Preferred metabolic pathway of bovine muscle fibre revealed by synchrotron–deep ultraviolet fluorescence imaging

International audienceThe different bovine muscle fibre types I, IIA and IIX are characterised by their preferred metabolic pathway, either oxidative (I, IIA) or glycolytic (IIX), and their contraction speed, either slow-twitch (I) or fast-twitch (IIA, IIX). These physiological specificities are associated with variations in intracellular composition and their fluorescence spectra signatures. We hypothesised that these slight differences in autofluorescence responses could be used to discriminate the muscle fibre types by fluorescence imaging. Serial histological cross-sections of beef longissimus dorsi were performed: the start set was used to identify the metabolic and contractile type of muscle fibres by both immunohistoenzymology and immunohistofluorescence, and the following set was used to acquire synchrotron–deep ultraviolet (UV) autofluorescence images after excitation in the UV range (275 nm and 315 nm). This strategy made it possible to explore the label-free autofluorescence of muscle cells previously subtyped by histochemistry. Glycolytic cells (IIX) showed more intense fluorescence than oxidative cells (I and IIA) with near-90 % accuracy. This discrimination is more specifically assigned to the fluorescence of nicotinamide adenine dinucleotide. UV autofluorescence was unable to discriminate contractile type

Muscle fibre type and beef quality

Muscle fibre type and beef qualit

Differentiation of bovine from porcine gelatines using polyclonal anti-peptide antibodies in indirect and competitive indirect ELISA

Gelatine is a collagen derivative obtained from bones and hides/skin mainly from bovine and pigs. As a consequence of the outbreak of bovine spongiform encephalopathy (BSE), the use of bovine gelatine in feed, food and pharmaceutical products has been restricted by regulatory authorities. However, no method was presently available for its specific detection. The large similarity in amino-acid sequences of collagens from different species make their immunochemical differentiation difficult when using polyclonal antibodies raised against the whole molecule [A. Venien, D. Levieux, J. Immunoassay Immunochem., in press]. To obtain bovine-specific antibodies, we immunized rabbits against putative species-specific sequences of the bovine collagen alpha 1 (1) chain. Using these antibodies, an indirect ELISA was developed to allow a quick and easy differentiation between bovine and porcine gelatines. Moreover, a competitive indirect ELISA was found suitable to detect bovine gelatine in porcine gelatine purchased from laboratory chemicals suppliers down to a dilution of 2-4 parts per 1000 with CVs ranging from 5.7 to 7.7%. When testing mixtures of the largest possible range of industrial batches of bovine and porcine gelatines (skin/hides or bones origin, acid or alkaline processes, high or low Bloom) the detection limi

Pepsin localization in digested meat by ToF-SIMS analysis

Meat, is a highly structured product consisting of muscle fibres, lipids and connective tissue sheaths. These differents compounds are suspected of modulating the diffusion of digestive enzymes in meat during its gastric digestion. To follow the pepsin diffusion in digested meat and better understand the role of meat structure organization we investigated the use of high resolution time of flight secondary-ion mass spectrometry (Tof-SIMS) imaging to detect pepsin in food. Two beef muscle samples (1.5 X 1.5 X 2.5 cm) were incubated in a pepsin solution at pH 2. After 2 hours incubation at 37°C under agitation, the samples were cryofixed, histological sections were collected on glass slides and used for chemical imaging. The entire surface area of the sections (1.5 X 1.5 cm) was mapped by ToF-SIMS imaging (ToF-SIMS V; ION-TOF GmbH, Münster,Germany). Mass spectra of commercial pepsin powder and aqueous solution were acquired using the same tool.Pepsin powder analyzes revealed signals of nitrogen species that seems to be characteristic of pepsin.Lipids and connective tissue were revealed by the map which also exhibit large amounts of chlorine on the periphery of the section associated with a depletion of phosphorus species. This result reflects the penetration of the digestion solution which is rich in HCl, and that probably extract the soluble endogenous phosphorus compounds.Finally, an enrichment in nitrogen species CxHyN, assigned to pepsin, was observed in the penetration zone of the digestive solution, but also along the connective tissue layers.The use of ToF SIMS allowed the visualization of pepsin in a food. This result opens great perspectives to understanding the mechanisms of digestion of solid foods

Structural changes of local thai beef during sous-vide cooking

Structural changes of local thai beef during sous-vide cooking. 64. International Congress of Meat Science and Technolog

Muscle fiber types identification by synchrotron fluorescence microspectroscopy

The skeletal muscle consists of three to four pure types of muscle cells (also called muscle fibers) identified as type I, type IIA, type IIX and/or type IIB in different proportions depending on the muscle function. They differ in their contraction speeds and metabolic pathways. Type I fibers are slow-twitch while type II fibers are fast twitch. The energy required to maintain cell homeostasis and muscle contraction is provided by the hydrolysis of ATP. The fibers IIX and IIB regenerate ATP by anaerobic glycolysis with lactate production. Fibers I and IIA favor cellular respiration (glycolysis + Krebs cycle). The latter are rich in mitochondria, where the cell respiration take place, and in myoglobin which carries oxygen to mitochondria. The intracellular composition of fibers therefore depends on their metabolic and contractile characteristics. The objective of our work was to study the impact of these slight differences in composition on the optical properties of muscle cells. Our hypothesis was, in part, based on the autofluorescence detection of NADH which is more concentrated in the mitochondria and thus in the oxidative metabolism fibers. Therefore, we studied the impact of cell type on the fluorescent response following excitation in deep UV. Rat soleus muscle consisting of I and IIA fibers and extensor digitorum longus (EDL) muscle consisting of I, IIA, IIX and IIB fibers were used as models. On each muscle, fibers, previously identified on their cell types by immunohistofluorescence, were analyzed by synchrotron fluorescence microspectroscopy on stain-free serial muscle cross sections. Muscle fibers excited at 275 nm showed differences in fluorescence emission intensity among fiber types at 302 (assigned to tyrosine fluorescence), 325, 346 (both assigned to tryptophan fluorescence) and 410 nm (assigned to NADH fluorescence). The 410/325 ratio decreased significantly with contractile and metabolic features in EDL muscle, ranked I>IIA>IIX>IIB fibers (p< 0.01). In a subsequent experiment, we acquired autofluorescence images for fast fiber types discrimination on label free histological sections. Computer processing of the images allowed us to improve the contrast and identify the metabolic types of fibers with a fairly good reliability. These studies highlight the usefulness of autofluorescence signals to characterize histological cross section of muscle fibers with no staining chemicals

Characterization of smoked salmon by infrared spectroscopy

Smoked salmon results from the salting of the fillets, followed by smoking. Salting and smoking methods may vary depending on different recipes specific to each manufacturer and can cause variability in the composition of the end product.The aim of this work was to try to classify a posteriori two processes of production of smoked salmon by simple analyzes of near and medium infrared spectroscopy.The experiment was carried out on 6 salmons. For each salmon, 1 filet was salted and smoked according to the process A, the other filet was salted and smoked according to process B. Processes from industrial production are confidential.Samples were taken and analyzed both in near infrared spectroscopy (Foss NIRSystems model 6500) and in medium infrared microspctroscopy (IN10 Mx Thermo Scientific). The spectra were pre-processed and the multivariate analyzes implemented using the software "The Unscrambler".Principal component analyzes (PCAs) reveal spectral signatures (NIRS and MIR) which are characteristic of each process and therefore allow to discriminate at posteriori the samples from each process (A and B).This experiment demonstrates the power of infrared spectroscopy methods to characterize smoked salmon fillets and identified their technological itinerary

Characterization of thai beef macromolecular changes under various sous-vide cooking conditions: an infrared microspectroscopy study

Characterization of thai beef macromolecular changes under various sous-vide cooking conditions: an infrared microspectroscopy study. 64. International Congress of Meat Science and Technology (ICoMST

Postmortem changes in muscle fibres autofluorescence

After slaughter, the muscle cells undergo biochemical and physicochemical changes which may affect their autofluorescence characteristics. The postmortem metabolism kinetic of rat EDL and soleus muscles was assessed by glycogen depletion determination while autofluorescent response of different muscle fiber types was investigated by Deep UV synchrotron microspectroscopy at slaughter and and 24 hours postmortem. Following a 275 nm excitation, emission fluorescence spectra showed discrimination depending on postmortem time (T0 versus T24h) on both muscles at 346 and 302 nm and to a lesser extent at 408 and 325 nm. Taken individually, all fiber types are discriminated but with variable accuracy, the type IIA showing better separation of T0 comparing to T24h than other fiber types. These results highlight the relevance of using the autofluorescent response of muscle cells to rapidly assess their state of postmortem changes