Athos Ottolenghi

Interaction of ascorbic acid and mitochondrial lipides

Abstract Incubation of rat liver mitochondrial suspensions in the presence of low concentrations of ascorbic acid results in the formation of lipide peroxides and the oxidation of the ascorbic acid. There is a quantitative relationship between the two reactions. The effects of added iron salts and ethylenediaminetetraacetic acid (EDTA) on the system have been studied. The results suggest that the mechanism involved is non-enzymic and consists of a cooxidation of ascorbic acid and unsaturated fat mediated by a metal ion.

Effect of Ethylenediaminetetraacetate on Lipide Peroxide Formation and Succinoxidase Inactivation by Ultraviolet Light.

Summary 1. EDTA inhibits lipide peroxide formation in rat liver homogenates and in rat liver mitochondria exposed to ultraviolet irradiation. 2. Ultraviolet light inhibits succinoxidase in mitochondria and EDTA protects it from inactivation. The protection is proportional to the inhibition of lipide peroxide formation. 3. EDTA does not inhibit lipide peroxide formation in pure methyl linolenate emulsions irradiated with ultraviolet light but does inhibit both the antioxidant effect of ascorbic acid and the iron catalysis of peroxide formation in these emulsions.

The inhibition of certain mitochondrial enzymes by fatty acids oxidized by ultraviolet light or ascorbic acid.

Abstract 1. 1. The relation between lipide oxidation and the activity of succinoxidase, choline oxidase, and aminoxidase of rat liver mitochondria has been examined following ultraviolet irradiation, incubation of mitochondria with ultraviolet-irradiated methyl linolenate and methyl linoleate, and incubation with ascorbic acid. 2. 2. Ultraviolet irradiation of mitochondria produced an oxidation of lipide which was directly proportional to the period of irradiation. Succinoxidase and choline oxidase were inhibited in direct proportion to the extent of lipide oxidation. Incubation of irradiated mitochondria with reduced glutathione gave a partial ( 3. 3. Incubation of mitochondria with ascorbic acid resulted in an inhibition of succinoxidase, choline oxidase, and aminoxidase which was directly proportional to lipide oxidation, as in the case of ultraviolet irradiation. Liver slices incubated with ascorbic acid also showed an increased lipide oxidation and decreased succinoxidase activity. 4. 4. Succinoxidase was inhibited by ultraviolet-irradiated methyl linolenate and its water-soluble oxidation products. The unoxidized ester had little or no effect. Methyl linolenate and methyl linoleate oxidized to the same degree by ultraviolet irradiation had equal inhibitory action on succinoxidase. 5. 5. The results indicate that the effects of ultraviolet light on cells may be mediated in part by the oxidation products of lipide constituents.

Comparison of methods for determining fatty acid oxidation produced by ultraviolet irradiation

SummaryThe thiobarbituric acid (TBA) reaction for fatty acid oxidation has been compared with Lundberg and Chipaults method for peroxides, the Kreis test for aldehydes, and with the degree of conjugation, using fatty acid esters exposed to ultraviolet light for various periods. The TBA test paralleled the other methods for methyl linolenate and methyl linoleate but was essentially negative for methyl oleate oxidation.The sensitivity of the TBA test for linolenate was 30–80 times that for linoleate at the same peroxide values. The TBA test appears to be a reliable method of estimating the oxidation products of linolenic and linoleic acids in tissues and other biological material.

Studies on bone marrow lipid in normal and irradiated rabbits.

Fatty acid peroxides are formed when liver, kidney, and brain homogenates or slices are incubated aerobically (1). The peroxides inhibit certain mitochondrial enzymes (2), and the inhibition is prevented by previous addition of glutathione but only partially reversed by it (unpublished). Peroxides react slowly with enzymes in vitro, and when small amounts are injected into animals death occurs in 20 to 40 hours, depending on the dosage (unpublished). Peroxides also inhibit the growth of certain bacteria (3) and cell division in marine worm eggs (4). Because of these effects the organism must have mechanisms to prevent accumulation of peroxides under normal conditions. It seemed likely that such mechanisms would be particularly effective in tissues where cell division is occurring, and it has already been shown (5) that Ehrlich tumor cells produce no peroxides on incubation. In the present study peroxide formation has been investigated in normal, actively dividing bone marrow and in the marrow of X-rayed animals. The peroxides were estimated by the thiobarbituric acid reagent (6).

Trichinella spiralis: phospholipase in challenged mice and rats

Mice challenged with 400 larvae of T. spiralis showed greatly elevated phospholipase levels in tissues of the small intestine from 5 to 29 days after infection, after which the levels returned to those of the uninfected controls. Related to these rises in enzyme levels were greatly increased numbers of eosinophils in the bone marrow. Increases strikingly above those in the controls were noted from 8 to 20 days after infection. In rats challenged with 3000 larvae, the enzyme levels were elevated from 4 to 18 days, and the eosinophils were increased during the same periods. A working hypothesis is proposed to explain these phenomena.

Inhibition of cell division by ultraviolet irradiated unsaturated fatty acid

Abstract Ultraviolet irradiation of eggs of Chaetopterus pergamentaceous resulted in the formation of lipid peroxides as measured by the thiobarbituric color reaction. Aqueous extracts of ultraviolet irradiated methyl linolenate inhibited cleavage, reversed furrow formation, and reduced spindle size. These effects were also brought about by ultraviolet irradiation. Irradiated methyl linolenate also reduced the size of the fully formed spindle of the eggs of Lytechinus variegatus . It is suggested that at least a part of the action of ultraviolet irradiation is mediated through the formation of lipid peroxides.

Oral toxicity of the cyclic polyethers--12-crown-4, 15-crown-5, and 18-crown-6--in mice.

Abstract The acute and chronic oral toxicities of a homologous series of three crown ethers in mice have been studied. The LD50 values and the partition coefficients (octanol/water) of the compounds have been determined. The LD50 values are as follows: 12-crown-4, 3.15 g/kg; 15-crown-5, 1.02 g/kg; and 18-crown-6, 0.71 g/kg. The compounds follow a general trend of increasing toxicity with increasing ring size and water solubility. There appears to be no cumulative effect of daily ingestion of small amounts (one-half to one-third the LD50 value) of the compounds over a 2-week period.

Nippostrongylus brasiliensis: Phospholipase in nonsensitized and sensitized rats after challenge

Abstract Rats given an initial infection with Nippostrongylus brasiliensis showed greatly elevated phospholipase B levels in the small intestines and lungs from 8 through 22 days after challenge. The rise in enzyme concentration occurred earlier (Days 8–11) in the proximal half of the intestine, but at Days 22, 29, and 36 the levels were much higher in the distal segments. This shift in activity correlates with the known elimination of worms and a diminishing inflammatory response in the proximal areas. The increase in enzyme activity in the intestine and lungs was associated with an increased production of eosinophils in the bone marrow 11–22 days after challenge. Rats sensitized with one stimulating infection before challenge showed an anamnestic type of response, as measured by enzyme levels in the small intestines and lungs and by the numbers of eosinophils in the bone marrow. The results are discussed in light of our similar data reported earlier from animals infected with Trichinella spiralis .

Phospholipase activity of rat tissues and its modification by trypsin1,2

Treatment with proteolytic enzymes before the addition of the phospholipid substrate increases the activity of the phospholipases of the spleen, thymys, bone marrow, lung, and liver of the rat. In contrast, the phospholipase activity of the intestine, which is higher than that of all other normal tissues, is not increased when incubated with proteases. The results of fractionation studies by high-speed centrifugation and gel filtration and differences in enzyme kinetics support the conclusion that the intestinal phospholipases differ substantially from phospholipases found in the other tissues.