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Two forms of α-galactosidase, I and II, exist in Vicia faba seeds and these have been purified 3660- and 337-fold respectively. They behaved as homogeneous preparations when examined by ultracentrifugation, disc electrophoresis and gel filtration. The apparent molecular weights of enzymes I and II, as determined by gel filtration, were 209000 and 38000 respectively. The carbohydrate contents of enzymes I and II were 25% and 2·8% respectively, and the enzymes differed in their aromatic amino acid compositions. Enzyme I was split into six inactive subunits in the presence of 6m-urea. α-Galactosidases I and II showed different pH optima and Km and Vmax. values with p-nitrophenyl α-d-galactoside and raffinose as substrates, and also differed in their thermal stabilities.

Aim : To study the various ocular anatomical and physiological parameters in presbyopia. Materials and methods : We studied the various ocular anatomical and physiological parameters like corneal curvature (keratometry readings: K1 and K2), central corneal thickness (CCT), anterior chamber depth (ACD), lens thickness (LT) and axial length (AL) in 100 presbyopic patients between 35 - 55 years of age. The patients were divided into two age groups: I (35 – 44 years) and II (45-55 yrs). ACD, AL and LT were measured using an Ascan. CCT was measured with ultrasonic pachymetry. Results : The CCT decreased (BE), LT increased and ACD decreased (RE) significantly with increasing age (p < 0.05). There was no significant difference in males and females. Nearly 3/4th of the total increase in lens thickness was responsible for the decrease in the anterior chamber depth and the rest, 1/4th , goes posteriorly. Corneal curvature and AL showed no significant change with age. Conclusions : The mean of CCT decreased significantly with advancing age. As age increased, the mean value of lens thickness increased and anterior chamber depth decreased. Nearly 3/ 4th of total increase in LT was anteriorly, decreasing the ACD. Corneal curvature and AL has no relation with age. Key words : Presbyopia, central corneal thickness, anterior chamber depth, lens thickness, axial length DOI: http://dx.doi.org/10.3126/nepjoph.v3i2.5269 Nepal J Ophthalmol 2011; 3(2): 155-158

Aim: To study accommodation in relation to different refractive errors, amblyopia and to measure the anatomical changes in the accommodating eye Materials and methods: We studied the amplitude of accommodation (AA) in 150 patients in the age group 11 – 30 years which included emmetropes, myopes, hypermetropes and hypermetropic amblyopes using the Royal Air Force (RAF) rule. The anterior chamber depth (ACD), axial length (AxL) and lens thickness (LT) changes during accommodation were measured using an A-scan. Myopes and hypermetropes were further divided based on the amount of refractive error : < 2D, 2 -4D and > 4D. Results: Corrected low myopes had the highest accommodation amplitude (p < 0.05) followed by emmetropes. Corrected hypermetropes were found to have the lowest amplitude of accommodation (p < 0.05). The amblyopic eye had a significantly low AA compared to the non-amblyopic eye (p < 0.05). ACD decreased (p < 0.05) and LT increased (p < 0.05) during accommodation. The AxL increase was maximum in myopes (p < 0.05) followed by hypermetropes but the change was not significant in hypermetropes (p > 0.05). Conclusion: The amblyopic eye has low amplitudes of accommodation proving the benefit of near adds in amblyopic patients. Prolonged near work might induce myopia in susceptible eyes by increasing the axial length. Key words: accommodation, refractive error, amblyopia, biometric changes DOI: http://dx.doi.org/10.3126/nepjoph.v3i2.5267 Nepal J Ophthalmol 2011; 3(2): 146-150

The presence of essential catalytic groups in sweet almond α-galactosidase (α- d -galactoside galactohydrolase, EC 3.2.1.22) has been investigated with the use of metal ions and photo-oxidation. Competitive inhibition was shown to occur with both Hg2+ and Ag+. From the inhibition data it was postulated that carboxyl and histidine groups in the enzyme active site were responsible for binding the metal ions. The latter group was destroyed by photo-oxidation resulting in inactivation of the enzyme. The enzyme was found to catalyse hydrolytic, transgalactosylation and synthetic reactions on the same active site. All the reactions proceeded yielding products with complete retention of configuration. Two possible mechanisms of action have been suggested and discussed.

A new enzyme, β- l -arabinosidase, has been detected in the dormant seeds of Cajanus indicus . This enzyme is different from α-galactosidase and does not hydrolyse an α- d -galactoside or an α- d -fucoside.

1. The hydrolysis of a variety of galactosides and other glycosides by α-galactosidases I and II of Vicia faba was studied. 2. The effect of temperature on kinetic parameters was also examined. 3. Both enzymes are inhibited by excess of substrate (p-nitrophenyl α-d-galactoside); with enzyme I this is competitive and is caused by the galactosyl moiety. 4. Enzyme I is inhibited by oligosaccharides possessing terminal non-reducing galactose residues and to a smaller extent by l-arabinose and d-fucose. 5. The effect of pH on Km and Vmax. values suggests that carboxyl and imidazole groups are involved in the catalytic activity of enzyme I. 6. Photo-oxidation experiments with enzyme I also suggest that an imidazole group is present at the active site.

Dormant seeds from various higher plant species have been examined for multiple forms of α-galactosidase. The properties of corresponding forms from Pisum sativum and Vicia faba have been compared. The high molecular from (I) from P. sativum has been purified 2770-fold.

1. alpha-Galactosidase from sweet almonds was purified about 2000-fold through eight steps. 2. The enzyme preparation was free from other related enzymes known to occur in sweet almonds, and behaved as a homogeneous protein on filtration through Sephadex G-75. 3. A molecular weight of about 33000 was determined from the gel-filtration data. 4. The ultraviolet-absorption spectrum and thermal inactivation of the enzyme are described. 5. The purified enzyme hydrolysed p-nitrophenyl alpha-d-galactoside at a much faster rate than melibiose. 6. The pH optimum was at 5.5-5.7. 7. Besides hydrolysis, it also catalysed transfer of galactosyl residues, chain elongation of melibiose and the synthesis of oligosaccharides from galactose.

The effect of temperature and pH on the activity of sweet almond α-galactosidase (α-galactoside galactohydrolase, EC 3.2.1.22) has been studied with melibiose and β-nitrophenyl α- d -galactoside (PNPG). Activation energies for the two substrates were found to be 12.4 and 19.0 kcal/mole, respectively. The p K m vs. t /T plots were linear for both substrates, and the enthalpies of binding were −7.3 and −10.2 kcal/mole for PNPG and melibiose, respectively. Changes in K m and v max of melibiose with pH suggested the participation of a group with p K 5.75 in the enzymep-substrate binding and of another group with p K around 3 in the catalytic step. Similar studies with PNPG also revealed the presence of two groups ( p K 3.4 or above and 6.7 or below), the p K values of which were strongly influenced by the substrate concentration. The group dissociating in the lower pH range appears to play a minor part only in the PNPG hydrolysis. In spite of apparently marked differences in the effect of temperature and pH, all the substrates appear to be hydrolysed at the same site on the enzyme molecule. The differences probably arose from differences in the mechanisms of the reaction or in the nature of the rate-limiting step in the two cases.

Two forms of α-galactosidase have been isolated from Vicia faba seeds and their physical and chemical properties compared.

Cajanus indicus α- and β-galactosidase which are inseparable by Sephadex gel-chromatography (Dey, P. M. (1973) Biochim. Biophys. Acta 302, 393–398), have now been separated by CM-cellulose chromatography. α-Galactosidase (EC 3.2.1.22) has been resolved into two isoenzymes and β-galactosidase (EC 3.2.1.23) into three. Various properties of the enzymes have been studied which have shown that they possess their own characteristic features.

1. The specificity of purified sweet-almond alpha-galactosidase has been investigated with 17 substrates. 2. Some of them exhibited inhibition at high substrate concentrations but others did not. Both substrate types were bound and hydrolysed at the same site on the enzyme. 3. The enzyme is specific for alpha-d-galactosides and beta-l-arabinosides. It did not hydrolyse beta-d-galactosides or alpha-d-glucosides. 4. Among galactosides the order of decreasing rates of enzymic hydrolysis was: aryl alpha-galactosides; sugars; alkyl alpha-galactosides. 5. All substituents in the aryl moiety of aryl alpha-galactosides enhanced V(max.), the electron-releasing (-sigma) groups being more effective than the electron-withdrawing (+sigma) groups. The substituent groups did not alter K(m) appreciably. 6. Implications of these results are discussed from a mechanistic viewpoint.

Liver fats ofCarcharias limbatus andPristis cuspidatus from the West Coast of India have been studied. Lithium salt-acetone and lead salt-alcohol methods were adopted for the resolution of the fatty acids. An efficient, electrically heated and packed column has been employed for fractionation of the methyl esters of fatty acids. These fats have been found to have a high content of saturated fatty acids and thus belong to the fourth group of Tsujimoto's classification of Elasmobranch fish liver fats. Analyses of the liver oils from the same species of fish show that there is a tendeney towards saturation or hydrogenation of polyethylenic acids accompanied by an increasing concentration of unsaponifiable matter, which is mainly alcohol ethers of the selachyl type, in accordance with Lovern's views.