McGill University

Faculty of Agricultural and Environmental Science Department of Food Science and Agricultural Chemistry Separation Techniques 2 FDSC 652

Differential Scanning Calorimetry as Analytical Tool

Seminar Coordinator:

Dr. Benjamin K. Simpson

By:

Chia-Ning Chang

McGill ID: 260440069

April 4th, 2012

TABLE of CONTENT 1. Introduction

2. Principles of DSC

3. Application of DSC in Food Analysis 3.1. DSC of Lipids 3.2. DSC of Starch 3.3. DSC of Protein 3.4. Thermal properties of frozen foods 3.5. food microbiology

4. Limitations and prospects

5. Reference

-2-

1. INTRODUCTION Differential scanning calorimetry (DSC) is a technique we use to study what happens to a substance when it is heated; a tool for observing thermal transitions. It is defined as the measurement of the difference in the heat flow rate to the sample and to a reference sample while they are subjected to a controlled temperature program (Hhne, Hemminger, & Flammersheim, 2003). The term differential scanning calorimetry was coined to describe the ability of the instrument to measure energy directly and allows precise measurements of heat capacity. This technique is first developed by E.S. Watson and M.J. O'Neill in 1962 and introduced commercially at Pittcon (Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy) in 1963. Later in 1964, P.L. Privalov and D.R. Monaselidze successfully developed the first adiabatic differential scanning calorimeter to be applied in biochemistry.

At present, DSC is the most widely applied thermal analytical technique in food research and quality assurance as it has the advantages of providing direct estimate of the overall enthalpy change of transitions in food requiring merely basic knowledge of the thermodynamic mechanism and minimum sample preparation requirements. For example, it allows the analysis of a piece of intact meat or plant tissues. Furthermore, it can simulate the heating conditions of a thermal processing procedure to certain food which permits us to examine the effects of different heating patterns on food. In the meanwhile, DSC may serve the function as a quality control tool to avoid excessive processing (Farkas & Mohcsi-Farkas, 1996).

2. PRINCIPLES OF DSC The basic principle underlying this technique is that when a substance undergoes a physical transformation such as phase transitions, it either absorbs or releases heat. A simplified scheme of DSC equipment is shown in Figure 1.

-3-

Figure 1. Simplified scheme of DSC devices (USM, 2005).

In DSC, the first step we do it to heat our samples by heaters where the temperature is controlled by the computer. As shown in Figure 1, there are two pans. One is the sample pan and the other one is the reference pan. We them put the sample of interest in the sample pan and leave the reference pan empty (or place a reference sample.) Underneath each pan, there is a heater which is connected to the computer. The heating rate can be set and adjusted by the computer at a specific pattern, for example 10 per minute. Throughout the experiment, the computer would

make sure that the heating the rate remains the same and more importantly, it makes sure that the two separate pans, with their two separate heaters, heat at the same rate as each other. In the sample pan, due to the existence of extra material, it will take more heat to keep the temperature of the sample pan increasing at the same rate as the reference pan. The extra heat taken is what measured by the differential scanning calorimeter. We will then obtain a plot as the temperature increases. The x-axis would be temperature and the y-axis is the difference in heat output of the two heaters at a given temperature. This is the plot of the heat absorbed by the sample against temperature (see Figure 2) (USM, 2005). Generally, the temperature program for a DSC analysis is designed such that the sample holder temperature increases linearly as a function of time. The reference sample should have a well-defined heat capacity over the range of temperatures to be scanned (Farkas & Mohcsi-Farkas, 1996).

-4-

Figure 2. Plot of DSC (USM, 2005)

The heat flow is presented in units of heat (q) supplied per unit time (t), and the heating rate is temperature increase (T) per unit time (t):

Divide the heat flow q/t by the heating rate T/t, we can get the heat supplied divided by the temperature increase, which represents heat capacity:

During the heating process, depending on the properties of every substance, substances may undergo thermal transitions such as glass transition (Figure 3a), crystallization (Figure 3b), and melting (Figure 3c); the latter two occur only when a substance is able to form crystals. In other words, amorphous polymers do not show all these features, yet polymers contain both crystalline and amorphous domains will display all the changes (Figure 3d) (USM, 2005).
-5-

a)

b)

c)

d)
Figure 3. DSC plots. a) Glass Transition; b) Crystallization; c) Melting; d) Substances that undergo all three stages (USM, 2005).

From Figure 3, we can see a big difference between the glass transition and the other two thermal transitions, crystallization and melting. For the glass transition, there is neither dip nor peak appears, this is because there is no latent heat given off or absorbed by the sample during the glass transition. Both melting and crystallization involve giving off or absorbing heat, yet we do see a change in the heat capacity at the glass transition temperature. We call the glass transition a second order transition and transitions like melting and crystallization first order transitions(USM, 2005). As a solid sample melts to a liquid form, it will require more heat flow to the sample to increase the temperature at the same rate as the reference. This is due to the absorption of heat by the
-6-

sample as it undergoes the endothermic phase transition from solid to liquid. Likewise, as the sample undergoes exothermic processes, less heat is required to raise the sample temperature. Figure 3b refers to an exothermic transition and Figure 3c refers to an endothermic transition where the release and absorption of heat is involved in respectively. By observing the difference in heat flow between the sample and reference, differential scanning calorimeter can measure the amount of heat absorbed or released during such transitions. It is also widely used in industrial settings as a quality control instrument due to its applicability in evaluating sample purity and for studying polymer curing (Farkas & Mohcsi-Farkas, 1996).

3. APPLICATION OF DSC IN FOOD ANALYSIS 3.1. DSC of Lipids The applications of DSC in lipids in food, e.g. fats, oils, and high fat content spreads, include the determination of onset temperature of melt, crystallization, polymorphic behavior, and oxidation stability (Heussen, 2011 ). As you can see the melting curves of various oils in Figure 4, the polymorphic behavior and overlapping melting range of most naturally existing oil makes it impossible to identify a single component from fat mixtures simply through DSC, yet it is still potential for the detection of adulteration (Farkas & Mohcsi-Farkas, 1996).

Figure 4. DSC melting curves of various oils: a) sunflower; b) linseed; c) olive; d) grape (Farkas & Mohcsi-Farkas, 1996).
-7-

In the study of crystallization and polymorphic behavior of oils, X-ray diffraction (XRD) pattern is sometimes applied together with DSC to confirm the formation of desirable or undesirable crystalline forms that may occurs during different stages of heat-cool-heat process or during storage (Szydowska-Czerniak, Karlovits, Lach, & Szyk, 2005). Understanding the melting and crystallization behaviors of lipids helps making decisions during production process that when and in what conditions a lipid should be added and when manufactures are seeking replacement ingredients for their products. This is very important, since they may affect the physical and textural properties of final products. DSC is also available for the measurements of lipid stability by the determination of oxidation induction time (OIT) in isothermal conditions. The high sensitivity for oxidation of unsaturated fatty acids is reflected in the onset of oxidation exotherms at a relatively low temperature as compared to hydrogenated /coconut oil (Farkas & Mohcsi-Farkas, 1996; Heussen, 2011 ).

3.2. DSC OF STARCH For the past thirty years, DSC has been used to study the thermal transitions of starches including retrogradation, gelatinization, and glass transition (Yu & Christie, 2001). A thermal transition in starches affect the cooking, textural and digestive properties of food, e.g., the gelatinization property allows starches to act as thinkers or stabilizers in several food products and retrogradation property which related to aging of gelatinized starches elevates the level of resistance starch (RS) may be preferred due to it has the property of dietary fiber. It is reported the amount of RS in final products may depend on the degree of gelatinization during processing (Sichina, 2000).

Retrogradation is an endothermic reaction that happens only in cooked or modified starches stored at lower temperatures. When native starch is heated in water, the crystalline structure of amylose and amylopectin molecules hydrate to form a viscous solution. If the viscous solution is cooled or left at lower temperature for a long enough period, the linear molecules: amylose and the linear parts of amylopectin molecules retrograde and rearrange themselves again to a more

-8-

crystalline structure. Retrogradation expels water from a polymer network which is also known as syneresis. It is directly related to the staling or aging of bread (Eliasson & Larsson, 1993).

Most starches and rice products contain fats and could form an amylose-lipid complex during gelatinization. The amylose-lipid complex is a thermo reversible complex which shows an exothermic peak on cooling. Sometimes the modification of the amylose with a lipid is performed to control the texture of the final starch (Heussen, 2011 ).

Different starches exhibit their own characteristics of transitions during heating, the temperature and intensity of transitions are significant parameters indicating particular reaction occurs within each starch system (Sichina, 2000). Figure 5 shows the DSC curves of gelatinization of different starches (Farkas & Mohcsi-Farkas, 1996) and Figure 6 and 7 shows the effects of heating rate and water content on gelatinization respectively (SII, retrieved 2012, Mar 28). Through DSC curves, we may determine the characteristics and types of starches, evaluate the rates of heating process, and observe the effects of various water contents in starches.

Figure 5. Heat gelatinization curves of aqueous suspensions of various starches and starchy flours (Farkas & Mohcsi-Farkas, 1996).

-9-

Figure 6. Heating rate and gelatinization of corn starch (water content: 50%). a : 3/min; b : 2/min; c : 1/min (SII, retrieved 2012, Mar 28)

Figure 7. Water content and gelatinization of corn starch (heating rate : 3/min). a : 43%; b : 50%; c : 60% (SII, retrieved 2012, Mar 28)

3.3. DSC OF PROTEIN

- 10 -

- 11 -

REFERENCES

1. R. L. Biltonen and E. Freire, CRC Critical Rev. in Biochem., Nov. (1978) 85. 2. S. D. Arntfield, M. A. H. Ismond, E. D. Murray, Thermal Analysis of Foods, (eds.) V. R. 3. Harwalkar and C. Y. Ma, Elsevier Appl. Science 4. Bertazzon and T. Y. Tsong, 1990. Biochemistry, 29, 6447. 5. S. D. Arntfield, E. D. Murray, (1981). Can. Inst. Food Sei. Technol. J., 14 289. 6. H. K. D. H. Bhadeshia. Differential Scanning Calorimetry, University of Cambridge, Materials Science & Metallurgy. 7. H. Levine and L. Slade, Thermal Analysis of Foods, V. R. Harwalkar and C. E Ma (eds.), Elsevier Appl. Science, London & New York (1990) 221.

REFERENCES

Eliasson, A. C., & Larsson, K. (1993). Cereals in breadmaking: a molecular colloidal approach (Vol. 55): CRC. Farkas, J., & Mohcsi-Farkas, C. (1996). Application of differential scanning calorimetry in food research and food quality assurance. Journal of Thermal Analysis and Calorimetry, 47(6), 1787-1803. Hhne, G., Hemminger, W., & Flammersheim, H. J. (2003). Differential scanning calorimetry: Springer Verlag. Heussen, P. (2011 ). Practical Food Applications of Differential Scanning Calorimetry (DSC). PerkinElmer, Inc. Sichina, W. J. (2000). Use of DSC for the Characterization of Starches. Thermal Analysis Application Note, PerkinElmer, Inc. SII. (retrieved from 2012, Mar 28). Application Brief: Measurements of Gelatinization of Starch by DSC. Szydowska-Czerniak, A., Karlovits, G., Lach, M., & Szyk, E. (2005). X-ray diffraction and differential scanning calorimetry studies of transitions in fat mixtures. Food Chemistry, 92(1), 133-141. USM. (2005). Differential Scanning Calorimetry. Polymer Science Learning Center, Department of Polymer Science. Yu, L., & Christie, G. (2001). Measurement of starch thermal transitions using differential scanning calorimetry. Carbohydrate Polymers, 46(2), 179-184. doi: 10.1016/s01448617(00)00301-5

- 12 -