Phenoloxidase from the leaves of the cotton plant

P H E N O L O X I D A S E F R O M T H E L E A V E S O F T H E C O T T O N P L A N T

T . S . Y u n u s o v a n d P . K h . Y u l d a s h e v UDC 577.15.06

There is informat ion in the l i t e r a tu r e on the p re sence of phenoloxldase in the l eaves of the cotton plant [1].

We ex t rac ted the p ro te ins f r o m f re sh ly f rozen l eaves with a tenfold amount of 0.1 M c i t ra te -phospha te buffer , pH 7.2, in the p re sence of 1% of a sco rb ic acid and of Kapron powder - 0.5 g of powder p e r g of l eaves . The ex t r ac t was f i l te red and centr i fuged at 18,000 r p m for 15 rain, and the supernatant liquid was cl ia lyzedagainst dist i l led wa te r for a day. The phenoloxidase act ivi ty in the dialyzate was de te rmined by D r a w e r t ' s method [2], using as the subs t ra t e mix ture 2 ml of a 10% solution of catechol , 2 rrd of 2% prol lne, and 1 rnl of enzyme solution. After the lapse of 20 rain, the reac t ion was fixed by the addition of 0.5 ml of a 5% solution of t r i ch lo roace t i c acid. A change in the absorpt ion (violet f i l ter) by 0.001 in 1 rain was taken as the unit of act iv i ty .

To de te rmine the highest specif ic act iv i ty of the phenoloxidase in the leaves of the cotton plant, we studied the dynamics of the accumula t ion of this enzyme in young cotton shoots . The expe r imen t s were p e r f o r m e d on cot ton-plant l eaves col lected in the e a r l y per iods of vegetat ion in the Tashkent . b l a s t in 1972. No phenoloxidase was found in the seeds and the co ty ledon.us l ea f le t s . The specif ic act ivi ty of the phenol- oxidase was 7 in May, 14 in June, 50 in July (chopping period), 40 in August, and 10 in September . Con- sequently, the g r e a t e s t phenoloxidase act ivi ty is obse rved in the per iod of vigorous growth of the cotton plant.

In addition to the soluble phenoloxidase, we found in the leaves of the cotton plant a la tent phenol- oxidase which was ex t rac ted by the buf fer solution mentioned but only in the p re sence of the detergent Tr i ton ~-100 . All the opera t ions were p e r f o r m e d at a t e m p e r a t u r e of +4-6°C.

Thus, the op t imum conditions have been se lec ted for the ex t rac t ion of phenoloxidase f r o m the leaves of the cotton plant of va r i e ty 108-F, and the vegetat ion per iod with the g r e a t e s t specif ic act ivi ty of this en- zyme has been de te rmined .

1. 2.

L I T E R A T U R E C I T E D

E. E. King, Phytochem., 1._00, 23370971) . F. Drawer t and H. Gebbing, Naturwiss . , 50, 522 (1963).

Insti tute of the C h e m i s t r y of Plant Substances, Academy of Sciences of the Uzbek SSR. Trans la ted f r o m Khimiya Pr i rodnykh Soedinenii, No. 3, pp. 446-447, May-June , 1973. Original a r t i c le submit ted January 5, 1973.

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