LIST OF FIGURES Figure Caption Page 1

Figure Caption Page
1.1 Molecular Structure of Starch
1.2 Molecular Structure of Succinic Acid
2.1 Molecular Structure of Amylose
2.2 Molecular Structure of Amylopectin
3.1 Flow Diagram for Preparation of Succinic Acid Modified
3.2 Flow Diagram for Synthesis of Succinic Acid Modified Sago
Starch Hydrogel
Starch is widely used in food industries as a thickening, stabilizing and gelling agent. While, hydrogel have biocompatible and biodegradable properties where usually applied in field of medicine, pharmaceutical and other industrial. Native starch is insoluble in water can undergo a process through chemical process to form a modified starch which soluble in water.


Figure 1.1 : Molecular Structure of Starch
The main source of starch are plant cells such as sago, potato, wheat, tapioca, rice and corn. The formations of starch in the plants occur in daylight during the process of photosynthesis from carbon dioxide and water, which produce glucose in a form of starch to the cell plant. The starch usually stored in the granules of green leaves, seeds, pulses and tuber. The starch usually stored in the granules of green leaves, seeds, pulses and tuber. The starch present in two components; amylose and amylopectin. Both of these components are stored in the granules (Yoshimura et al., 2006).

Both components of starch are connected by a glycosidic linkage shown in the Figure 1.1. The glycosidic linkage is the bond that joins the unit of the glucose with one to another. In general, the starch is insoluble in cold water or in alcohol. By modification process, the starch will be able to soluble in water. According to Yoshimura et al.(2006), the starch and succinic acid anhydride undergo a process of esterification which resulted in formation of hydrogel by modification of starch.

In past year, the application of hydrogel has been committed to drug delivery, clinical application, as well as scaffolds for tissue engineering and regenerative medicine. Regarding to the increase of attention for the hydrogel, it was a material applied for regenerative application due to ability to entrap large amount of water, good biocompatibility and the ability to mimic tissue environment (Barbucci,2005).

In Malaysia, sago starch flour was applied in production of many various kind of food. Sago starch mixed with water may be baked to make bread or pancake; where in Sarawak, local biscuits tabaloi and lempeng were produced while in Peninsular Malaysia it was used as fish crackers and keropok lekor by mixing sago flour with fish meat (Mohamed et al.,2008). It shows that sago starch normally used as functional ingredient in food industry.

Hydrogel is hydrophilic natured there-dimensional network. Regarding to this property, it has the ability to absorb large amount of water or saline solution (Yoshimura et al., 2006). The starch hydrogel prepare by esterification process and will be investigated in this study to characterize it properties.


Figure 1.2 : Molecular Structure of Sucinic Acid
Succinic acid also known as amber acid with IUPAC name butanedioic acid was used in this study. Succinic acid is dicarboxylic acid with four carbon atoms (Figure 1.2). This acid present naturally in plant and animal tissue especially it is important to play a main role in the citric acid cycle or mostly familiar with the Kreb’s Cycle. Moreover, succinic acid is soluble in water but it is dissolve in ethanol, ether and does not dissolve in benzene.

1.2Objective of Study
To synthesize succinic acid modified starch
To characterize the succinic acid modified starch hydrogel properties, morphology, structural analysis, swelling behaviour and solubility
In this study, sago starch was used. This starch produced great abundance in South-East Asia where it has been applied in food and non-food formulations. It was derived from the pith of certain palm trees especially Metroxylon species (Fakirov and Bhattacharyya, 2005) especialy M.sagu and M.rhumpii (Ahmad and Williams, 1999). Sago starched is produced from CYCAS, the pith of the cycads of genus. It is also known as rumbia where this is important resource for different uses especially in production of starch as sago flour or sago pearl (Mohamed et al., 2008).

Starch consists of glucose which linked with ?-D-(1-4) or ?-D-(1-6) linkages. There are two types of structural component which are amylose (Figure 2.1) and amylopectin (Figure 2.2). Amylose usually in linear or slightly branched has lower molecular weight and degree of polymerization. In the other hand, the amylopectin is the largest molecule present in nature ( Sajilata et al., 2006).

Both amylose and amylopectin from sago starch with low (280 BU,low-v) and high (735 BU, high-v) viscosities on amylography differed in molecular structures . The degree of polymerization of low-v amylose was 5090, which was half those of high-v amylose. Both of starch granules had the same crystalline type and amylose content with 24% (Takeda et al., 1989).


Figure 2.1 : Molecular Structure of Amylose

Figure 2.2 : Molecular Structure of Amylopectin
Starch usually become the reactive carbohydrate if they undergo modification. Based on Erickson (2006) , starch has four physicochemical properties. They are thickening, ability to disperse and suspend, gel formation and ability to produce strong adhesive films. This properties are depending on the type of starch that was used.

In addition, starch can form a gelatin as starch granules are heated in water. This process is called gelatinization process where the reaction allows water to enter into the starch molecule. During water is heated, the hydrogen bonding within starch molecule becomes weak causing them to swell until their thickness is reached (Brown, 2008).

2.2 Modified Starch.

Starch has been modify to give variety of modified starch leading their usefulness in food processing and other industry. The modification of starch refer to starch that has been chemically or physically modified to create unique functional characteristics (Brown, 2008).

The modified starch can be divided into three types : cross-linked starch, oxidized starch, and instant or pre-gelatinized starch.

The acid modification of starch granules in alcohol influence and give an impact to the granules by changed the temperature, acid concentration and starch concentration (Robyt et al., 1995).From their studies, they found that the starch granules present a different morphology of starch granule with different kind of condition. The reaction on modification of starch granules involved the hydrolysis of glycosides bond where water present in the granule.

A series of starch ester has been prepared and studied by Fang et al. (2001). Where they modified a potato starch with a different degree of substituent and side-chain lengths. The acylation process are used to develop acylated starch by controlled free substituent or hydroxyl group in starch. From their studies, they found that lot of properties may be prepared to modify fatty chains, temperature and degree of substituent.

2.3 Starch Ester
Esterification is a condensation process where the reaction involved between the carboxylic acid with alocohl to form an ester. In this study, the synthesis of starch ester will be done first before further study continue to form a starch hydro gel.

The previous studies by Hui et al . (2008) showed that the potato starch can be prepared by esterification process with the octenyl succinic anhydride (OSA). They stated that the modification of starch by OSA may cause physical change especially to the crystalline pattern of potato starch. By their research, they stated that the hydrophobic side chain was added to the hydrophilic starch. These prove that the starch may absorb to the interface of water and oil for stabilizing the emulsion.

According to Zhou et al. (2009) , the surface esterification of corn starch films by reaction with dodecenyl succinic anhydride (DDSA) showed that starch films can be modified through the esterification process . Based on their study, esterification process with DDSA shows that the thermoplastic starch product can be improved without changing the bulk composition and properties.

2.4 Starch Hydrogel.
Hydrogels are water-swollen polymeric materials that maintain a distinct three-dimensional structure. They were first bio-materials design synthesis by traditional method include cross linking co-polymerization, cross-linking of reactive polymer pre-cursors and cross linking via polymer-polymer reaction. These hydrogel have potential applications especially in tissue engineering, synthetic extracellular matrix, implantable devices, bio-sensors and other (Barbucci, 2005).

Yoshimura et al., (2006) reported on synthesis and characterization of bio-degradable succinic anhydride modified starch hydrogels . In their studies, the succinic anhydride undergo an esterification process where the dimethylsulfoxide (DMSO) is used as solvent starch and catalyst 4-Dimethylaminopyridine followed with the neutralization step by sodium hydroxide. On their previous study, they are investigated a novel biodegradable hydrogels by the esterification process but using a cotton cellulose or chitin with succinic anhydride as catalyst. They showed that the hydrogel can be obtained without any specific cross linking agent.

2.5 Swelling Behaviour.

Swelling is the process that occurs when starch is heated in present of water and hydrogen bonds stabilizing the structure of the double helices in crystallites are damaged and substituted by the hydrogen bonds of water (Tester and Karlakas, 1996) It occurs in the presence of enough water, the starch granule swells and its volume was increased during the heating process.

Vandeputte et al . (2002) investigated the structural aspects provide insight into swelling and pasting properties on rice starch. It has been found that short amylopectin chains with the degree of polymerization 6-9 led to increase swelling power and decreased close packing concentration at 55 0C and 65 0C where longer amylopectin chain with degree of polymerization 12-22 had the opposite effect.

Fabian et al (2010) also found that the swelling power of the starch gradually increased at temperature range 250C to 60 0C followed by very sharp increase at 80 0C to 90 0C. It shows that the branched polymer, amylopectin swells more extent with protein as individual component while amylose the linear polymer is more resistant to swelling.


Sago starch brand OPAC will be use for this study, Succinic acid (SA) supply by fisher Chemicals and absolute ethanol will be apply for preparation of the succinate starch hydrogel. The ultra pure will be water use to prepare succinate starch hydrogel (Water Purifying system, Ultra Gentic, Model: ELGA , UK).

3.2 Methods
3.2.1 Preparation of Succinic Acid Modified Starch.

The reactions will be carried out between the reactions of sago starch and succinic acid. First, sago starch will be added into the water to form a gelatinized starch solution. The mixture will be stir and sodium hydroxide solution will be added as a catalyst. Then, the mixture will be heated in the water bath at 80 0C for 10 minutes. Then, succinic acid will be added and the heating process continued for another hour (Pang et al, 2010). The parameters of preparation of succinate starch are reaction time and amount of acid.

1191491261505Corn Starch + Ultra Pure Water
Adding 3 mL NaOH, 1.0 M.

Heat at 80 °C for 10 min in water bath under magnetic stirring
00Corn Starch + Ultra Pure Water
Adding 3 mL NaOH, 1.0 M.

Heat at 80 °C for 10 min in water bath under magnetic stirring

11430007562Gelatinized Starch Solution
Adding 20 mL amount of succinic acid
00Gelatinized Starch Solution
Adding 20 mL amount of succinic acid

1170709402936Succinic Acid Modified Starch
Heat at 80 °C for 8 hours in water bath
00Succinic Acid Modified Starch
Heat at 80 °C for 8 hours in water bath
1177636215438Succinic Acid Modified Starch Solution
00Succinic Acid Modified Starch Solution

Figure 3.1 : Flow Diagram for Preparation of Succinic Acid Modified Starch
3.2.2 Synthesis of Succinic Acid Modified Sago Starch Hydrogel
Succinic acid modified sago starch hydrogel can be obtained where succinate starch obtained by previous step will be suspended to form a hydrogel by dropping the solution into absolute ethanol. After ethanol added, the mixture will be centrifuged to obtain the starch hydrogel.

1634836151245Succinic Acid Modified Starch Solution
Succinic Acid Modified Starch Solution

right345786Precipitate with absolute ethanol
Precipitate with absolute ethanol
14962333804978Characterization :
SEM : Morphology
FTIR : Functional Group
00Characterization :SEM : Morphology
FTIR : Functional Group
16621992821305Characterisation of Succinic Acid Modified Starch Hydrogel
Characterisation of Succinic Acid Modified Starch Hydrogel
16344901879196Purification Succinic Acid Modified Starch Solid
Purification Succinic Acid Modified Starch Solid
1634490916651Suspension of Succinic Acid Modified Starch Hydrogel
Suspension of Succinic Acid Modified Starch Hydrogel

Figure 3.2 : Flow Diagram for Synthesis of Succinic Acid Modified Sago Starch Hydrogel
3.2.3 Characterization of Succinic Acid Modified Starch Hydrogel Functional Group Analysis.

The functional groups of the samples will be examined by using FT-IR spectroscopy (SHIMADZU Model FTIR-8201 PC). The sample in powder form will be mixed with dry potassium bromide (KBr) by ratio of 1:100 to from the KBr pellet. Then, the pellet was scanned through FT-IR at the wave number in the range between 400 and 4000 cm-1 . Morphology Studies
The morphology of native sago with starch hydrogel will be characterized by Scanning Electron Microscopy SEM (JEOL Model JSM 6390 LA) at different magnifications. The sample in form of powder will be coated onto a stainless steel and the morphology will be observed by a scanning electron microscope. Swelling Measurements
The swelling could be described as water absorptivity of starch hydrogel based on equation below:

The sample will be immersed in water for 24 hours at room temperature until the hydrogel reached the equilibrium state of swelling. Based on Paranhos et al. (2007) in their study, it is confirmed that 24 hours of equilibration for the absorption in water inside the network of the solid and the formation of swelled sample. Then, the water on the surface of the swollen gel was removed by tissue paper and weight of the sample was recorded. The wet of hydrogel sample was also recorded at specific time (Tchong, 2009). Solubility Test
The solubility of succinic acid modified starch hydrogel in water will be studied at different weight of sample used. The dry succinate starch hydrogel samples will be weighed before immersed into 20 mL of water. The samples will be immersed under magnetic stirrer and the time will be taken for sample completely dissolved in water will be recorded.