Carnosine in Amphibian Voluntary Muscle

Abstract

1. Three modifications are suggested in the KOESSLER and HANKE colorimetric technique (based on the Pauli reaction) for the estimation of pyrimidine bases: (a) the substitution of bicarbonate for some of the carbonate in one of the reagents (leading to greater stability in the pigment formed); (b) the use of carnosine nitrate as a standard of reference for estimation instead of a mixture of dyestuffs; and (c) the use of an appropriate light filter in conjunction with the colorimeter. This leads to an eightfold increase in sensitivity. 2. Extracts from symmetrical pairs of muscles from the legs of the frog give Pauli reactions of equal intensity. This equality is not destroyed by previous fatiguing or killing by heat of one of the pair. 3. The cutaneo‐crural, sartorius, gracilis, semitendinosus, semimembranosus, and gastrocnemius muscles contain increasing concentrations of carnosine (measured by the Pauli reaction) in the order named. 4. Different frogs from one batch differ considerably in the apparent carnosine content of any one muscle; variations in the gastrocnemius, for example, were observed from 124 to 293 mg. per 100 grm. 5. The Pauli reaction given by histidine, histamine, carnosine, and extracts of living and dead muscles have been studied. In the conditions chosen, all save histamine were alike in the tint given and in the rate of colour production and fading. 6. No substance capable of giving the Pauli reaction was found to diffuse from living muscles into a surrounding saline whether the muscles were resting or fatigued, and regardless of the presence or absence of oxygen. 7. Histidine can diffuse from saline into an excised resting muscle and out again. The equilibrium reached appears to be a simple osmotic equilibrium. 8. All substances capable of giving the Pauli reaction diffuse freely out of a muscle in rigor (induced by heat or by iodoacetic acid). 9. Carnosine has been isolated in the form of its copper salt in comparable amounts both from muscles killed while in the resting state and from those previously sent into heat rigor. 10. Carnosine diffuses freely and completely into a dead muscle, but if the muscle is alive only 30 per cent. of its bulk appears to be permeable to carnosine diffusing in. 11. It is concluded that the carnosine in living muscles of the frog is contained within some series of structures occupying only 70 per cent. of the bulk of the muscle. It is retained in virtue of some property of the tissue which is lost at death. 12. No evidence was found for the existence either in dead or living muscle of substances other than carnosine capable of giving the Pauli reaction. This work has been financed by the Medical Research Council.

Diffusion of inorganic phosphate into and out of the skeletal muscles and bones of the frog

“…Recently it has been found that the same statement is true for the amino acid histidine [Eggleton and Eggleton, 1933], but the behaviour of the dipeptide carnosine in similar experiments indicated that only part of the water of the muscle was available to carnosine diffusing in from surrounding saline.This fraction was of the order of 30 p.c. of the total water of the muscle.…”

The Water and Base Balance of the Body 1

“…These figures are corroborated by recent analyses of dog muscle made by Hastings (24). Eggleton (25,26) has shown that certain solutes diffuse through all the water of the frog muscle, while other substances diffuse through only 20 to 30 per cent of the water of vital frog muscle, presumably the extracellular portion. After rigor mortis has set in this selective permeability disappears.…”

Distribution of chloride in frog's skeletal muscle immersed in saline solution

“…5. This technique has been described by Eggleton et al [1928]. The behaviour of agar gels followed very closely the "square-root law"; the slight but invariable deviations from the law exhibited by living and dead muscles must be attributed to their structure, in so far as it is not accounted for by the [Eggleton et al 1928].…”

“…4. This method has already been used in determining the equilibrium value of creatine [P. Eggleton, 1930], inorganic phosphate [M. G. Eggleton, 1933] and lactate [Ghaffar, 1935] Fig. 5).…”

“…The question as to whether this quarter of the muscle is identical with the anatomically defined interspaces is open to serious doubt, for no direct histological estimate has yielded so high a figure for its size. The evidence that one-quarter of the muscle can be regarded as interspaces in a physiological sense is the well-established fact that when muscles are brought into equilibrium with Ringer's solutions containing the substance under investigation, the apparent concentration of the latter in the muscle is only one-quarter the concentration outside [Conway & Kane, 1934;Eggleton & Eggleton, 1933]. It is obvious that although this result is most simply explained on the "interspace" theory, two others, at least, are equally possible.…”