Thermal Degradation and Decomposition Kinetics of Freeze Dried Cow and Camel Milk as well as Their C

K. V. Sunooj1，2， J. George2， V. A. Sajeev Kumar2， K. Radhakrishna2 and A. S. Bawa2

1. Department of Food Science and Technology， Pondicherry University， R. Venkataraman Nagar， Pondicherry 605014， India

2. Defence Food Research Laboratory， Siddarthanagar， Mysore 570011， India

Received: March 17， 2011 / Published: July 20， 2011.

Abstract: A study was conducted to evaluate thermal properties and degradation kinetic parameters of FD cow milk and camel milk powder. FT-IR was used to confirm the fat removal from the whole milk powder. Differential Scanning Calorimetry (DSC) was used to study the thermal transitions. DSC thermograms of WMP showed a two-step endotherm， the former at lower temperatures (cow milk 16-35 °C， camel milk 25-60 °C) and the later at higher temperatures (cow milk 75-125 °C， camel milk 90-160 °C). The main difference observed between cow and camel milk was peak maximum temperature for fat melting， ΔH and other decomposition temperatures. The enthalpy of fat melting was 2.314 J/g and 3.397 J/g for cow and camel milk respectively. Thermogravimetric Analysis(TGA)/derivative thermogravimetric analysis (DTG) also showed two steps degradation. The first step involves lactose degradation and second step corresponds to combined degradation of protein and fat. Hence logβ vs 1000/T was plotted separately for individual components to determine cumulative value of activation energy using Flynn-Wall-Osawa method.

Key words: Thermal degradation， differential scanning calorimetry， thermogravimetric analysis， FT-IR， Flynn-Wall-Osawa method.

1. Introduction

Milk and milk products are widely accepted by consumers throughout the world， because of their nutritive value and easy availability. Since milk is a highly perishable commodity， it is generally converted into more shelf stable powder form. Milk powder itself is available in various forms like whole milk powder， skim milk powder etc. It is manufactured using various drying processes such as spray drying， freeze drying etc. The main advantage of milk in powder form is that it can be easily transported from areas of surplus to those of deficit and can be reconstituted later at the user end. Milk powder is a common ingredient in several bakery products， soup powders， snacks， chocolates and confectionaries. More recently milk powder has been used as a major component in extruded food products for specialty applications like therapeutic foods [1]， appetite suppression [2] etc.

Maintaining the natural taste， flavour and nutritional attributes of milk powder based products like chocolate， cookies etc. even after processing is always a challenging task [3]. Deterioration in nutritional quality owing to high temperature exposure during baking and extrusion processes is very common [4， 5]. Processing techniques involving elevated temperatures lead to denaturation [6] and reduction in water binding capacity [7] of proteins. Such conditions may also lead to loss of other constituents like sugar [8] and even cause undesirable chemical reactions [9]. Thermal stability of each component in food systems is different. Hence a detailed knowledge with respect to thermal stability and decomposition temperatures is essential for rigorous process and strict quality control during thermal processing of foods.

Thermal analyses using Thermogravimmetry/ Derivative Thermogravimetry (TGA/DTG) and Differential Scanning Calorimetry (DSC) have been extensively carried out by the food industries [10]. These techniques have been widely used to measure changes in the properties of foods as a function of temperature by subjecting it to a controlled temperature programme. DSC is the major technique used for studying glass transition temperature (Tg) and the amounts of heat flow occurring during various thermal transitions in foods [11]. TGA can be used to determine kinetic parameters that govern thermal decomposition reactions as well as for estimating the activation energy required for decomposition of each individual constituent in the food [12].

The objective of this study was to evaluate the thermal properties of freeze dried cow and camel whole milk powders (FD-WMP). Various kinetic parameters of these milk powders were studied and the activation energy of decomposition (Ea) for the major constituents of milk powders was estimated. Freeze-drying was employed to avoid any temperature-induced changes in milk powders. Since the fat content in the milk can influence thermal properties of its other constituents like protein and lactose， defatted cow and camel milk powder (DMP) was also used in this study.

2. Materials and Methods

2.1 Sample Preparation

Cow milk samples were collected from the local market， while the camel milk was obtained from a farm in Rajasthan (India). The collected samples were refrigerated immediately and transported to the laboratory. Both the cow and camel milk samples were divided in to two lots. The first lot was refrigerated until freeze-dried. The second lot was centrifuged at 2500 rpm for 30 min at 4 °C to separate fat and the isolated fat was used for thermal studies. The whole and defatted milk samples were freeze-dried and used for further studies. Similarly protein was isolated from milk powders as per AOAC [13] methods (AOAC 15.120).

2.2 Freeze Drying

Freeze drying was carried out in a pilot scale freeze dryer (Hull Corporation USA) equipped with rapid freezing and drying facilities. The milk samples were pre-frozen to -40 ± 2 °C for 4 hours and drying carried out in a chamber maintained at 1-3 m bar pressure and temperature in the range of 30-60 °C， so that the final moisture content of the milk samples was reduced to 2%-3%. After drying vacuum was released and the dried material taken to low humidity room (23 ± 2%) and packed.

2.3 Chemical Composition

Milk powder samples were analyzed for moisture， protein， fat， lactose， ash and titratable acidity according to the procedures (15.111， 15.120， 15.125， 15.028， 15.122， and 15.004) respectively of AOAC [13]. pH was measured using pH meter (Model pH 510， M/s Eutech Instruments， India) at room temperature.

2.4 Fourier Transform Infrared (FTIR) Spectroscopic Analysis

Freeze-dried milk powders (whole milk powder and defatted milk powder) were analyzed using Fourier transform infrared (FTIR) spectroscopy. The IR spectra were recorded using Thermo Nicolet FTIR spectrometer (Model 5700， Madison， WI) fitted with single bounce Attenuated Total Reflectance (ATR) accessory with ZnSe crystal. Sixty-four scans were averaged to reduce the noise. All the spectra were recorded at 4 cm-1 resolution and analyzed using the software (OMNIC) provided along with the instrument.

2.5 Differential Scanning Calorimetry (DSC)

A DSC 2010 differential scanning calorimeter (TA Instruments， New Castle， DE， USA) was used to determine the Tg values. Samples were enclosed in hermetically sealed aluminum pans just before analysis and then loaded into the equipment at room temperature. An empty pan was used as reference. Nitrogen gas at a flow rate of 60 mL/min was employed in the purge line to control the local environment around the sample. The calibration of the cell was made following the DSC manufactures recommendations and consisted of three procedures namely baseline， cell constant and temperature calibration. The base line calibration involved heating the cell through the entire temperature range i.e.， from-30 to 250 °C. The cell constant was determined by heating the standard material (indium) through its melting temperature. The temperature calibration was performed using a multiple-point calibration with indium (mp 156.60 °C， ΔHm 28.71 J/g)， distilled water(mp 0 °C， Hm 335 J/g) and heptane (mp， -91.0 °C ΔHm 140 J/g). In our experiments samples were scanned at 20 °C /min in the temperature range of -30 °C to 250 °C. The DSC measurement was repeated thrice. The results of the experimental run were analyzed with the universal analysis software provided along with DSC instrument. DSC was also employed to determine the thermal behaviour of different constituents in the milk powder. The enthalpy of fat melting was determined by integrating the peak maximum values. 2.6 Thermo Gravimetric Analysis (TGA)

Thermo gravimetric analyzer (TGA Q50， TA Instruments， USA)， a thermal weight change analysis instrument was used in conjunction with a thermal analysis controller. The Thermogravimetric analyzer was employed to measure the amount and rate of change in weight of the material either as a function of increasing temperature or time， in a controlled atmosphere. The initial weight of each sample was approximately 10 mg. The samples were kept in platinum crucible flushed with nitrogen gas at the rate of 40-60 mL/min and heated in a furnace from 30 °C to 600 °C using different heating rates like 5， 10， 15 20 °C/min. The percentage and derivative weight loss were recorded against the temperature for all the samples.

2.7 Calculation of Kinetic Parameters Using TGA Analysis

Thermogravimetric measurements together with Flynn-Wall-Osawa method were employed to assess the kinetic parameters and to compare the thermal behavior. This method is a non isothermal method where the rate of heating (β) is controlled by a programmed linear heating. Generally the degradation of food samples followed a first order rate Eq. (1) given in the form: dα/dt = k (1-α) (1)

Where k is the Arrhenius rate constant， which can be expressed as Eq. (2): k = Ze-Ea/RT (2)

Where Z is the pre-exponential factor， Ea is the activation energy， R is the universal gas constant and T is the temperature. In Flynn-Wall-Osawa method， by the introduction of β， the equations can be rearranged as Eq. (3)， where a and b are constants: Log β = a/T + b (3)

When log β vs. 1000/T is plotted， a straight line is obtained， which can be used for calculation of activation energy， Ea. 2.8 Statistical Analysis

Statistical analysis was performed using SPSS software (SPSS Inc.， 1996) and the effect of various parameters on activation energy conversion was estimated for significance at 1% level (P < 0.01).

3. Results and Discussion

3.1 Chemical Composition

The chemical composition of cow and camel milk powders has been presented in Table 1. Percentage of all the major constituents like fat， protein and lactose in camel milk powder was slightly lower than that in cow samples， while ash content was higher. The average pH value and moisture content of camel milk powder were comparable to those for cow’s milk powder， while acidity was lower in the former.

3.2 FT-IR Analysis

FT-IR was used to characterize whole milk powder(WMP) as well as the major constituents present in cow and camel milk. Characteristic IR peaks for fat， protein and lactose were evident from the spectra of WMP of cow and camel samples as shown in Fig. 1A and 1B. The IR peaks at 1745 cm-1 and 1150 cm-1 corresponded to the stretching vibration of C = O and C-O bonds of milk fat respectively [14]. The intense bands observed at 1640 cm-1 and 1540 cm-1 corresponded to the stretching vibration of amide-I and amide-II groups respectively in milk proteins. The CH2 stretching bands at 2921 cm-1 and 2853 cm-1 were due to aliphatic groups present in major milk constituents. A major band and its associated peaks at the area from 800-1200 cm-1 corresponded to the characteristic peaks of various vibrations in lactose[15]. Other IR bands like those observed at 1443 cm-1 and 1390 cm-1 were associated with CH2 bending [16]， while the peaks at 1245 cm-1 corresponded to the CH2 was bending and CN stretching with contribution from NH bending [17]. All these characteristic peaks for cow and camel milk WMPs were in agreement with those reported in literature.

The spectra of defatted cow and camel milk powder shows the absence of two main peaks (1745 cm-1 and 1150 cm-1)， characteristic of fat， indicated the effective removal of fat from WMP. Another difference observed in defatted milk powder spectra was the reduction in intensity of CH2 stretching bands at 2921 cm-1 and 2853 cm-1， which was also attributed to the fat removal. The absence of other bands corresponded to the fat and lactose in the spectra indicating the purity of the samples isolated.

3.3 Differential Scanning Calorimetry (DSC) Analysis

DSC is widely used to characterize various first and curves of freeze dried cow milk and its constituents like fat， protein and sugars. The DSC curve of FD cow milk powder showed a two step endotherm， one at lower temperature (25-50 °C) and another at higher temperatures (110-150 °C). The endotherm occurring at lower temperature was mainly due to the melting of fat in milk， while the broad endotherm occurring at 110 to 150 °C was due to two endothermic events occurring in that temperature range. At these elevated temperatures， protein denaturations [19， 20] as well as lactose dehydration [21-23] have been found to occur and the same was confirmed by the DSC thermograms of the individual constituents. In freeze-dried milk powders lactose exist as amorphous form. DSC curve of fat， isolated from cow milk showed that the melting occurred in the temperature range of 16-35 °C and the enthalpy for the same was estimated to be 2.314 J/g. DSC thermogram of milk protein isolated from FD cow milk powder has a broad endotherm in the range of 90-150 °C. This corresponded to the denaturation of protein and removal of moisture. DSC thermograms of lactose showed two endotherms， one occurring in the range of 140-160 °C， while the other occurred at around 210-230 °C. The first endotherm having a peak maximum temperature of 156 °C corresponded to the dehydration of lactose while the second one having a peak maximum temperature of 226 °C corresponded to the decomposition of lactose. These DSC thermograms of the individual constituents were used to assign the thermal events occurring in the whole milk powder， which implied that the first transition was due to fat melting and the second one was as a result of combined effect of denaturation of protein and dehydration of lactose.

Similarly camel milk powder as well as its isolated major constituents was also investigated using DSC. The main difference observed between cow and camel milk powder was at the peak maximum temperature observed for fat melting as well as other decomposition temperatures. In case of camel milk the fat melting was found to occur in the range of 25-50 °C and the enthalpy of fat melting was estimated as 3.397 J/g， while the broad endotherm correspond to the protein denaturation and lactose dehydration occurred at 145 °C. This may be attributed to the difference in nature of fat and protein composition of cow and camel milk.

3.4 Thermogravimetric Analysis (TGA) and Degradation Kinetics

TGA was used to study the degradation profile of FD WMP from cow and camel milk. Fig. 3 depicted the thermogravimetric curves for cow and camel milk powders respectively. TGA of cow milk powder showed two major thermal degradation events occurring at 150-250 °C and 270-420 °C. Derivative thermogravimetric (DTG) curves clearly showed the peak maximum temperatures to be 214 °C and 377 °C respectively. Out of these， first decomposition event corresponded to the degradation of lactose， while the second one corresponded to the combined degradation of protein and fat. In the case of camel milk powders， there events were found to occur at 221 °C and 382 °C respectively.

The main aim of this study was to determine the kinetic parameters associated with the thermal degradation of milk powders， using Flynn-wall-Osawa method. In this method， cow and camel milk powders were subjected to a non-isothermal TGA at four different， constant heating rates of 5 °C/min， 10 °C/min， 15 °C/min and 20 °C/min. As mentioned earlier， the thermal decomposition of milk powders involved a two step process consisting of the first step as degradation of lactose while the second corresponded to the combined degradation of proteins [24] and fats. Hence log β Vs 1/T was plotted separately (i.e. from 10-25% and from 30%-77%) to determine the cumulative values of activation energy (Ea). Fig. 4 showed parallel and narrowly spaced linear plots of cow milk powder. From the slope of these plots， activation energy (Ea) levels required for thermal decomposition were calculated and plotted against percentage conversion as shown in Fig. 5. Activation energy required for thermal decomposition depended on the energy barrier preventing the decomposition of individual constituents and was influenced by the chemical composition of milk. During the initial stages， the activation energy needed for lactose degradation was almost similar for both the milk samples. However in the second stage， camel milk powder required more activation energy for achieving the same percentage conversion.

Since the thermal degradation of fat and protein occurred in almost the same temperature range(270-420 °C)， activation energy needed for thermal degradation could not be determined separately. Hence similar TGA studies conducted Fig. 6A using defatted cow and camel milk revealed that the peak maximum temperature shifted towards lower temperatures. For example， in the case of cow milk powder， fat removal resulted in the reduction of the peak maximum degradation temperature from 377 °C to 327 °C. A similar result was observed in case of camel milk powders， where peak maximum degradation temperature was reduced from 382 °C to 334 °C. This may be due to the lower thermal stability of milk proteins as compared to that of fat. Hence the second degradation step observed in the case of defatted milk powder corresponded to the protein fractions.

In order to verify the above， the thermal decomposition behaviour of isolated milk protein and milk fat was also investigated as shown in Fig. 6B. The peak maximum temperature for fat decomposition the kinetic parameters associated with degradation of individual constituents like lactose， protein and fat in cow and camel milk powders and statistically analyzed(Table 2). Activation energy at different percentage conversion levels for lactose， fat and protein were statistically significant (P < 0.01). Thermal events as well as the decomposition kinetics of individual constituents like lactose， fat and protein of cow and camel milk could be investigated.

4. Conclusions

Thermal properties and degradation kinetics of cow and camel milk were studied using differential scanning calorimetry and thermogravimetric analysis. DSC studies revealed various thermal events occurring to the whole milk powder as well as its constituents. TGA results depicted that the thermal degradation in whole milk powder occurred in two different steps， out of which the former one corresponded to the degradation of lactose while latter was due to the combined degradation of fat and protein. In order to separate these two degradation events， similar TGA studies were conducted using defatted milk powder， which helped to evaluate the thermal degradation of milk protein alone. The activation energy for the degradation of all the individual constituents of cow and camel milk powders were calculated. These studies revealed that the constituents of camel milk were more thermally stable and need more activation energy for thermal decomposition.

Acknowledgments

The authors thank Mr. K. R. Vijay， Scientist， Defence Food Research Laboratory， Mysore， and Dr. A. M.V.Sc. Shijin for their valuable guidance and help rendered during the study.

References

[1] C. Hoppe， G.S. Andersen， S. Jacobsen， et al.， The use of whey or skim milk powder in fortified blended foods for vulnerable groups， Journal of Nutrition 138 (2008) 145s-161s.

[2] R. Child， The ways of weight loss， Functional Foods and Nutraceuticals 5 (2005) 62-71.

[3] K. Franke， K. Heinzelmann， Structure improvement of milk powder for chocolate processing， International Dairy Journal 18 (2008) 928-931.

[4] S. Singh， S. Gamlath， L. Waheling， Nutritional aspects of food extrusion: a review， International Journal of Food Science and Technology 42 (2006) 916-929.

[5] A. Choudhury， M. Mukhurjee， B. Adhikari， Thermal stability and degradation of post-use reclaim milk pouches during multiple extrusion cycles， Thermochimica Acta 430 (2005) 87-94.

[6] C.H. Kim， J.A. Maya， Properties of extruded whey protein concentrate and cereal flour blends， LWT Food Technology 20 (1987) 311-318.

[7] J.M. Aguilera， F.V. Kosihowshi， Extrusion and roll cooking of corn-soy-whey mixtures， Journal of Food Science 43 (1978) 225-227.

[8] C.W.P. Carvalho， J.R. Mitchelle， Effect of sugar on the extrusion of maize grits and wheat flour， International Journal of Food Science and Technology 35 (2000) 569-576.

[9] J.C. Cheftel， Nutritional effects of extrusion cooking， Food Chemistry 20 (1986) 263-283.

[10] J.C.O. Santos， I.M.G. Dos Santos， A.G. De Souza， S. Prasad， A.V. Dos Santos， Thermal stability and kinetic study on thermal decomposition of commercial edible oils by thermogravimetry， Journal of Food Science 67 (2002) 1393-1398.

[11] M. Le Meste， D. Champion， G. Roudaut， G. Blond， D. Simatos， Glass transition and food technology: a critical appraisal， Journal of Food Science 67 (2002) 2444-2458.

[12] P.M. Madhusudanan， K. Krishnan， K.N. Ninan， New equations for kinetic analysis of non isothermal reactions， Thermochimica Acta 221 (1993) 13-21.

[13] AOAC (Association of Official Analytical Chemistry)， Official Methods of Analysis， Association of Official Analytical Chemists， Washington， D.C.， 2000.

[14] Q. Zhou， S.Q. Sun， L. Yu， C.H. Xu， I. Noda， X.R. Zhang， Sequential changes of main components in different kind of milk powders using two dimensional infra red correlation analysis， Journal of Molecular Structure 799(2006) 77-84.

[15] C.S. Pappas， P.A. Tarantilis， E. Moschopoulou， G. Moatsou， I. Kandarakis， M.G. Polissiou， Identification of goat and sheep milk based on diffuse reflectance infrared fourier transform spectroscopy (DRIFTS) using cluster analysis， Food Chemistry 106 (2008) 1271-1277.

[16] H. Zaleska， P. Tomasik， C.Y. Li， Formation of carboxymethyl cellulose casein complexes by electrosynthesis， Food Hydrocolloids 16 (2002) 215-224.

[17] A. Hewavitharana， B. VanBrakel， Fourier transforms infrared spectrometric method for the rapid determination of casein in raw milk， Analyst 122 (1997) 701-704.

[18] M.S. Rahman， State diagram of foods: its potential use in food processing and product stability， Trends in Food Science and Technology 17 (2006) 129-141.

[19] M. Paulsson， P. Dejmek， Thermal denaturation of whey proteins in mixtures with caseins studies by differential scanning calorimetry， Journal of Dairy Science 73 (1990) 590-600.

[20] I. Nicorescu， C. Loisel， A. Riaublanc， C. Vial， G. Djelveh， G. Cuvelier， J. Legrand， Effect of dynamic heat treatment on the physical properties of whey protein foams， Food Hydrocolloids 23 (2009) 1209-1219.

[21] M.K. Haque， Y.H. Roos， Differences in the physical state and thermal behavior of spray-dried and freeze-dried lactose and lactose protein mixtures， Innovative Food Science and Emerging Technologies 7 (2006) 62-73.

[22] K. Jouppila， Y.H. Ross， Glass transition and crystallization in milk powders， Journal of Dairy Science 77 (1994) 2907-2915.

[23] K. Jouppila， Y.H. Ross， Water sorption and time dependent phenomena of milk powders， Journal of Dairy Science 77 (1994) 1798-1808.

[24] H.G. Kessler， H.J. Beyer， Thermal denaturation of whey protein and its effect in dairy technology， International Journal of Biological Macromolecules 13 (1991) 165-173.