Original ContributionsQuantitative magnetic resonance imaging of fresh and frozen-thawed trout
Introduction
There is ample precedent in the literature for the use of the Nuclear Magnetic Resonance (NMR) parameters of water to study bulk specimens of meat, and it is known that those parameters are sensitive to changes in meat structure such as those due to the development of rigor mortis.1, 2, 3 Recently MRI has been used to study changes in those parameters induced by freeze–thawing of red meat, and it has been suggested that such measurements may distinguish fresh from frozen–thawed meat.4
Authentication of fresh fish is of great importance to the consumer for two reasons; as there is a price differential between fresh and frozen produce, retailing of fish as ‘fresh’ when it has been previously frozen is fraudulent; also many fish and shellfish can be hazardous with regards to their microbiological safety if they have been stored carelessly. The main purpose of this project was to evaluate the effects of freeze–thawing on the MRI parameters of water in trout flesh. Specifically, we have measured the values of water concentration (M0%), the spin-lattice relaxation time (T1), the spin-spin relaxation time (T2), the magnetisation transfer parameters Msat/M0, T1sat and MT rate for the protons of water. In order to support those measurements we have evaluated different MRI protocols for visualising the anatomical structures of intact trout, and also the spatial distribution of lipid- and collagen-rich tissue. Rainbow trout belongs to the super-order Teleostei, the bony fish with vertebrae; various imaging contrasts have been used previously to show selective enhancement of different tissue types in cartilaginous fish, Elasmobranchii.5
Freezing is an important method for reducing chemical and biological spoilage in fish, but can result in irreversible changes in its quality compared to unfrozen ‘fresh’ fish. The loss of quality as judged by changes in texture, toughening and loss of water-holding capacity,6 is thought to be mainly due to two processes; protein denaturation/aggregation and lipid oxidation. When fish is frozen, the solutes present in the muscles become concentrated as water crystallises during freezing; this increases the ionic strength of the liquid which leads to breaking of electrostatic bonds in proteins and thereby to their denaturation and aggregation. It is believed that myosin and actin, the main contractile proteins, are largely responsible for the functional properties of the muscle and that on frozen-storage, myosin in particular, undergoes denaturation/aggregation reactions.6, 7, 8, 9, 10 Although it is thought that these protein changes upon freezing are the main causes of quality loss in frozen fish,6 oxidation of lipids can lead to rancid flavours. In addition, lipid oxidation is thought to enhance the deterioration of proteins.11, 12 Lipids have been shown to be prevalent in trout by the use of MRI weighted images.13
Changes associated with freeze-storing cod, as well as thermal and high pressure treatment, have been followed14 at 20 MHz (0.47T). It was found that the transverse relaxation time of the water protons showed multi-component behaviour, with frozen-stored cod having a longer relaxation component that increased with the temperature and duration of frozen-storage. This component was attributed to the water exudate associated with protein denaturation, as well as that from physical damage and enzymatic reaction. A similar MR-relaxation study was undertaken on frozen-storage of post rigor minced cod,15 which also showed bi-exponential relaxation behaviour for the water protons. The MR parameters showed variation on frozen-storage between different quality minces, which became more marked in the cooked state. These changes were ascribed to the aggregation of myofibrillar proteins during frozen-storage as opposed to intra/extra-cellular compartmentalisation since the bi-nodal distribution persists into the cooked state.
Since the precise history of the fish flesh is of paramount importance to any study of temporal changes,16 attention was focussed in the present study on freshwater rainbow trout which were purchased live from a local trout farm. Initially, a fully automated MRI protocol was used to study a small number of whole trout. However, when those results were analysed it was realised that a much larger number of samples needed to be studied to account for biological variations and so a multiple-sample, bulk-MRI approach was used. Given that inter-animal variation was likely to be large, it was also decided to investigate the feasibility of a comparative method, which involved repeated freeze–thawing of the sample. The hypothesis is that the change in the MR parameters on re-freezing a sample which had been previously frozen would be smaller than those on freezing a fresh fish, since the major damage had already been inflicted.
Section snippets
MRI hardware
All 1H MRI images were acquired using a Bruker BMT (Bruker Medzintechnik Biospec II) imaging console (Karlsruhe, Germany) connected to a 2.35 Tesla, 31 cm horizontal bore super-conducting magnet (Oxford Instruments, Oxford, UK). A 20 cm gradient set built ‘in-house’ with each axis powered by a Techron gradient amplifier (Model 7790, Crown International Inc., Elkart, IN, USA) provided gradient strengths up to 100 mT m−1. An ‘in-house’ built, cylindrical, eight strut, bird-cage radio-frequency
Anatomical images
Segmented anatomical images were obtained using T1-weighted multi-slice imaging for the head of a fresh rainbow trout in the coronal direction (Fig. 1). Those images revealed numerous anatomical features, including the gill structures, the eyes, nasal passage and vertebrae. They gave high contrast between muscle and fat tissue because the T1 of fat is approximately half that of muscle. Comparison in Fig. 2 with the fat-resolved image obtained by a three-point Dixon technique39, 40 clearly
Conclusions
The original aims of this work were 2-fold; first, to demonstrate the type of anatomical resolution that MRI can achieve between individual organs of intact trout. Second, to explore the potential use of MRI as a basis for the authentication of fresh trout from that which had been frozen–thawed.
It is clear that MRI enabled the anatomical details of rainbow trout to be visualised by highlighting various soft tissues using a combination of protocols that gave different contrasts. This has
Acknowledgements
The authors thank the UK Ministry of Agriculture Fisheries and Food (MAFF) and the Herchel Smith Endowment for collaborative funding of this work; Miss A. Kshirsagar for providing the edge detection software; Dr. J.J. Tessier for the MRI quantitation protocols; Dr. T.A. Carpenter, Mr. C. Bunch, Mr. S. Smith and Mr. C. Harbird for supplying and maintaining the MRI facilities.
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