Oxidative and Phosphorylative Activities of Mitochondria Isolated From Cotton Hypocotyls

Abstract

Abstract. Mitochondria isolated from 2 strains of cotton plant hypocotyls (Gossypium hirsutum L. var. Rex smootlh leaf and Rex glandless) were examined for their oxidative phosphorylation activities. Bovine serum albumin at a relatively high concentration was essential in the extraction medium for the isolation of oxidatively active mitochondria from both strains of cotton. Phosphorylation was obtained only with Rex glandless cotton mitochondria. This activity was low in comparison to the mitochondria isolated from soybeans (Glycine max L. var. Lee). The endogenous gossypol content was found to be much higher in the Rex smooth leaf tissue than in the Rex glandless tissue. In turn, comparable gossypol differences were found associated with their respective mitochondrial fractions. Exogenous gossypol uncoupled succinate oxidation with active mitochondria isolated from soybeans. Gossypol as a possible uncoupler is discussed and compared to carbonyl cyanide, m-chlorophenvl hydrazone and 2,4-dinitrophenol.Metabolically active mitochondria have been isolated and studied in a xvide variety of plant tissues (7), but verv little xvork has been reported on cotton mitochonclria. Throneherry (13) reported that an extract fromii cotton tissue inactivatedc mitochondria prepared fromii sovbean seedlin,gs. He showed that the inclusion of bovine serum albumin (BSA) in the extractioni medium facilitated the isolation of active mitochondria from cotton hypocotyls. Later, Throneberry (14) found that the phosphorylation mechanism was probably more sensitive to endogenous inhibitors than were the oxidative enzymnes. Plants of the genus Gossvpium, of which cotton is a member, normally containi several pigments unique to the plant kingdom. Some of the pigments are enclosed in glands that are distribuited throughout the above ground part of some strains of cotton. An important component of the glands is gossypol (1.1 ', 6,6',7,7'-hexah. droxy-3,3' -dimethvl-15,5' -diisopropvl-2,2' -binaphthyl-8,8'-dialdehyde), a polyphenolic. toxic, velloNv compound (2). MAvers and Throneherrv (9) reported that gossylpol affected mitochondria prepared from sweet potato and beef heart tissuies. Gossypol was showvn to inhibit oxidative phosphorylation at concentrations which had no effect on oxidation. This prompted the investigation of a glandless strain of cotton. relatively free of gossypol. Rex glan(dless, hereafter referred to as glanded and glandless, respectively) and soybeans (Glycine mlax L. var. Lee) were used as the experimental material. The seeds were surface sterilized with 0.5 % (w/v) sodium hvpochlorite. After rinsing thoroughly, the seeds wvere germinated in paper towels, moistened w\ith n0.1 mAi CaCI.., at '0O in the dark. Five-dav old hypocotyls were harvested, cut into 3 to 5 cm segments, and chilled at 0 .A slight modification of the Bonner and Sikes procedure (1) was used for mitochondria isolation.The hypocotyl segments were disrupted in a WaringBlendor at a reduced speed using a volume of extraction medium equal to the weiglht of ...

Some Factors Affecting the Hill Reaction Activity in Cotton Chloroplasts

“…Cotton chloroplasts isolated in a sucrose-phosphate medium exhibited low rates of ferricyanide reduction when assayed for the Hill reaction ( Table I). The findings are similar to reports of low oxidative activity in mitochondria which were isolated from cotton hypocotyls in a sucrose medium (3,7). These authors assumed that an endogenous inhibitor was present, possibly gossypol, occurring primarily in the pigment glands of the cotton hypocotyls.…”

“…Recent reports show that when adsorptive agents are added to the isolation medium, it was possible to isolate particles with higher activities. Mitochondria with active enzyme (7) and electron transport systems (3) have been isolated from cotton hypocotyls when bovine serum albumin was added. Active enzymes have been isolated from cotton roots, leaves, and cotyledons in the presence of an insoluble form of polyvinylpyrrollidone (H. B.…”

Electron Transport in Plant Respiration

“…New isolation procedures of plant mitochondria are thus still appearing in the literature (e.g. 24,47,50,57,59,75,76,83,87,89,91). The preparatory problems may strengthen the view presented by Packer et al (82) In this article the ensuing discussions will be limited to the electron trans fer system in plant mitochondria and to an intriguing problem on the devel opment of cyanide resistance in storage tissues.…”

Properties of Wheat Mitochondria. Study of Substrates, Cofactors and Inhibitors

“…Mung bean hypocotyl mitochondria oxidize succinate, L-malate and externally added NADH with good respiratory values (Ikuma and Bonner 1967 a). Other workers have prepared mitochondria which utilize a-ketoglntarate, succinate, lio-citrate and pyruvate plus malate (Lance et al 1965, Hanson et al 1965, Drury et al 1968, Killion et al 1968).…”