flavors In this study,

we used a combination of both rennet and passion fruit juice as coagulants for fresh cheese

+ Evaluate the qualities of passion fruit mozzarella (Chemical composition: total dry matter content, proteins, lipids, acidity, pH, microstructure of products, sensory test, microbiological criteria); the qualities of passion fruit mozzarella during 21 days of storage

1.3 Subject and scope of research

 Subjects:

 Fresh cow's milk imported from the breeding unit of Nong Lam University, Ho Chi Minh City

 Passion fruit juice which added to the Mozzarella production process

 Passion fruit mozzarella production process

 Research scope:

 The study will investigate the change of conditions during the production of mozzarella such as: coagulation temperature, passion fruit volume, rennet volume

 Study the influence of passion fruit juice on some quality characteristics of mozzarella

 The research was carried out at the laboratory of the food technology industry, Faculty of Food and Chemical Technology, Ho Chi Minh City University of Technology and Education

1.4 Research content

Effect of passion fruit juice on qualities of mozzarella includes the following contents:

+ Analysis of the chemical composition of materials (ash, moisture, protein, lipid, pH, titrable acidity and microbiological indicators, fruit weight and diameter)

+ Investigate the influence of passion fruit juice volume on properties of passion fruit mozzarella cheese (%H, %TSwhey, %TScheese, CSY (soild cheese yield), colorspace measurement and texture analysis)

+ Investigate the influence of coagulation temperature on passion fruit mozzarella qualities + Investigate the influence of rennet concentration on the qualities of mozzarella cheese qualities

+ Evaluate the qualities of passion fruit mozzarella (Chemical composition: total dry matter content, proteins, lipids, acidity, pH, microstructure of products, sensory test, microbiological criteria)

+ Evaluate the qualities of passion fruit mozzarella during 21 days of storage

1.5 The scientific and practical values of the topic

 Scientific significance:

- The process of producing fresh cheese with the addition of passion fruit juice can be used

as a reference for research and study at universities

Chapter 2: LITERATURE REVIEW 2.1 Cheese

2.1.1 Overview of cheese

Cheese is called name for a group of highly nutritious dairy products (Le, 2010) The collection name of chese is derived a group of food products made from fermented milk, produced in a variety of flavors and forms around the world (Fox, 1993) (Khanal et al., 2019) Cheese is produced by coagulating milk with the help of yeast or suitable probiotics or vegetable coagulants, vinegar and lemon and partially depleting the whey (Hickey, 2017) Initially, this was simply a method of preserving milk (Patrick F Fox, 1993), until now cheese has developed into a popular food due to its variety of applications, value nutritious, convenient and palatable to consumers This diversity is due to developments in cheese production technology, biochemistry and microbiology of cheese ripening (Farkye, 2004) Since ancient times, cheese has been an neccesary ingredient in the diet in a lot of country on the world Cheese is made from food product rich in proteins, fats, minerals and vitamins, besides containing bioactive amino acids, peptides and lactic acid bacteria (Malcata et al., 2016)

Figure 2 1 Illustrations of cheese products Fresh cheese is a cheese without biochemical ripening (Thanh, 2003), produced by

coagulating milk, cream or whey through acidification, acidification with small amounts of rennet, or a combination of acid and temperature, can be used immediately after production is complete (P F Fox et al., 2017 )

There are many high nutritional value of fress cheese in dairy product In which, the protein content is about 15-20% including all kinds of important amino acids such as metionin, lizin, In addition, fresh cheese also has a significant amount of minerals Ca, P, Fe, Mg, needed for cell growth, bone formation and metabolism in the body Whey is separated from fresh cheese by gravity rather than mechanical pressure to help maintain a smoother texture and

higher moisture content Therefore, fresh cheese has the highest moisture content of all cheeses

In our country, fresh cheese has begun to be produced but not much Currently, on the market, there are many fresh cheese products from different companies with unique quality and taste

2.1.2 Classification

Currently, there are many types of cheese in the world Cheese products differ in structure, odor, taste, color, physicochemical and microbiological parameters (Le, 2010) There are different methods for classifying cheese: grading based on texture (depending mainly on moisture content), grading based on coagulation method and grading based on ripeness of the cheese indicators (Khanal et al., 2019)

Table 2.1 Classification of cheese by water content

Product Type Water content (%)

Very hard cheese < 41

Product Type Fat content (%)

Cheese has a very high fat content > 60

Cheese with high fat content 45 – 60

Cheese with medium fat content 25 – 45

Cheese with low fat content 10 – 25

Source: (Le, 2011)

Table 2 3 Classification of cheese by processing

Product Type Characteristics

Cheese has gone through the

- On the surface of the cheese block

- In the cheese block depth

- On the surface and in the depth of the cheese block

Source: (Le, 2011) Table 2 4 Classification of cheese by coagulation method

Coagulation method Product

Coagulation by Rennet Most major international cheeses

Acid coagulation Cottage, Quark, Queso, Blanco

Heat/acid coagulation Ricotta, Manouri, Sapsago, Ziger, Schoottenziger,

some forms of Queso Blanco

Source: (P F Fox et al., 2017)

2.2 Mozzarella Cheese

Mozzarella cheese is type of a soft, unripe of the Pasta-filata family that originates from the Battipalia region of Italy Cheese in general and mozzarella in particular is made by freezing and fermenting (incubating) the milk of some animals, such as cows, buffaloes, sheep, goats Traditionally, mozzarella is made from buffalo milk Buffalo milk is a high Vitamin A, protein and low cholesterol, thus this traditional cheese made from buffalo milk has more preferred characteristics

Mozzarella is taken into account a fress cheese as well as it is famous for a mild flavor and rubbery texture Mozzarella cheeses dated back in the southern areas of Italy, from the Battipalia region manufacturing from milk of cow however traditional cheese with main ingredient is that from buffalo milk, i.e., Mozzarella di Bufala Until now, mozzarella is created throughout Italy, another European countries and the United States from a different of dairy materials , but the

2008) Mozzarella cheese is sort of a soft, unripened and white cheese whose melting and

stretching propertie, especially mozzarella cheese is highly suitable for pizza making (Kindstedt,

1995) These functional properties of mozzarella cheese are effected by a variety of elements that contains cheese composition particularly moisture, fat content, pH, coagulating enzyme, starter culture, cooking and stretching, salt content and the changes occurring during aging and storage

(McMahon et al., 1996)

The finished cheese is lightly salted, white, soft with a very lively surface sheen and has unique propery of stretchability Due to this, it can be used to make pizza toppings, lasagna, and veal cutlet alla Parmigiana Since mozzarella cheese naturally stretches, it is often used as a topping on pizza The popularity of pizza joints, especially among young people, has increased mozzarella cheese manufacturing In Italy, 78% of the country's entire cheese production is made

up of mozzarella and pizza cheese (Merrill et al, 1994) In the production of this cheese, after fermentation the curd (pH∼5.1 – 5.4) undergoes a plasticization step at the end of production by kneading and spinning in hot water ( ∼80 – 85℃) or dilute saline (Guinee et al., 2002) It was the low pH and high temperature that facilitated the formation of para-casein fibers with relatively high tensile strength (Kimura et al., 1992; Taneya et al., 1992) After the cheese is cooked on the pizza, these threads give the melted cheese its spinning, stringiness, and chewiness (Pf Fox et al., 2000)

Figure 2 2 Mozzarella cheese

2.2.1 Classification

According to Codex standards (WHO/FAO) mozzarella is divided into two main types based on usage:

1 Mozzarella has a high relative humidity, produced for direct consumption

Mozzarella has high moisture content (52 – 60%), high pH (5.6 – 5.8), high moisture content, low salt content (0.1 – 0.5%) and short shelf life (21 days) (Clark et al., 2009) Fresh mozzarella is usually originated by adding acid directly to acidify milk rather than acidifying bacteria Products are usually formed into small spheres with the size of different sizes and are consumed immediately after production Fresh mozzarella is frequently consumed raw because

of its delicate milky flavor and tender texture, which make it a good choice for salads, sandwiches, and appetizers This cheese is not commonly used as an accompaniment to other

dishes because the protein matrix contracts when heated, releasing the whey, making the dish overly watery (Clark et al., 2009)

2 Mozzarella has low moisture content and is frequently used as a component in foods (Clark et al., 2009)

Low moisture mozzarella (45 - 52%) is a product with lower pH (5.2 -5.4), lower moisture content, produced by using bacterial culture method (Clark et al., 2009) Because of its lower pH, lower moisture content, and higher salt content, it has a longer shelf life Low moisture mozzarella can be produced to have a variety of spinning and melting properties, is solid enough to be chopped or sliced, and can be cooked without the need of excessive water Due to its special processing and cooking qualities, mozzarella is frequently used Low-moisture mozzarella that has not been cooked can be used in sandwiches, as a topping for pizza, as a filler for lasagna, and even breaded for use as an appetizer (Clark et al., 2009)

Beside that, due to code of Federal Regulations, mozzarella can devide into 4 groups:

Table 2 5 Mozzarella identity standards from CFR (Code of Federal Regulations

(CFR) (b) Title 21, Part 133.155 thru 133.158)

Type of cheese Moisture Fat-on-dry matter (%)

Mozzarella ( Low moisture, part skim) 45.0-52.0 30.0 - 45.0

2.2.2 Casein coagulation

The method of changing casein from a colloidal state to a curd with a gel structure occurs during casein coagulation in milk (Le, 2010) This is one of the important stages of cheese production in general and mozzarella in particular The structure of casein micelles is very sensitive to changes in factors such as pH, temperature, and ionic strength Therefore, the study

of the modification of casein micelles has been applied in the food industry to improve product quality There are many different methods applied and in the cheese industry, milk coagulation methods are often used: isoelectric (acid) coagulation, heat and acid coagulation, rennet (enzyme) coagulation

2.2.2.1 Coagulation by acid

By lowering their charge, dissolving part of the insoluble calcium phosphate crosslinks, and altering internal protein bonds, acidification has a direct impact on the stability of casein micelles At some crucial point, when electrostatic repulsion is diminished and is unable to repel attractive forces like hydrophobic contacts, aggregates and eventually gels begin to develop Acid-induced milk gels become more rigid over time as a result of continuing casein particle-to-casein link formation inside the network

Normal milk contains between 80 and 95 percent casein in the type of micelles, which are colloidal dispersions that precipitate at a pH of 4.6 (isoelectric pH) at 30ºC (Brunner, 1977; Dalgleish, 1982; Patrick F Fox et al., 1998) Milk acidification can be performed using bacterial cultures, fermenting lactose to lactic acid, direct addition of acids, such as HCl, or using glucono-δ-lactone (GDL) (J Lucey et al., 1997) is hydrolyzed to gluconic acid, leading to a decrease in pH We bring the milk pH to the isoelectric value of casein 4.6, it will reduce the electrostatic repulsion between the molecules, then the interaction between the hydrophobic radicals of the protein molecules will prevail, the casein will switch to insoluble state and the appearance of milk coagulation (Le, 2010) The process of coagulating casein in milk by adjusting the milk pH to the casein isoelectric point, acid coagulation (lactic or citric) method is used in production technology of some fresh cheeses such as Cottage, Quarg, Cream cheese, etc Acid coagulation represents 25% of total cheese production and is commonly used fresh (Mcsweeney et al., 2004) Manufacturers will use lactic acid bacteria to create lactic acid, decrease the pH value of milk, causing coagulation of casein (Le, 2010) In addition, the use of lactic acid bacteria also serves the purpose of biosynthesis of some metabolic products to contribute to the characteristic flavor of cheese

2.2.2.2 Coagulation by acid and heat

Review of component and processing parameters' influence on the textural characteristics

of fresh acid cheeses and acid milk gels (Lucey & Singh, 2003) Fresh acid cheeses use a mix of heat and acid to coagulate milk, cream, or whey Pretreatments, such as heating the milk or homogenizing it, delayed acidification often using mesophilic starter cultures and gelation, whey separation, and/or curd treatment are frequently required for the creation of fresh acid cheeses (Sheehan et al., 2004) Coagulation by a combination of heat and acid is used for a limited number of cheeses, including ricotta and manouri (Fox et al., 2017a) Queso blanco is a cheese from Latin America that is created by boiling milk to a temperature of 80 to 85°C, adding acid, and then gently churning the milk to create a curd Acetic, citric, phosphoric, among other acids,

or simply lemon juice, might be

The κ-casein molecules are crucial for maintaining the milk's micelle structure (Le, 2010)

It has been discovered that during the rennet coagulation of milk, only κ-casein is hydrolyzed and that it is hydrolyzed to para-κ-casein and macropeptides also known as glycomacropeptides because they include all or most sugar groups associated with κ-casein (Fox et al., 2015) At this stage, only κ-casein molecules are hydrolyzed at the above specific site, hydrophilic macropeptides diffuse into the surrounding medium while para-κ-casein remains attached to the micelle moieties, makes micelles “easy to assemble” together in preparation for agglomeration

(Le, 2010) The micelles become less stable and start to coagulate when 85% of the total casein in milk has been hydrolyzed (the coagulation temperature is usually 30℃) During the coagulation process, the micelles will drag fat particles and some other compounds in milk including lactose, vitamins, salt, etc

κ-Casein

Figure 2 3 The mechanism of coagulation by rennet With the right temperature and time, the milk curd has been formed, the milk serum separation stage is performed For milk casein coagulation agglomeration of milk with a particle gel structure, casein gels are induced by enzymes formed by filaments, each consisting of many interacting casein particles, and each granule considered as aggregates of micelles in milk This granular gel structure contains straight fibers, through which part of the milk serum from within the coagulants will escape Therefore, cutting the curd will make more milk serum escape, the finished cheese has low moisture content, high hardness (Le, 2010) Besides the curd mass formation process, the enzymatic coagulation of casein is also influenced by external factors They depend on temperature, at <15℃, the milk solution will not be able to coagulate (Le, 2010), at a temperature of >55-60℃ depending on the pH and concentration, the enzyme will be denatured When the pH of milk is decreased, calcium phosphate is dissociated from the micelle structure, reducing the charge of the micelle, the enzyme moves closer and closer to its optimal

pH, increasing the reaction rate (Fox et al., 2015) Increasing the concentration of Ca2+ also changes the equilibrium of the system, they reduce the charge of the casein, reduce the electrostatic repulsion between the micelles and promote coagulation The higher the amount of enzyme used, the faster the hydrolysis reaction of κ-casein, the more micelles that can release macropeptides on the surface, the faster the coagulation rate (Le, 2010)

The complex group of enzymes generated by the rennet in the stomachs of young ruminant mammals, mainly calves (Uniacke-Lowe & Fox, 2017) The most important component of rennet is Chymosin - a protease enzyme used to coagulate caseins in milk (A Kumar et al., 2010) Pepsin and lipase are two more enzymes found in rennet in addition to chymosin

Rennet's main functions include the protease chymosin Its cleavage cleaves the caseinomacropeptide from the κ-casein chain, causing destabilization of the casein micelles (Corredig & Salvatore, 2016) Milk coagulates as a result of the casein's adhesion to other cleaved casein molecules and formation of a lattice (Corredig and Salvatore, 2016) They can clot better in the presence of calcium and phosphate, which is why these minerals are occasionally added when making cheese (Lucey, 2002) The network of solid casein proteins is cut that holds together the other components of milk, such as fats and minerals, to make cheese

Animal rennet: is a complex mixture of enzymes, the main one being chymosin, which is

normally obtained from the stomach of calves (O’Connor, 1993) Young animals' gastric glands secrete animal rennet; when adults consume milk, rennet also occurs in their stomachs Because

Para-casein + macropeptides

Gel

Ca2+ ,  20oC Rennet

rennet from each species has somewhat varying chemical properties, utilizing sheep's rennet to coagulate milk is the best option, but the difference is not significant Milk coagulates quickly with animal rennet at a pH that is natural and with minimum degradation of milk proteins (Garg

& Johri, 1994) Acids, neutral calcium and barium salts, and the occurrence of coagulation all speed up this process Slower when mixed with distilled water, potassium, sodium, or base salts

In addition, animal rennet added during cheese making improves whey separation and increases density hardening of curd mass (Fox et al., 2017b) Based on the above advantages, we selected animal rennet sources to use for this study

Plant rennet: There are numerous recognized milk-derived proteolytic enzymes, including

bromelain (EC 3.4.4.24) from pineapple, ficin (EC 3.4.4.12) from Ficus species, and papain (EC 3.4.4.10) from papaya These enzymes appear to be present everywhere in plant structures Buds, flowers, fruits, latex, leaves, roots, sap, and seeds all contain them Ash gourd, fig, milkweed, papaya, pineapple, prickly artichoke, pumpkin, and soybean are a few of the most well-known plants Sadly, the majority of plant coagulants have shown to have non-specific proteolytic action Most plant rennets are unsuitable for manufacturing cheese because the ratio of milk-clotting function to proteolytic activity is so low; excessive proteolysis results in bitter products, but bitterness lessens over the course of 3-6 months of storage (Garg & Johri, 1994)

Microbial rennet: used for milk fermentation has also been developed through genetic

engineering by genetically modifying the fermenting microorganisms, providing the market with many innovative products (Islam et al., 2021) However, in certain nations, microbial coagulants are not permitted for the manufacturing of cheese because it is thought that they will result in poorer yields and poorly performing (Garg et al., 1994)

Although rennet's primary role in cheese making is to coagulate milk, it also contributes to improved flavor and texture by participating in proteolysis during cheese ripening (Fox, 1988) Coagulation is directly affected by the type of enzyme used, the ratio and composition of substances present in milk fat, protein, whey and casein, the presence of calcium ions and pH (Fox et al., 2017a)

In the Mozzarella production process, coagulation of milk casein to create curd is an important process Nowadays, many coagulation agents have been studied and found Among them, the most common is the enzyme coagulant (rennet) with 75% of the total cheese yield (Fox

et al., 2017d), along with previous studies by D'Alessandro (2019), Mme (2018) about Mozzarella, the coagulation agent is rennet In addition, we use passion fruit juice as the source

of acid for this process to lower the pH of the milk, it is effective in the coagulation

2.3 Raw material

Traditional mozzarella is made from buffalo milk (McSweeney et al., 2017) and Mozzarella produced from buffalo milk is the best quality cheese until now (Zicarelli, 2004) Nevertheless, the source of raw milk depends on the region that to use the vary of mammals like buffalo, cow, sheep, goat, or camel

2.3.1 Raw milk

Milk is a liquid secreted from the mammary glands of animals, containing high levels of nutrients The variations in breed, environmental factors, and breeding conditions all affect the essential ingredients in milk The milk from various animals varies greatly Although dairy cattle and goats have been raised in our country for a long time, for many reasons, the production of these types of milk is still very low So far, the raw material for the dairy industry in Vietnam is still cow's milk There are several factors that significantly influence milk composition including breed, feed composition and lactation stage

Milk is a very complicated liquid that is made up of four primary parts namely water, lipid, protein, lactose and about 100 minor components Among them, the most important ingredient for cheese production is calcium phosphateThe characteristics of one of the milk's casein and protein groups, in particular, affect the creation and quality of cheese Whey, that contains soluble components, removes the majority of the water present in milk 90% of it, i.e whey protein, lactose and some inorganic salts (P F Fox et al., 2017)

Good quality milk serves as the foundation for making cheese of top quality, which has a clean taste, is low in somatic cells, contains no antibiotics, and has a relatively low bacterial count The yield, quality, and functional attributes of cheese are influenced by the milk's composition, which is influenced by the breed of the lactating animal, season, lactation stage, disease, and genetics To guarantee constant cheese quality and depending on the raw milk's microbiological quality and its source, hydrogen peroxide (H2O2) can be added to the raw milk (Moatsou & Moschopoulou, 2014) In addition, the production of mozzarella is also impact by the lipid and protein/casein content of the milk The higher the casein content in milk, the greater the cheese recovery in the cheese making process (Le, 2010)

Table 2 6 Chemical composition (Means  SD) of raw camel’s and cow’s milk

Parameter Camel Cow

Milk protein

Cow's milk usually contains about 2.9÷5.0% protein, with an average value of 3.4% Milk proteins can be divided into 75÷85% of total cow's milk protein as casein and 15÷25% as whey protein (based on their solubility at pH=4.6) and enzymes ( Le, 2010) In fact, the properties of many dairy products depend on the presence of milk proteins, although fat, lactose, and salt also exert significant effects The majority of cheeses are created by coagulating casein or using proteolytic enzymes to change proteins, dairy products that withstand high processing temperatures are only possible thanks to their thermal stability milk protein, casein (P F Fox et al., 2015)

Casein is the main protein found in milk They exist as micelles, each containing about 65% water, the rest being caseins and minerals (Le, 2010) Casein micelles are easily destabilized by proteolytic enzyme treatment or by acidification to produce coagulants, which are the basis of cheese and other products (Douglas et al., 2012) When the pH of the milk is reduced to 4.6, the caseins agglomerate and, if acidified under static conditions, form a coagulant (Pf Fox, 2003) This agglomeration of milky juices can occur at various temperatures, but their tendency to agglomerate will be different and into different forms, depending on the purpose of product formation At a temperature of 30-35℃, the curd is quite hard, coarse and easily precipitated from the initial milky solution (Pf Fox, 2003) By processing milk with a particular proteinase, casein can be coagulated and converted to rennet casein (rennet) One of the caseins, κ-casein, is hydrolyzed during renneting, resulting in fundamentally different characteristics between rennet casein and acid casein (made by acidification with lactic acid, hydrochloric acid, sulfuric acid)

Casein and whey protein groups both are heterogeneous, each containing a variety of proteins (P F Fox et al., 2015b) Whey protein is a mixture of proteins, isolated from whey, the liquid left over when milk is coagulated during cheese making, containing milk soluble substances (Tunick, 2008) with diverse functional properties, the main proteins being β-lactoglobulin and α-lactalbumin, serum albumin, Immunoglobulin, proteosepeptone and among many others (Cayot and Lorient, 2017) Whey protein refers to milk proteins that dissolve at pH=4.6, which can form fibrous, spherical aggregates, etc depending on the conditions, heat treatment method (Nicolai et al., 2013) The ability to stabilize, difficult to denatured whey protein in cheese depends on the temperature change with 95℃ (Guyomarc'h et al., 2014), however, for the studied product Mozzarella, the collection whey is not required

Milk Fat

Milk fat is usually in the type of globules surrounded by a lipoprotein membrane, which maintains the integrity of the fat globules and stabilizes the emulsion in the milk (Christie, 1983) Fat globules are composed mainly of glycerides, phospholipids and the protein Phospholipids are a small but important part of the lipids in milk and are found mainly in the milk fat globules, where they bind glycerides and proteins The phospholipid molecule has two ends: the lipophilic (non-polar) end will interact with the glyceride molecules in the center of the fat globule and the hydrophilic (polar) end will combine with the protein membrane surrounding the fatty globule (Le, 2010) Therefore, fresh milk as a raw material, it is necessary to maintain the milk storage temperature at 4℃ to limit the growth of bacteria in milk as well as limit the breakdown of fat in

milk (Muir et al., 1978), avoiding the fatty globules from combining together to make them float

to the surface of the milk, causing the preserved milk to separate

2.3.2 Passion fruit

Milk coagulation is the first step of cheese making and is of importance, as it affects the whole process and the final cheese (Wium et al., 2003) So, during cheese making, the pH of the milk needs to be reduced a bit before renneting, which reduces the electrostatic repulsion between the casein micelles and promotes close contact, increasing the concentration rate synthesis of paracaseins and accelerated coagulation (Zoon et al., 1989) However, the optimal coagulation of rennet is at around pH 6.0 and the pH of milk is in the range of 6.7 (Ali A S Marouf, 2018), so it is necessary to add an amount of acid to lower the pH of milk for rennet to work optimally Therefore, in this study, we used passion fruit juice to adjust the pH, and at the same time enhance the sensory properties of the product

2.3.2.1 Description

The Passion fruit vine is a shallow-rooted, woody, perennial, climbing by means of tendrils The alternate, evergreen leaves, 3 to 8 in (7.5-20 cm) long, deep-green and glossy above, paler and dull beneath, and, like the young stems and tendrils, tinged with red or purple, especially in the yellow form The nearly round or ovoid fruit, 4-7.5 cm in width, has a tough rind, smooth, waxy, ranging in hue from dark-purple with faint, fine white specks, to light-yellow or pumpkin-color Within is a cavity more or less filled with an aromatic mass of double-walled, membranous sacs filled with orange colored, pulpy juice and as many as 250 small, hard, dark-brown or black, pitted seeds The flavor is appealing, musky, guava-like (Sushma Kumari, 2016)

2.3.2.2 Origin and classification

 Origin

It is known as passion fruit in the United States, passion fruit in the United Kingdom and Commonwealth, and purple granadilla (South Africa) It is widely farmed in South America, Central America, the Caribbean, Africa, Southern Asia, Vietnam, Israel, Australia, South Korea, and the continental United States for its sweet, seeded fruit (Mary KawenaPukui, 2003) Passion fruit was introduced to Hawaii in 1880, but it was first commercially farmed in Kenya in 1933, and it quickly gained popularity in home gardens Early in the 20th century, Australians introduced the purple passion fruit to Israel, where it is now regularly grown in private gardens along the entire coastal plain and sold in small numbers to processing plants In the mid-1950s, interest in yellow passion fruit cultivation increased in Colombia, Venezuela, and Surinam in South America

 Classification

Passion fruit is a short-lived, evergreen perennial vine that bears aromatic,

tropical-flavoured fruit The purple passion fruit (Passiflora edulis Sims) and, to a lesser extent, the