Copper

Copper is an essential element found in the blood and other tissues of Crustacea. It forms a part of cuproprotein required for oxygen transport and electron transport systems. It also forms as prosthetic group in many enzymes such as phenol oxidase and acts as activator for enzymes such as malate dehydrogenase. It is toxic in free state (Holden, 1970) and dialysable copper is absent in the blood of crustaceans (Arumugam & Ravindranath, 1980). The copper exists in blood both in the cuprous and cupric state and is linked with protein through sulphydryl groups (Klotz & Klotz, 1955). Several methods are in vogue for determination of copper. Some of them are suitable for vertebrate tissue where it is 100 times lower than the crustacean blood or other tissues. Here the suitability and consistency of 3 spectrophotometry methods were analysed for determination of crustacean blood or other tissue copper concentration

Proteins

The nitrogen of the protein precipitate is converted to acid ammonium sulphate by digestion with sulphuric acid and various catalysts. On making the reaction mixture alkaline, ammonia is liberated, which is removed by steam distillation. The ammonia liberated by steam distillation is gathered in acid solution containing an indicator. Change in pH of the acid solution due to addition of ammonia is indicated by the indicator dye. This solution is back-titrated with 0.01 N HCI, and the original acidic condition indicated by the indicator dye, is taken as the end point. The amount of HCl consumed in back titration is proportional to the amount of ammonia liberated. Percentage of nitrogen is calculated from the titre value, which is converted into gm% of protein present in sample by multiplying the percentage nitrogen with a factor 6.2S. The factor 6.25 is used for conversion because in average, protein contains 16% of nitrogen

Total free sugars, reducing sugars and glucose

Carbohydrates in the tissues of crustaceans exist as free sugars and as bound with proteins (Saravanan, 1979). The free sugars in haemolymph consist of mono, di and oligosaccharides. All monosaccharides, maltose and its oligosaccharides constitute the total reducing sugars. Trehalose constitutes the non-reducing sugar fraction of the total free sugars. The total free sugars are estimated by Anthrone method and reducing sugar by Nelson-Somogyi method. The difference in the values obtained by these two methods indicates total non-reducing sugar value which is primarily trehalose in crustacean blood. The glucose can be determined by Glucose-oxidase method. The difference between values of glucose and reducing sugars would indicate the concentration of non-glucose reducing sugars

Biochemical Variability

A perusal of previous literature on the biochemistry of crustaceans would reveal a remarkable intra-specific variability in biochemical composition of tissues. A critical assessment of factors responsible for biochemical variability of tissues of individual species is necessary, in order to understand the physiological role of the biochemical components in question. Halberg (1973) has broadly classified the factors influencing individual biochemical variability as inter-individual and intraindividual. The inter-individual factors would include sex, size, moulting, nutritional and reproductive status in addition to pathological and environmental conditions. The intraindividual factors which may bring about biochemical variability include techniques and time of day

Ammonia excretion and uptake

Ammonia is one of the major excretory products of. .aquatic crustaceans. The rate of ammonia excretion may reflect the activity of the animal (Subhashini, 1981). Ammonia excretion in crabs as well as in several isopods show diurnal variation (Kirby & Harbaugh, 1974; Subhashini, 1981). The amount of ammonia excreted by the animal can be determined by placing the animal in ammonia free artificial sea water, following the method as mentioned in 8.0. Crabs are known to take in" ammonia from the medium (Mangum & Towle, 1977 ; Subhashini, 1981)

Blood Volume

the principle underlying the methodology of blood volume determination involves the determination of degree of dilution of an injected dye by comparing the concentration of the injected dye and the concentration of the dye in the blood after a particular period of injection. The degree of dilution reflects blood volume (Lee, 1961)

Phosphatases

The phosphatases are the group of enzymes of low substrate specificity and are characterised by the ability to hydrolyse a large variety of organic phosphate esters with the formation of an alcohol and phosphate ions. This group is composed of those enzymes which attack only monoesters of orthophosphoric acid. The alcohol esterified to the orthophosphoric acid, (HO)» P=0, may be a simple aliphatic alcohol, a polyhydric alcohol such as sugar or any one of a variety of aromatic hydroxyl compounds such as tyrosine. The phosphatases are not one enzyme but a group of related enzymes. In crustaceans in general, two types of enzymes are recognised : alkaline phosphatase and acid phosphatase. In Scylla serrata, the optimal activity of acid phosphatase is at pH 5.0 and that of alkaline phosphatase at pH 9.0 (Mercy, 1979). The probable function of the phosphatases is the transfer of the phosphate group from a donor substrate to an acceptor compound containing an (OH group). If the acceptor is water, the net effect is hydrolysis