Luận văn thạc sĩ Công nghệ thực phẩm: Fabrication of red palm oil emulsion gel using combination of plant-based starches

LIST OF ABBREVIATIONSRPO: Red palm oil CPO: Crude palm oil EG: Emulsion gel EG 10%: Emulsion gel with 10% concentration of starch EG 15%: Emulsion gel with 15% concentration of starch EG

MINISTRY OF EDUCATION AND TRAINING

NONG LAM UNIVERSITY - HO CHI MINH CITY

Faculty of Chemical Engineering and Food Technology

FABRICATION OF RED PALM OIL EMULSION GEL USING

COMBINATION OF PLANT-BASED STARCHES

A Thesis submitted in partial fulfilment

of the requirements for admission to the degree of

Bachelor of Engineering in Food Technology

ByStudent: Huynh Thi Phuong Uyen

Supervisor: Prof Dr Tan Chin Ping

Dr Tan Tai BoonAssoc Prof Dr Kha Chan Tuyen

Ho Chi Minh City, 2024

MINISTRY OF EDUCATION AND TRAINING

NONG LAM UNIVERSITY - HO CHI MINH CITY

Faculty of Chemical Engineering and Food Technology

FABRICATION OF RED PALM OIL EMULSION GEL USING

COMBINATION OF PLANT-BASED STARCHES

A Thesis submitted in partial fulfilment

of the requirements for admission to the degree of

Bachelor of Engineering in Food Technology

ByStudent: Huynh Thi Phuong Uyen

Supervisor: Prof Dr Tan Chin Ping

Dr Tan Tai BoonAssoc Prof Dr Kha Chan Tuyen

Ho Chi Minh City, 2024

TABLE OF CONTENTS

LIST OF ABBREVIATIONS wasssisssssnsssssusesarsieamacnnwsimenancenernannnenrnnneneamnrenes 1 LIS TOR TABLES hinaosptstoissen 04615592 ngi09650 885038588: iaAirlsjsstgtastbossAlqoiil9stlssGddirlisll\lv3voqltÄ8NgBS43495643600489Ngs0s80, 2 Mis TSS SNF CO) Fe Go RS i a a me Re 3 ACKNOWLEDGEMEN TD ossesssessnenesoammenaaszresoanennnumennenavescunm enna ecnenannveneenivaasceunrenunreonuecteansenoeesned 4 ABSTRACT siisscassncscsemncnuecennvnncrsinecnvencnas aren ananmanrnrne ema uar TR 5 CHAPTER 1 LR KD LJ CC HO Nhac y5u224115084g01686113NcEEE13-40000GS85632453 16-0 nasa Seat eR 6 Non 1 6 l2: (ẬẲHssiawesdsdoesdinniiDieniiiiADGEDAEI4100910055090044091S851S00L5X134490319S0IS413093591945390193139328913060391199879568 §

lộ, - QUISEHVESnearnsagasernsdiadrbiODODUEGSLIEHIMMEIASUHINQIAUIGHONIDERRISHRRHSEERHUGNROSVNNGSIRRRRHBSURG 8 CHAPTER 2: LITERATURE REVIE W LH HH TH TH HT HT HT Hư 9 Dol, ROd ĐAlHfiốilESBASEGERNHURIGusaoenamddnnndididunondpndlidindidbisaodthaositeaidvausdiekolilBesesagmdieneoisôdfed 9 21s Réd palm 01) vsswsnsisteannvarinnnimuraa enc inate 9 3.12 Red palin oil extraction sesceccsstns66566811580506301358556618885330438855538ESE543205615E645ES5SE/306408388G8318 10 BED, — THHUNIOÍbi¿sossusunsnsoaoiieorioinitisosnuiisiesebvodiBii6fAS/074880064649u8i63020ã2ci800n5ui03ugnj6aã0gđSumststiSlGletSbslawEsfluEm 13 2:3 Hiph-Pressure Homopenization PTOCESS : cicscccibiiciibiiioiiciiaeLi-xesllSbsgEx166s163020.3 15 2c - BHHBHGTEEÏusssesesesaeindtodsonndlBiAEDDDDEHOEASAIAHGLONOSDUEOIRESSVOIAHSEVNENGVEEIAGBESSSSĐESRIGC.S0SĐASEASUEM 16 2:5, PlantbasedstarclsssssssssespassnsasanooondttirsidrdddsDleDAEELSLDSESEEEDISSESG3030114551301481599/383992138 20 25:1, - JPO(RIORSIREEHsszxsaoaggrttointiiGEDOIIROGGUEIRSGEEAAGGISGSSESEHIBSEEEEERHERNIGNUSIHHGIADMEGUEURMR 20

"ta 22 CHAPTER 3: MATERIALS AND METHODS ssssiesessescannecroenesescnnsenrerravrnnnucnneasinaeserenaeerseesey 24 Bil; - Material Siassccscusvamanrmeaninem nance anne near eumranre aR 24 3.2 ốc ha 24 3.2.1 Red palm oil-based emulsion pDr€parafiOII - 5 65+ 2+ St S3 +2 EEEEEkkskkekrrkskrrkrrke 24 3.2.2, Préeparationof starch-based emulsion ĐEÏssssssasseaodiedodiooioislloeddrodlisailstedtloasteasstoe 25 3.23 Investigation of the different formulations for red palm oil starch-based emulsion gel 25

3.2.4 _ Investigating the selected optimum emulsion gel during storage - ‹- 27 3.2.5 Statistical aaly sis wcssssscscssssesesenennessemnnrenrsenseevrensmrneainveemennomnmerrasaeoeeens 27

CHAPTER 4: RESULT AND DISCUSSION song Ra nh HH Đi Là 81A S45018290556113455G01/38144858 28 4.1 Investigation of the different formulations for red palm oil starch-based emulsion gel 28 4.1.1 Gel appearance (Tube Inversion method) - óc 2c 2212322 3 1512111111 ree 28

412, 'LEXHITECHAFESGETIRHGoysrmoarndoadoStlitvgrrogilitdtlgtGHGSEIEREGOEDOIQGIRGESRINSRSISOIiNOUASnuSR 29 4.1.3 Water Holding Capacity and Water ACLVIẨY LH HH “HH HH H4 2n 35 4.2 Investigating the selected optimum emulsion gel during storage - ‹ +<-+++ 37 4.2.1 Wisual ZDfiEafaifoe:0EeiSÍOH'EBÌSsescoiesi:3öc0gi06GIESEDSELGSXSDGISIERSSPEEGEHEEEgSiESi4SEsiSERS 37 4.2.2 Scanning electron microscopy (SIEEM|) - - tk TH HT HH 384.243 Water holding Capacity bsseeseaosoioieviseL0109199015623L0033016093504141439155086/391851800433000402480900590sS6 404.2.4 Texture characteris te sr vercie bit g2 HRGEHSULEBIGIEIEBIEESIEEGRISGSEHGEIEVEGESISGEISHASRDERERĐSfS-idpiE 43 4.3 Potential application in plant-based che€€se - - c3 1v 1 9 19 1111 111118 12x re, 46 CHAPTER 5: CONCLUSION AND RECOMMENDATIƠN SẶ St te 49

Sl, CÔNG.USIONGassvassbsasdseotirotsoboltSsdiBliseyesidistdsnddteodiadtbfulitotvesienaytesedi 49

52, RECOMMENDATION cussseuossaaaiceoiasaoodtiioioiieioiliagloiSG03400466836524403S00039469040.40A63036Aaeg8d 49 KEEEREERNE S is csccssssanssenennsrona nnenin saad atk se vii hee nen Nea ARNOT 51 APPENDICES

LIST OF ABBREVIATIONS

RPO: Red palm oil

CPO: Crude palm oil

EG: Emulsion gel

EG 10%: Emulsion gel with 10% concentration of starch

EG 15%: Emulsion gel with 15% concentration of starch

EG 20%: Emulsion gel with 20% concentration of starch

PS: Potato starch

CS: Corn starch

WPI: Whey Protein Isolated

HPH: High Pressure Homogenization

TPA: Texture Profile Analysis

Aw: Water activity

WHC: Water Holding Capacity

LIST OF TABLES

Table 2.1 Health benefits of phytonutrients found in red palm oIÏ - 10Table 4.1 Water holding capacity (WHC) of emulsion gels of different concentrationsand at different storage temperatures after 1 day of sfOraØe - -++-+++<++2 41Table 4.2 Water holding capacity (WHC) of emulsion gel in different concentrationsatid teifipEfaffeS ALLEL SOAVS Of SOTA LES sss ccssseenssuseenavecansrnxaenasmaannrenmenenanmausvarmnenn’ 42Table 4.3 Water activity (ay) of emulsion gel in different concentrations and

temperatures after 1 day of SfOFÀ€ 2c 2 1211121121 1151 12 1 111211111111 11101 1x rret 42Table 4.4 Water activity of emulsion gels in different concentrations and temperaturesafter 30 40:01 Ả ố 42Table 4.5 Texture profile analysis (TPA) of emulsion gel in different concentrationsand temperatures atter 1 day Of SON cá scsnissssasissenncasncexsamasnanevs swavarnanstannastaas cacteenncanee 44Table 4.6 Texture profile analysis of emulsion gel in different concentrations and

temperatures after 30 days Of Storage cccecccecesssceeesseeseeseeeseeseeeseeseeeseeseenseeeeeseeneenes 44Table 4.7 Spreadability of emulsion gel in different concentrations and temperaturesmiter lida Ol SLOTEđĐE sat seme ceeeeatraures cic veras sca eee eae eRe 45Table 4.8 Spreadability of emulsion gel in different concentrations and temperaturesafter 3U: daws @f SÍOTĐĐibicscscssoi con euccactaassnscteadvancersenansqunauaneencantuems <eenseea tae apeeamaaeed 46Table 4.9 Ingredient list, and photographs, of dairy and plant-based products 47

LIST OF FIGURES

Figure 2.1 Simple processing flowchart of red palm oil and refined, bleached and

deodorized Palit Othe wicssecesweesneennnecemmmoneeeseamennennevaumemamenenenenasannenmencesenmnees 13Figure 2.2 Summary of various methods, in which plant-based proteins,

polysaccharides, and lipids are used in the formation of plant-based emulsion gel 18Figure 4.1 Gel appearance of emulsion gel at different concentrations and ratios

between CS and PS: A) 100:0; B) 75:25; C)50:50; D) 25:75; E) 0:100 28Figure 4.2 Texture profile analysis (TPA) of Emulsion gel with 10% concentration ofstarch in different ratios of CS and PS ccsesescsnsesessesnsstensesseesdsecscsnsencsssenesssensessees 30Figure 4.3 Texture profile analysis (TPA) of emulsion gel with 15% concentration ofstarch in different ratios of CS and PS wo eee eeeseeeseesesseeseeseeseeseeeeeseeeeseeeeeseeeeeaeens 31Figure 4.4 Texture profile analysis (TPA) of emulsion gel with 20% concentration ofstarch in different ratios of CS and PS - - c2 11129 v12 ng ng ng rưy 31Figure 4.5 Spreadability of emulsion gel with 10% concentration of starch in differentPALO SCS ANd BS xuonsbctsoagtadizlkSiaSug68Ä05g:0ã308,l55E88g5ã8u8ã:40:g83G2608ãS0g5303885g883ö18003:G882ãx/85Si25800:563G00034Ex 33Figure 4.6 Spreadability of emulsion gel with 15% concentration of starch in differentTAUOS'CS BNE PS wescercesmreasusesenvaansnstemstae raven we ames snete woul volnls gE1880S8300183/8889883/3Ba88088p05i88108 34Figure 4.7 Spreadability of emulsion gel with 20% concentration of starch in differentratios CS and PS siresnnemenesmwnss ewer eemman arene 34Figure 4.8 The water activity of emulsion gel in different concentrations and ratiosbetween: CS and PS cecrcscnmssseseuescuseamanrnueaeeacmasae eer ecReEE eT ES 36Figure 4.9 The water holding capacity of emulsion gel in different concentrations andratios between CS atid PS ¡unesosoaneinni00116604154164598864019383011631831181833) 6143154 0885113483688 36Figure 4.11 Visual appearance of emulsion gel in different concentrations and

temperatures after30 daysofst0rate nnn memnmnmnnenninendieecntermminneninowns 38Figure 4.10 Visual appearance of emulsion gel in different concentrations and

temperatures aiter 1 day-of storage’ sisceescsacnnsercwetgenwamaeneddewensemesmarentinnes 38Figure 4.12 SEM images of emulsion gels after 1 day of storage: (A) EG 10% at 4°C;(B) EG 10% at 25°C; (C) EG 15% at 4°C; (D) EG 15% at 259C -. c©+c<+cs++ 39Figure 4.13 SEM images of emulsion gels after 30 days of storage: (A) EG 10% at4°C; (B) EG 10% at 25°C; (C) EG 15% at 4°C; (D) EG 15% at 259C 39Figure 4.14 Micrograph of emulsion gel after 30 days of storage: (A) EG 15%

(75PS:25CS) at 4°C under ultrasonic treatment; (B) EG 15% (75PS:25CS) at 4°C

without ultrasonic ff€afIT€TIÍ - - c6 2c 3263351151153 1123 1218511511511 11 11 811 01 1 rệt 40

Firstly, I would like to demonstrate my gratitude to the Deans of the Faculty of

Food Science and Technology at Nong Lam University and Universiti Putra Malaysia

for allowing me to complete my final year project in Malaysia

I could not complete my journey without the support and encouragement of my

supervisors, Prof Tan Chin Ping and Dr Tan Tai Boon, for giving me a chance to work

and study in the Fats and Oil Laboratory In addition, I am deeply thankful to Dr Tan

Tai Boon for his invaluable advice, continuous support, and patience during my research

at UPM His knowledge and experience were invaluable in helping me in both my

adjustment to living overseas and in my final year project

Moreover, I also received support from my Vietnamese instructor — Assoc Prof

Dr Kha Chan Tuyen His advice and guidance led me to complete this project

A special thanks to Dr Khor Yih Phing and the Food 5 members — Thong

Shuen, Li Ann, Somayeh, Atikah, Wana, Khai Yi, and Ming Yang, who supported me

and my classmate throughout the time we did the research at UPM They were always

willing to guide us on how to use the machines in the lab and share the most advanced

knowledge in the field to do my research

Thanks to the Food 5, Food 3, and IBS officers who created conditions for me

to use the equipment for my graduation thesis

Lastly, I am grateful to my family, who raised and supported me Their belief

me has kept my spirits and motivation high during this process

The advancement In using plant-based emulsion gel for low-fat products orvehicles to deliver functional food ingredients via encapsulation is currently noteworthy.The study presents the fabrication of an emulsion gel from red palm oil (RPO) combinedwith the mixture of corn starch (CS) and potato starch (PS) as the gelling agents Thisstudy investigated the effects of the concentrations of plant-based starch complex, theratio between CS, PS, and temperature in fabricating the emulsion gel with RPO Theeffect of different concentrations, ratios of CS and PS on the water activity, waterholding capacity (WHC), texture (assessed through texture profile analysis andspreadability), and microstructure observation using scanning electron microscopy.Subsequently, the optimum formulations was selected for storage study for a period of

30 days at 4°C and 25°C to evaluate their storage

Based on the result, the emulsion gel with a ratio of 75% PS and 25% CSexhibited favorable physical properties At this ratio, three different concentrations ofplant-based starch (10, 15, and 20%) also yielded satisfactory results after one night ofstorage The water-holding capacity and textural properties of the emulsion gelcontained 15% of plant-based starch showed no significant difference (p>0.05) whenstored at 4°C However, the result indicated that a significant decrease in the physicalproperties of the emulsion gel stored at 25°C (p<0.05), especially WHC and texturalproperties These findings justified the potential of CS and PS complexes in formingemulsion gel and their promising applications in various food sectors Nevertheless, it

is noteworthy that this research did not employ any preservatives during the storageperiod, leading to biochemical changes in the samples that could affect their physicalproperties Therefore, the addition of preservatives is recommended for further research

Keywords: Red palm oil; Corn starch; Potato starch; physical properties; Emulsiongel; Whey protein isolate; Water holding capacity

CHAPTER 1: INTRODUCTION

1.1 Introduction

Recently, there has been a growing demand for reduced-fat food products,

driven by the need to address diet-related diseases such as diabetes and obesity Within

the food industry, emulsion gels play a significant role in creating desired texture or

sensory experiences for fat-reduced food products, as well as serving as vehicles for

delivering functional food ingredients via encapsulation Emulsion gels combined

elements of both emulsions and hydrogels, forming semi-solid systems with a gel

network structure that typically contains dispersed oil droplets These gels exhibit

gel-like characteristics, including: (i) a high concentration of oil droplets of closely packed;

(11) strong attractive interactions between these oil droplets; and (iii) the presence of a

three-dimensional network within the aqueous phase (Hu, Li, Tan, McClements, and

Wang (2022))

Animal fats contain many medium-chain saturated fatty acids (SFAs) and

cholesterols, which may cause metabolic disorders if consumed beyond the daily

requirement Consequently, substituting of animal fats with vegetable oils in meat

products has gained popularity Vegetable oils offer monosaturated fatty acids

(MUFAs), polyunsaturated fatty acids (PUFAs), and fewer SFAs Palm fruit stands out

as one of the most economically significant crops in Malaysia, Indonesia, and Thailand

because of its high productivity and the excellent functional properties of the oil (Tan

et al., 2021) Red palm oil (RPO) is particularly noteworthy as a rich source of

increasingly focused on the antioxidant properties of carotenoids (as provitamin A) and

Vitamin E in RPO The fatty acid composition of RPO is well-balanced, comprising

approximately 50% saturated fatty acids, 40% monosaturated, and 10% polyunsaturated

fatty acids (Ayeleso, Oguntibeju, and Brooks (2012)) The distinctive characteristics of

RPO, including low levels of free fatty acids and high levels of carotene, particularly

beta- and alpha-carotene

As a natural polysaccharide, starch can be used as a thickening, texturizing,

gelling, and stabilizing ingredient in emulsions due to its many functional properties

There is a growing trend in utilizing starch as a fat replacer due to its ability to interact

with various components within a food system, including polysaccharides, lipids,

proteins, and low molecular weight ingredients such as sugar, flavor compounds, and

preservatives In this research, the combination of potato starch (PS) and corn starch

(CS) was used as the gelling agent for emulsion gel These starches are used for their

favorable thermal gelling properties, abundance, cost-effectiveness, and compatibility

with labeling requirements PS granules possess a large size, facilitating easy swelling

and gelatinization

Moreover, the gel produced from PS is characterized by its transparency and

lack of odor, qualities that are highly desirable for starch-based gelling agents in the

food industry Recently, many researchers have successfully applied PS in the

production of emulsion gels CS, on the other hand, are widely used as thickeners,

binders, and food additives in the food industry However, the application of regular

corn starch in emulsion gel production remains limited due to its high amylopectin

content, necessitating modification prior to gel formation (Zhang et al., 2023)

Generally, the preparation of a food-grade emulsion gel involves two essential

steps: emulsifying and gelling During the initial stage, the formation of an oil-in-water

emulsion necessitates the use of an appropriate emulsifier, such as whey protein isolate

(WPI) Then, the emulsions are gelatinized by gelling agents, achieved through

processes such as heating, cooling, acidification, salt addition, or enzyme crosslinking

In this investigation, the combination of CS and PS is used as the gelling agent to

enhance the quality of the emulsion gel

1.2 Aim

This study investigated the impact of the different concentrations and the ratios

of potato starch and corn starch on the physical (including water activity, water holding

capacity), texture and morphological-properties of the RPO emulsion gel In addition,

the storage stability of the optimum emulsion gel was also investigated

1.3 Objectives

e To investigate the impact of the concentration of starch on the physiochemical

properties of red palm oil emulsion gel

e To evaluate the storage stability of the optimized red palm oil emulsion gel using

a combination of plant-based starch over 30-day period at both 4°C and 25°C

e To investigate the optimal formulation and explore potential applications of

emulsion gel within the food industry

CHAPTER 2: LITERATURE REVIEW

2.1 Red palm oil-based emulsion

2.1.1 Red palm oil

Red palm oil is extracted from the fruit of the oil palm tree (Elaeis guineesis)

The oil palm is a perennial crop tree and has the highest oil yield compared with other

leading oilseed crops in terms of oil yield per hectare (Mielke, 2018) It has been

cultivated in Malaysia since 1917 and is now the most important economic crop in

Malaysia, with the export revenue from the crop reaching more than RM 67.12 billion

in 2018 (Parveez et al., 2020) Thailand and Indonesia are two more top producers of

palm oil

The oil palm is a unique crop that can produce two types of oil, namely palm oil

from the fibrous mesocarp (which has a brilliant, deep red-orange pulp) and palm kernel

oil (which resembles coconut oil) from the kernel (Sundram, Sambanthamurthi, & Tan,

2003) Red palm oil is obtained through the novel processes of pretreatment,

deacidification, and deodorization using molecular distillation, which allows about 80%

of the carotenes and vitamins present in crude palm oil to be retained (Nagendran,

Unnithan, Choo, & Sundram, 2000) Red palm oil has a distinctive red color because it

contains a high level of carotene and a low free fatty acid level Moreover, it is also rich

in other phytonutrients such as vitamin E, phytosterols, squalene, and coenzyme Q10

About 600 types of naturally occurring carotenoids are known, 12 to 13 of which are

found in palm oil: phytoene, phytofluene, 6-carotene, /-carotene, a-carotene,

cis-carotene, ¢-carotene, y-carotene, d-carotene, neurosporene, /-zeacarotene,

a-zeacarotene, and lycopene

Table 2.1 Health benefits of phytonutrients found in red palm oil

Anticancer, antiangiogenic, antioxidant, antiatherosclerotic,

cardioprotective, and neuroprotective properties; inhibits cholesterol

synthesis; aids in diabetes management

Provitamin A activity; cardioprotective and anticancer activity;

prevents night blindness

Cholesterol-lowering properties, anticancer activity, enhanced

immune function

Cardioprotective, radioprotective, and anticancer activity; inhibits

cholesterol synthesis

Enhances the production of cellular energy; antioxidative properties;

cardioprotective and anticancer activity

Adapted from (Loganathan et al., 2017)

2.1.2 Red palm oil extraction

The processing of oil palm fruit into edible oils involves many different and

complex steps Besides using traditional ways of processing, there are also applications

of small, medium, and large-scale mills (Poku, 2002) The processing steps of oil palm

fruit can be broken into a few steps: bunch reception, fruit sterilization, fruit digestion,

pulp extraction, and oil as shown in Figure 2.1 Bunch reception involves grading the

Faborode, & Ajibola, 2002) After the fruits have been graded, the sterilization process

will take place in the sterilizer Sterilization is a crucial step that inactivates and destroys

the enzymes to prevent free fatty acids (FFA) by using high-temperature steam This

process also softens and loosens the fruit structure for easier fruit digestion and

extraction of oil The mesocarp (flesh) and the kernel (seed) are separated in the digester

The steam-heated vessels with attached rotating shafts and a few stirring arms help

destroy the exocarp of the fruit and reduce the oil’s viscosity

Palm oil extraction has two common methods: the “dry” method and the “wet”

method The “dry” method uses mechanical presses such as hydraulic presses and screw

presses to extract the oil from the digested material The hydraulic presses are usually

used in the batch system, while the screw press is used in a continuous system more

often (Poku, 2002) The “wet” method, on the other hand, uses water to draw out the oil

from the fruit The hot water introduced to the fruit will break down gums and resins

that cause foaming of the oil during high-temperature frying The gums and resins will

soon be removed through the oil clarification process The mesocarp fiber will retain

about 5-6% of oil after the pressing (Obibuzor, Okogbenin, & Abigor, 2012) (Poku,

2002)

Oil clarification is to separate the impurities from the oil A mixture of oil,

water, and solids from the bunch fibers is transferred to the tank, and the separation of

the oil is based on the density of the materials Hot water is added to provide a barrier

to the lighter oil droplets and the heavy solids The oil droplets will stay at the top of the

tank, and the solids will sink to the bottom The crude palm oil (CPO) is decanted into

a reception tank and the moisture content is reduced to 0.15% to 0.25% to prevent FFA

increase through the autocatalytic hydrolysis of the oil CPO is subjected to

centrifugation for purification, followed by a drying step The purified and dried oil is

then transferred to the oil storage tank (Poku, 2002), (Obibuzor et al., 2012), (Hameed,

Ahmad, & Hoon, 2003)

Red palm oil can be obtained from the mild processing of crude palm oil while

refined, bleached, deodorized (RBD) palm oil is obtained by physical refining or

chemical refining of the crude palm oil (Nagendran et al., 2000) There are two stages

of processing for the refining of red palm oil from crude palm oil The first stage

involves the CPO’s pre-treatment, which uses phosphoric acid for degumming the oil

and treatment with bleaching clay The main purpose of the pre-treatment is to remove

the impurities in the CPO while retaining the carotenes The bleaching clay is removed

by filtration The next stage of the process is deacidification and deodorization The

pre-treated oil is passed through the short-path distillation unit at about 150°C to 170°C

under vacuum to remove the free fatty acids (FFA) without destroying the carotenes

(Choo et al., 1996)

Refined, Bleached and Deodorised Palm Oil Red Palm Oil

Figure 2.1 Simple processing flowchart of red palm oil and refined, bleached and

deodorized palm oil

2.2 Emulsion

Emulsions are colloidal dispersions that consist of at least two immiscible fluids

(normally water and oil), with one of them being dispersed in the other in the form of

small droplets (McClements, 2004) Emulsion science and technology principles are

commonly employed in the food industry to create a wide variety of emulsified food

products, such as beverages, milk, creams, dips, sauces, desserts, dressings, mayonnaise,

margarine, and butter The nature of emulsions confers these foods with distinct

functional attributes, such as desirable appearances, textures, mouthfeels, and flavor

profiles Moreover, emulsions are a widely used vehicle for the encapsulation and

delivery of bioactive agents, such as vitamins and nutraceuticals (C Tan & McClements,

2021)

Food emulsion consists of an oil phase containing hydrophobic compounds and

an aqueous phase containing water-soluble components One is dispersed into the other,

defined as oil-in-water (O/W) emulsions or water-in-oil (W/O) emulsions, in which the

aqueous solution and oil are the continuous phase, respectively Moreover, they also

have another type which is water in oil in water (W/O/W), which is, in effect, an O/W

emulsion whose droplets themselves contain water droplets (Friberg, Larsson, &

Sjoblom, 2003) They are very complex in composition and structure Besides lipids and

water, they contain proteins, polysaccharides, small surfactant molecules, and molecular

and ionic solutes (sugar, alcohol, salts, preservatives, colorants, flavorings, etc.).The

distribution and arrangement of these components, determined in part by their respective

chemical affinity, are such that they permit a partial reduction of the system's free

energy Some proteins and surfactants (amphiphilic compounds) are in the O-W

interface Water-soluble polysaccharides are mainly solvated in the continuous phase,

contributing to their viscosity, which affects product stability At the O/W interface,

solid particles (such as caseins in milk fat globules), and small surfactant molecules or

proteins arranged in mono or multi-layers may be found Emulsions can also be

classified such; nanoemulsions, high internal phase emulsions (HIPEs), Pickering

emulsions, multilayer emulsions, solid lipid nanoparticles, and multiple emulsions (C

Tan & McClements, 2021)

2.3 High-Pressure Homogenization Process.

High-pressure homogenization (HPH) is used in the food industry frequently,

due to the high efficiency in industrial scale up (Yihong Wang et al., 2015) This has

been employed for unit operations like comminution, mixing, and stabilization of

pharmaceutical solids and nanoparticles (Vinchhi, Patel, & Patel, 2021) In this process,

fluid materials are subjected to high pressure through the narrow homogenization gap at

high speed The shearing forces from the process of HPH can bring about molecular

refinement (Martinez-Monteagudo et al., 2017) HPH has the distinct advantage of being

one of the most versatile and scalable processing methods for the preparation of different

vesicular and non-vesicular lipid-based nanosystems such as nanoemulsions, solid lipid

nanoparticles (SLNs), nanostructured lipid carriers (NLCs), nanocrystals, as well as

polymeric nanoparticles

HPH is a widely applied food technology in protein modification, milk

homogenization, beverage preparation, and emulsion preparation When the material

quickly passes through a cavity with a special internal structure, the material is subjected

to mechanical forces such as high-speed shearing, high-frequency oscillation, cavitation

effect, and convection impact (Dumay et al., 2013) The large emulsion droplets could

be broken into small ones under these mechanical forces HPH is the most used

emulsifying method to prepare a protein-stabilized fine emulsion

In a high-pressure homogenizer, the oil and water mixture is subjected to intense

turbulent and shear flow fields Turbulence is said to be the predominant mechanism

(Walstra, 1975), even though laminar shear and cavitation may also play an important

role Turbulence leads to the breakup of the dispersed phase into small droplets The

relative motion between the drops results in their collision, leading to their coalescence.

There is usually a dynamic equilibrium between breakage and coalescence The shelf

life, as well as the texture of the emulsion, greatly depends on the drop size distribution,

which can be adjusted by controlling the rate of drop breakage and coalescence during

emulsion formation (Floury, Desrumaux, & Lardiéres, 2000)

2.4 Emulsion gel

Food emulsion gels are ubiquitous in the food industry to create texture or a

sensory experience for low-fat products or as a vehicle to deliver functional food

ingredients (Farjami & Madadlou, 2019), (Guo, Cui, & Meng, 2023) Emulsion gels are

semi-solid systems with a gel network structure, often embedded with oil droplets

(Dominguez, Munekata, Pateiro, López-Fernández, & Lorenzo, 2021) They integrate

the dual characteristics of emulsion and gel, improving both the stability of the emulsion

system and the rheological and nutritional properties of a hydrogel, making them a

unique and versatile format for developing new foods (Yong Wang & Selomulya, 2022)

The difference between an emulsion gel and a simple emulsion with oil droplets as the

inner phase lies in the presence of the gel network In an ordinary emulsion, oil droplets

are dispersed in a continuous phase with emulsifiers to lower the interfacial tension

between the two immiscible liquids However, in an emulsion gel, the dispersed oil

droplets are not only stabilized by emulsifiers but are also trapped within a continuous

gel network, which provides additional stability and unique characteristics to the system

(Dickinson, 2012)

Emulsion gel systems were originally applied to reduce the fat content of food

products by incorporating the gelled water phase while still retaining sensory properties

(C Tan & McClements, 2021) Emulsion gels can be used to replace fats in a variety of

food products, including baked goods, processed meat, dairy products, functional foods,

and edible 3D printing inks (Yu, Wang, Li, Wang, & Wang, 2022) In addition to their

potential as a fat replacer, emulsion gels show promising prospects as a delivery medium

for functional ingredients through encapsulation They can effectively inhibit the release

of the ingredients and improve the efficiency of bioactive substances, while also

controlling the release rate of the encapsulated ingredients According to research, the

use of a protein-polysaccharide for encapsulation may yield better results than systems

that utilize only protein or polysaccharides (Mao, Lu, Cui, Miao, & Gao, 2020)

The composition of an emulsion gel is fundamental to the stability and physical

and mechanical characteristics of the product Emulsion gels are typically differentiated

by the material(s) from which they derive their structure Emulsion gels may be formed

by both proteins and polysaccharides These hydrocolloids may be used in isolation or

in combination to stabilize oil-in-water emulsions and provide structure (Yiu et al.,

2023) A summary of major techniques for emulsion gel formation and mechanism is

presented in Figure 2.2 (Adapted from (Yiu et al., 2023))

Plant-based protein/polysaccharide

stabilized emulsion Mixed formulation

Typical plant protein

sources for emulsion

gels: Addition of polysaccharide

* Chickpea k C Lipid phase

+ Fava

- Pea nO Aqueous

° Soy oO phase

+ Lentils Enzyme-induced cross- Heat-induced © Plant-based

- Salt/acid- linkage (Protein) or emulsifier

Heat-induced induced Cold-set (acid/salt-induced)

@ saltiAcia Emulsion Gel @ Plant-protoin

% Plant polysaccharide

Heat to above gelation E.g.GDL, lactic acid, E.g Bacterial transglutaminase, E.g SPI, PPI/Starches, KGM, Guar

temperature CaSO, MgSO, tyrosinase gum

- Formation of continuous - Cold-set - Cold-set ~ Polysaccharide-dependent gel properties

protein fibrils structure/ - Non-covalent interactions - Covalent bonds between protein (eg thermal-irreversibility, gel strength)

polysaccharide polymerization between macromolecules - Typically stronger than heat- or acid

induced by salt/acid induced gels

Figure 2.2 Summary of various methods, in which plant-based proteins,

polysaccharides, and lipids are used in the formation of plant-based emulsion gel

Generally, emulsifying and gelling are the two essential steps to prepare a

food-grade emulsion gel (R Wang & Duan, 2023) Specifically, the oil-in-water emulsions

are prepared first through emulsifying using emulsifiers Emulsifiers including small

molecular surfactants, amphiphilic biopolymers, and colloidal particles have interfacial

activity so that they can absorb the oil-water interface and form an interfacial layer,

thereby inhibiting the coalescence of the dispersed oil droplets (Berton-Carabin &

Schroén, 2015) Then, the prepared emulsions are gelatinized by gelling agents through

heating, cooling, acidification, salt addition, or enzyme crosslinking (Hu et al., 2022)

Gelling agents, such as proteins and polysaccharides, can gelatinize the continuous

phase of the oil-in-water emulsions to make the liquid emulsions transform into

semi-solid state, blocking the migration of oil droplets and thus inhibiting the coalescence of

Heat treatment: In the process of emulsion gel formation, heat treatment is the

most common method Heat treatment can denature proteins, which would aggregate

and form three-dimensional structures through chemical forces (i.e., disulfide bonds,

electrostatic interactions, hydrophobic interactions, hydrogen bonds, and ionic bonds)

under appropriate conditions (e.g., concentration, pH, and ionic strength) (Guo et al.,

2023) In the gel formation process, the emulsion is first heated and then placed at a low

temperature to cool to form a gel The heating process makes the protein molecules fully

expand and denature, and the sulfhydryl and hydrophobic groups are fully exposed and

then form a network gel structure through hydrogen bonding and hydrophobic

interaction (Spotti, Tarhan, Schaffter, Corvalan, & Campanella, 2017)

Enzymatic treatment: The advantage of enzymatic cross-linking is that it does

not cause unpleasant odors and can preserve essential nutrients Furthermore, reaction

conditions of enzymatic treatment are mild, as heating, change in pH, and/or ionic

strength are not required (de Souza Paglarini et al., 2020)

Salt treatment: Salt (e.g., Ca”', Mg”') treatment coagulation mechanism of

calcium sulfate is complicated, the addition of calcium ions not only neutralizes surface

charges but also forms salt bridges between protein aggregates, which leads to the

formation of a three-dimensional network (Lu, Lu, Yin, Cheng, & Li, 2010), (Zhao, Li,

Qin, & Chen, 2016) And salt-induced emulsion gels usually combine preheating to

expose the hydrophobic groups, and then the protein molecules gradually aggregate to

form a stable system under electrostatic attraction After that, the pre-denatured proteins

are induced to aggregate to form fibrous or granular gels by adding salt ions (Bryant &

McClements, 1998)

2.5 Plant-based starch

Starch is a “green” polymer widely used in foods It is a native polysaccharide

that has a variety of functional attributes for utilization in emulsions, including

thickening, texturing, gelling, and stabilizing agents (Alcazar-Alay & Meireles, 2015)

More and more studies have used starch to make food emulsion gels due to its abundance

in quantity, low cost, high biocompatibility, non-allergenicity, modifiability, and

eco-friendliness (Shao, Zhang, Niu, & Jin, 2018) Relevant studies have proven that starches

of various biological origins, such as potato, taro, acorn, wheat, and corn, can also be

used as emulsifiers to stabilize emulsion gels In addition, Starch is naturally present as

semi-crystalline granules composed of two biopolymers, amylose and amylopectin

Amylose has an essentially linear structure, some with a few branches; whereas

amylopectin is a highly branched a-glucan with 5% of a-(1-6) branch linkages When

heated in excess water, the granular structure of starch is irreversibly lost, leading to the

release of amylose (Zhang et al., 2023) The linear structure of amylose enables the

formation of helical structures in the aqueous systems, and the hydrophobic cavity of

the helix can encapsulate lipids (Oyeyinka, Singh, & Amonsou, 2021) Specifically, the

formation of starch-lipid complexes has been demonstrated to be an effective way to

modify the functionality of starch, e.g., reducing the solubility and swelling power,

increasing thermal stability, and retarding retrogradation (Oyeyinka et al., 2021)

2.5.1 Potato starch

Potato starch is used in a wide variety of products, as a food ingredient or as an

industrial material The total annual world production of all starches was approximately

source of carbohydrates, potatoes also contain significant amounts of protein, ascorbic

acid, and other vitamins, phenolic substances, and minerals such as phosphorus,

potassium, and calcium (Kadam & Wakier, 1991) The chemical composition and

structure of potato components, such as starch, non-starch polysaccharides, sugars and

other carbohydrates, organic and inorganic compounds, and proteins influence the

quality of potatoes and potato products Potato starch is a mixture of two

polysaccharides, amylose and amylopectin Amylose is a relatively long, linear a-glucan

with few branches, containing approximately 99% a-(1,4) linkages and up to 1% a-(1,6)

linkages, whereas amylopectin is a heavily branched structure, containing

approximately 95% a-(1,4) and 5% a-(1,6) linkages in a hierarchical structure (Dupuis

& Liu, 2019) Potato starch from various cultivars typically contains 20-33% amylose

on a total starch basis, with the balance being amylopectin (Dupuis & Liu, 2019)

Potato starch can hydrate quickly and form a paste with higher viscosity when

heated, and a clearer gel when cooled, which differs from those of other starches such

as corn, wheat, and rice (Xu et al., 2021) This can be gelatinized in hot water by

breaking the intermolecular hydrogen bond and destroying the arrangement of the

micelle structure in starch granules After being fully gelatinized, the starch granules are

porous with broken hydrogen bonds, birefringence disappears, and the crystalline order

is lost through rapid drying at high temperatures (Han et al., 2019) The natural

properties of potato starch are exploited by the food industry to provide products with

the required texture, appearance, density, and storage stability According to previous

reports, high amounts of phosphate groups in potato starch tend to enhance the starch

paste viscosity, high swelling power, and increased paste clarity and extent of

retrogradation (Karim et al., 2007) However, native potato starch has some drawbacks,

such as poor solubility and poor stability against heat and shear during pasting

Digestibility may be reduced by complexing a hydrophobic compound (most

commonly a fatty acid or lipid) into an amylose helix Potato starch forms complexes

with lipid tails of 10 to 22 carbons with varying degrees of unsaturation (Tufvesson,

Wahlgren, & Eliasson, 2003) Potato starch has a highly crystalline nature, minimizing

penetration of the complexing group into the granule Thus, complex formation is best

performed on starches in which the granular order is altered (swelled) or destroyed

(gelatinized starch), giving the complexing group better access to the amylose chains

Amylose-lipid complexes prevent staling and improve crumb texture in gluten-free

bread as well as replace fat in products

2.5.2 Corn starch

Corn provides a high-quality starch used widely in the food industry in many

applications requiring particular viscosities and textures Basic corn starches have a

small amount of protein (0.35%), lipid (0.8%), ash, and >98% of two polysaccharides,

namely amylose and amylopectin (Palanisamy, Cui, Zhang, Jayaraman, & Kodiveri

Muthukaliannan, 2020) Corn starch is composed of two _ polymers

(homopolysaccharides), amylopectin and amylose, which differ in their chain length and

degree of branching Amylopectin is more highly branched (a chain of D-(1-4) and

a-D-(1-6)-glucosidic linkages) and normally constitutes about 75% of the starch granule

Amylose is mostly linear (a-D-(1-4)-linked glucose residues) and constitutes 25% of the

starch granule (Jeon, Ryoo, Hahn, Walia, & Nakamura, 2010), (Nelson & Pan, 1995)

amylopectin plays a key role in the appearance, structure, and quality of industrial

processing

The difference among corn starches in granule swelling (onset of viscosity),

peak temperature, peak viscosity, shear thinning during pasting, and gel firmness during

storage, have been mostly attributed to the difference in amylopectin structure (Ring &

Stainsby, 1985), whereas differences in setback and final viscosity during pasting have

been attributed to amylose structure The preparation of starch-lipid complexes mainly

utilizes high amylose starch as substrate, because the limited chain length and branched

structure of amylopectin would restrain the formation of the complexes (Zhang et al.,

2023)

CHAPTER 3: MATERIALS AND METHODS

3.1 Materials

Red palm oil was purchased from a local food ingredient supplier (online store,

Selangor, Malaysia) Both potato and corn starches (Bestari, Synerchem Sdn Bhd,

Selangor, Malaysia) were purchased from Aeon Mall (Taman Equine, Selangor,

Malaysia) BiPRO whey protein isolate (WPI) was obtained from Davisco Foods

International Inc (Eden Prairie, MN) Deionized water was used throughout the study

WPI Red-palm oil Potato starch Corn starch

3.2 Methods

3.2.1 Red palm oil-based emulsion preparation

An aqueous phase containing 2 wt% WPI was prepared by dissolving the

powdered protein into deionized water and stirred using a magnetic stirrer (MR

Hei-Standard, Heidolph, USA) at 750 rpm overnight at room temperature The oil phase (red

palm oil) and aqueous phase (WPI solution) were then mixed at a ratio of 1:9 (w/w)

using a high-shear homogenizer (Silverson-L4RT, Silverson Machines Inc., MA, USA)

operating at 7000 rpm for 5 min to form the coarse emulsion Subsequently, the coarse

emulsion was subjected to high-pressure homogenization (PandaPLUS 2000, GEA Niro

emulsions These emulsions were later on used to prepare the red palm olein

starch-based emulsion gels

3.2.2 Preparation of starch-based emulsion gel

WPI-stabilized emulsions were added with different concentrations (10, 15, and

20%) of potato and corn starches at ratios of 0:100, 25:75, 50:50, 75:25, and 100:0 The

resulting emulsion-starch mixture was continuously stirred and heated in a water bath at

90°C for 15 min to gelatinize the starch granules The mixture was then stored overnight

at 4°C to form emulsion gels

3.2.3 Investigation of the different formulations for red palm oil starch-based

emulsion gel

3.2.3.1 Gel appearance (Tube Inversion Method)

The emulsion gel samples were poured into a serum bottle After the samples

cooled down to room temperature, the self-sustaining (gel-forming) ability of the

samples was assessed visually by inverting the serum bottle Samples were categorized

as a gel, thickened liquid, or liquid based on their appearance (Fayaz et al., 2017)

3.2.3.2 Texture characteristic

3.2.3.2.1 Texture Profile Analyzer (TPA)

This analysis was performed using the TA-XT Plus Analyzer (Stable

Microsystems Ltd., Surrey, UK) at room temperature (25°C) The emulsion gel was first

filled in a container The hardness of the sample was then determined using a cylindrical

probe (P/10 with 10 mm) The measurements were carried out using the compression

mode, in which the probe penetrated to a depth of 15 mm into the sample (at a rate of 2

mm/s) and retreated at the same pace In addition, the gumminess, chewiness, and

adhesiveness of the samples were also measured

3.2.3.2.2 Spreadability

For the spreadability test, a male 45° cone probe and a container were used

Before the test, emulsion gel samples were filled into a container and placed on the

platform of the instrument The sample was compressed using the 45° cone probe, which

shifted by a distance of 10 mm at a test speed of 3 mm/s The probe then retreated to its

initial position at a speed of 10 mm/s

3.2.3.3 Water holding capacity

An empty 50 mL centrifuge tube was filled with 2.5 g of emulsion gel After

that, the mass of the centrifuge tube and gel was weighed and recorded as Mo The

sample was centrifuged at 8,000 x g for 15 min, and expulsed moisture was dried using

filter paper The centrifuge tube and sample were then weighed and its mass was

recorded as Mi The WHC was then calculated based on the following formula:

Mo—MnWHC (%) = (1 — “°”) x 100

where M; is the mass of water in the original sample

(Feng, Jia, Yan, Yan, & Yin, 2021)

3.2.3.4 Water activity

Before measuring water activity, the emulsion gel was first poured into the

sample cup Then, the water activity was measured by the water activity analyzer ( Pre

3.2.4 Investigating the selected optimum emulsion gel during storage

The selected optimum formulation was freshly prepared and kept at 4°C and

25°C for 30 days to characterize its storage stability The gel's appearance, texture

characteristics, water-holding capacity, water activity, and microstructure will be

analyzed at the beginning and the end of the storage period One of the sample of the

optimum fomulation was ultrasonicated for 15 minutes in a sonicator (Powersonic 420,

Hwashin Technology Co., Ltd., Korea), with an ultrasonic power of 700W This sample

was used to compare with the sample without ultrasonic after 30 days of storage

3.2.4.1 Microstructure

The microstructure of the emulsion gel was analyzed using scanning electron

microscopic (SEM) The sample was prepared around 2-3g then put in the plate After

that put in the SEM machine (JSM-IT 100 InTouchScopeTM) to take the micrograph of

the sample The micrograph of the sample was taken at the pressure of 60Pa, and

magnification of x1000 The best pictures were selected

3.2.5 Statistical analysis

All measurements were performed in duplicate on each replicated sample

Two-way ANOVA was used to test the significant diffeence between samples, with

significant differences established using Turkey’s test at p < 0.05

CHAPTER 4: RESULT AND DISCUSSION

4.1 Investigation of the different formulations for red palm oil starch-based

The visual appearance of these emulsion gels was ofa yellow color because of

the red palm oil-based emulsion The appearance of emulsion gel was affected by the