|Year : 1980 | Volume
| Issue : 2 | Page : 73-75
Free amino acids in various stages of human cataractous lenses
BS Chauhan1, NC Desai1, Ramesh Bhatnagar2, SP Garg1
1 Department of Ophthalmology, S.N. Medical College, Jodhpur, India
2 Department of Chemistry, University of Jodhpur, Jodhpur, India
B S Chauhan
Deptt. of Ophthalmology, Medical College, Jodhpur (Rajasthan)
Source of Support: None, Conflict of Interest: None
|How to cite this article:|
Chauhan B S, Desai N C, Bhatnagar R, Garg S P. Free amino acids in various stages of human cataractous lenses. Indian J Ophthalmol 1980;28:73-5
Dische, has stressed that changes in the transparency of the lens are fundamentally associated with changes in the physical nature of the constituent proteins of the lens. These changes can consist of de-aggregation of the protein sub-units which go to make up the soluble proteins of the lens; such changes amounting in some cases to a denaturation of the proteins. They can also be associated with decrease in solubility of the proteins or alterations in the composition of such functional constituents of the lens as glutathione or mineral contents, such changes obviously tending to affect the integrity of the lens in a sequential manner.
The free amino-acids composition of lenses is derived from studies on experimental animals; calf, cat & monkey. The concentration of free amino acids in the lens is more than the aqueous humour which surrounds it. The relation of free amino acids in human lenses to cataract has also been reported.
We undertook the present study of free amino acid pattern qualitatively in immature, mature and hypermature cataractous human lenses by thin layer chromatography.
| Material and methods|| |
Human lenses at various stages of cataract formation were taken out intracapsularly.
These had been graded clinically, as seen on slitlamp examination (i) Immature cataract having some non-cataractous clear lens fibres (ii) Mature Cataract having entire lens showing cataractous changes. (iii) Hypermature Cataract - only Morgagnian cataracts have been studied in this group.
For experimental purposes lenses were divided into the following groups :
Group I - 7 lenses of Immature cataract.
Group II - 25 lenses of Mature cataract.
Group III - 10 lenses of Hypermature cataract.
Weighed lenses were homogenized in 5 ml. of 80% alcohol by grinding. The homogenized material was kept in refrigerator at 4°C for 24 hours. This material was centrifuged at 12,000 r.p.m. for 20 minutes. The supernatant liquid was transferred into another tube and boiled at 90'C for 30 minutes in a water bath. The residual solution after evaporation was measured by pipette. 30 I (micro liters) solution was spotted by lambda pipette on 20 x 10 Cms. thin layer silica plates. The plates were run with butanol : acetic acid : water (4:1:1) for 3-4 hours. The dried plates were sprayed with ninhydrin (0.1 mg/ 100ml of butanol) and were kept in hot air oven at 110°C for 30 minutes.
The amino acids were identified by comparing the Rf values of the unknown with the Rf values of standard amino acids under identical conditions.
| Results|| |
Although an almost same amount of lens was analysed in each case, the thin layer chromatographic patterns varied significantly.
The concentration of free amino acids in immature, mature and hypermature human cataractous lenses varies considerably [Figure - 1]. 6 free amino acids were present on the thin aver chromatographic plates, 5 of which were identified, namely cystine, threonine, glutamic acid, methionine and leucine. One ninhydrin positive spot could not be identified.
The results showed gradual decrease in the concentration of various free amino acids with respect to the advancement of cataract formation.
| Discussion|| |
Amino-acids have been shown to be actively transported from plasma into the posterior aqueous humour and from there into the lens (Kern, Kinsey et al,). It has been observed that the concentration of most of the free amino acids presents in the immature cataract decreases gradually with the development of cataract. The methionine was present in highest concentration in immature cataract while its concentration decreases immature and hypermature cataract. Similarly the concentration of leucine, unidentified amino acid, glutamic acid, cystine and threonine which seems to be higher in the immature cataract decreases in mature and hypermature cataract respectively.
Agarwal et a1 showed that threonine, glutamic acid, cystine are predominant free amino acids present in crystalline lenses. Our finding also supports the above study.
The findings in the present study show, therefore, that methionine, leucine, unidentified amino acid, glutamic acid, cystine, and threonine are significant in cataract formation. Therefore theory of cataract formation due to the deficiency of certain amino-acids such as leucine, histidine (Mann et a1) phenylalanine, valine (Hall et a1) does not appear to be very significant.
The observation of Bles, that methionine was responsible for lens opacity, support the present findings. The work of Schaeffer & Shankman, who reported about the high values of glutamic acid and glycine in young transparent lens, is in agreement with the present finding as concentration of these amino-acids decreases significantly with the lens opacity.
The absence of tryptophan in the present study shows that its role in cataract formation is insignificant. Gupta et al have also observed the absence of tryptophan in their study.
Calam et a1 reported unidentified ninhydrin positive compounds and Reddy also reported unidentified ninhydrin positive compounds. The present study also supports the above findings.
| Summary and conclusion|| |
Free amino acids of immature, mature and hypermature human cataractous lenses (42) have been identified by thin layer chromatographic method.
There was a gradual decrease in the concentration of various amino acids with respect to cataract formation.
Due to decrease of concentration of methionine, leucine, unidentified amino acid, glutamic acid, cystine and threonine, it was concluded that these amino-acids may be responsible for the cataract formation.
| References|| |
Dische, 1965, cited by Sorsby, 1,II,371, 1972.
Calam, D.H. and Waley, S.G., 1964, Biochem. J.93, 526.
Kirsten. G. and Dardenne, U. 1961, Ber. deutsch. ophthal. Gesellesch, 15, 474.
Reddy, D.V.N., 1967, Invest. ophthalmol. 6, 478.
Agarwal, L.P., Kwatra, M. and Mathur, RE., 1978, East. Arch. ophthalmol, 6, 88.
Kern, H L., 1962, Invest. ophthalmol. 1: 368,
Kinsey, V.E. and Reddy, D.V.N. 1962, Invest.ophthalmol 1: 355,
Kinsey, V.E. and Reddy, D.V.N,, 1963, Invest. ophthalmol 2,229.
Mann, I 1945. Cited by Pirie, A. VanHeyningen, R: "Biochemistry of the eye". Blackwell, oxford, 1956.
Hall, W. and Bowles, 1, 1948, J. Nutr. 36 277.
Bles, 1947, cited by Pirie, A. VanHeyningen. R. "Biochemistry of the eye", Blackwell oxford, 1956,
Schaeffer, A.J. and Shankeman, S; 1960 A.J.O. 33 :1049, cited by Angra, S.K. lenticular amino acid, Proc. of all India ophth. Soc. 29 :55-60, 1969.
Gupta, A.K., and Sarin, G.S., 1976, Ind. J. ophthalmol, 24, II, 1.
[Figure - 1]