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Evaluation of microstructural changes and their relations to physical changes of shrimp during boiling using fractal analysis.


Frozen cooked shrimp is a high-value product of Thailand. Boiling is an important process of frozen cooked shrimp production. Boiling affects several qualities of boiled shrimp, especially, shrinkage and texture due to protein changes and cooking loss, which are in fact a result of microstructural changes. Relationships between microstructural changes and physical changes of shrimp during boiling are required to improve the quality of boiled shrimp. Some studies have reported relationships between microstructure and physical properties of muscle foods (Mizuta et al., 1999). However, it is not easy to describe changes of microstructure quantitatively without the use of other appropriate evaluation techniques. Recently, image analysis has been used to quantify microstructural changes of foods (e.g., Quevedo et al., 2002). Relationships between various physical changes and microstructural changes of some foods have also been established successfully using coupled image and fractal analysis (Kerdpiboon & Devahastin, 2007). The aims of the present work were to evaluate quantitative changes of microstructure of shrimp during boiling using fractal analysis. In addition, the relationships between some physical changes, namely, shrinkage and hardness, of shrimp and fractal dimension were established.


Fresh wild white shrimp (Penaeus indicus) of the size of 150-160 shrimp/kg was graded, washed and weighed; 150 [+ or -] 0.5 g of raw deheaded shrimp was used in each experiment. Shrimp was boiled in tap water in a 14-cm diameter stainless steel vessel at boiling temperature (100[degrees]C). The boiling condition was boiling time of 1, 3, 5 and 7 minutes at a mass ratio of shrimp to tap water of 1:2. All boiling experiments were performs in duplicate.

2.1 Microstructural imaging and image processing

Firstly, the boiling shrimp sample was fixed to preserve its tissues in Bouin's solution. After that, water from the tissues was removed. This was done using a series of alcohols at different concentrations. Alcohol within the sample was subsequently removed by flushing in xylene. Finally, the sample was infiltrated and embedded with paraffin. This tissue processing and embedding process was that of Humason (1979).

Each embedding sample was sectioned by a microtome (Jung, RM2025, Heidelberg, Germany) into a 8 [micro]m thick slide. Azan stain method was used to dye the sample (Humason, 1979). The microstructural images were obtained using a light microscope (Olympus, BX51, Tokyo, Japan) at a 20x magnification level. The image of sample was captured by a digital camera (Olympus, C-5060, Tokyo, Japan) at image sizes of 520x520 pixel. The image was then transformed from a red, green and blue (RGB) format to a black and white format. The edge of the muscle was detected to determine the perimeter of each muscle fiber, which was in turn used to calculate the fractal dimension.

2.2 Fractal dimension (FD) calculation

The fractal dimension of black and white image was calculated using the box counting method (Quevedo et al., 2002) using MATLAB[TM] software (version 7.01). Ten light microscopic images were used for each sample. Cubic boxes with different sizes (r) were mounted into the images. The number of boxes ([N.sub.r]), which the edges of the muscles appear on, was then counted. Fractal dimension was calculated by:

FD = log([N.sub.r])/log(l/r) (1)

To obtain the changing values of FD of the sample undergoing boiling, the normalized change of FD was reported as [DELTA]FD/[FD.sub.0] (Kerdpiboon & Devahastin, 2007). [DELTA]FD is the difference between the fractal dimension at any instant during boiling and the fractal dimension of the fresh sample.

2.3 Shrinkage measurement

The volume of deshelled shrimp (V) was determined using a liquid pycnometer with n-heptane as the working liquid (Devahastin et al., 2004). Shrinkage is the normalized changes of volume of deshelled shrimp sample ([DELTA]V/[V.sub.0]) obtained from different boiling time.

2.4 Texture analysis

A texture analyzer (Stable Micro Systems, TA.XT.Plus, UK) with 50 kg load cell with a compression probe was used to evaluate the texture of deshelled shrimp following the method of Niamnuy et al. (2007). The maximum compression stress values were reported as the hardness (H) of boiled shrimp. To obtain the change of hardness of boiled shrimp, the normalized change of hardness ([DELTA]H/[H.sub.0]) was reported. [DELTA]H is the difference between the hradness of sample at any instant during boiling and the hardness of the fresh sample ([H.sub.0]).


3.1 Microstructural changes

Light microscopic images of raw and boiled shrimp, focusing mianly on the transverse inner part of the shrimp meat, indicating muscle fibre, endomysium and perimysium, are shown in Fig. 1. The muscle fiber of raw shrimp was plump although there were gaps between muscle fibers (as shown in Fig. 1a). The gaps might occur due to small degradation in the muscle structure post mortem. It can be seen that thermal denaturation and aggregation strongly affected microstructure of shrimp during boiling. As boiling time increased, dense protein structure occurred due to protein denaturation and aggregation; the gap regions were also more frequently observed around the muscle fibres. Collagen, around the muscle fibers (endomysium) and around the muscle bundles (perimysium) reduced due to thermal shrinkage.

Fractal dimension (FD) was used to represent the quantitative changes of microstructure of the samples. FD of raw shrimp (Fig. 1a) was about 1.85 while FD of boiled shrimp was in the range of 1.87-1.91. It was found that [DELTA]FD/[FD.sub.0] of the samples increased with the boiling time as shown in Fig. 2. The rate of increase of fractal dimension was higher during the early stage of boiling because most proteins denatured since the early stage of boiling.

3.2 Relationship between FD and physical changes

During boiling shrinkage of shrimp occurred due to shrinkage of muscle protein and collagen, which denatured and aggregated. Additionally, shrinkage was also due to the loss of moisture and water soluble protein (sarcoplasmic protein). It was found that shrinkage of shrimp increased with boiling time. When protein denatured and coagulated muscle proteins lost their water holding capacity causing the hardness of shrimp to increase. It was found that hardness of shrimp increased with boiling time.

Fig. 3. illustrates the relationship between physical changes, namely, shrinkage ([DELTA]V/[V.sub.0]) and the normalized change of hardness ([DELTA]H/[H.sub.0]) of shrimp, and the normalized change of fractal dimension ([DELTA]FD/[FD.sub.0]). It can be seen that well established relationships between both observed physical changes and [DELTA]FD/[FD.sub.0] existed. It can be seen that shrinkage and hardness of boiled shrimp increased with an increase in the fractal dimension. Linear relationships between both physical changes and [DELTA]FD/[FD.sub.0] are shown in equations (2) and (3). The results show that the fractal dimension was highly correlated with the physical changes of muscle food. However, form of correlations may depend on kind of muscle foods and type of processing.




[DELTA]V/[V.sub.0] = 4.956 [DELTA]FD/[FD.sub.0] + 0.006 [R.sup.2] = 0.99 (2)

[DELTA]H/[H.sub.0] = 27.218 [DELTA]FD/[FD.sub.0] - 0.047 [R.sup.2] = 0.98 (3)


Boiling process affected protein denatuaration and aggregation, which in turn influenced the microstructure, shrinkage and hardness of shrimp, as represented by the changes in [DELTA]FD/[FD.sub.0]. An increase in boiling time led to an increase in [DELTA]FD/[FD.sub.0], shrinkage as well as hardness. The changes of both physical properties were highly correlated with [DELTA]FD/[FD.sub.0] values. The future research plan is using the fractal analysis to quantitative describe the microstructural changes and their relations to physical changes of muscle food during other processes such as fermentation of meat.


Devahastin, S., Suvarnakuta, P., Soponronnarit, S. & Mujumdar, A.S. (2004). A Comparative Study of Low-Pressure Superheated Steam and Vacuum Drying of a Heat-Sensitive Material. Drying Technology, Vol. 22, pp. 1845-1867, ISSN 0737-3937

Humason, G. L. (1979). Animal tissue techniques, Freeman Press, ISBN 0716-702991, San Francisco

Kerdpiboon, S., & Devahastin, S. (2007). Fractal characterization of some physical properties of a food product under various drying conditions. Drying Technology, Vol. 25, pp. 135-146, ISSN 0737-3937

Mizuta, S., Yamada, Y., Miyagi, T., & Yoshinaka, R. (1999). Histological changes in collagen related to textual development of prawn meat during heat processing. Journal of Food Science, Vol. 64, pp. 991-995, ISSN 0022-1147

Niamnuy, C., Devahastin, S., & Soponronnarit, S. (2007). Quality changes of shrimp during boiling in salt solution. Journal of Food Science, Vol. 72, pp. S289-S297, ISSN 0022-1147

Quevedo, R., Carlos, L. G., Aguilera, J. M., & Cadoche, L. (2002). Description of food surfaces and microstructural changes using fractal image texture analysis. Journal of Food Engineering, Vol. 53, pp. 361-371, ISSN 0260-8774
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Article Details
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Author:Niamnuy, Chalida; Devahastin, Sakamon; Soponronnarit, Somchart
Publication:Annals of DAAAM & Proceedings
Article Type:Report
Geographic Code:4EUAU
Date:Jan 1, 2009
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