2,4-Dinitrophenol, uncoupler oxidative phosphorylation

A. Pentachlorophenol and dinitrophenols uncouple oxidative phosphorylation in the mitochondria. Substrates are metabolized but the energy produced is dissipated as heat instead of producing adenosine triphosphate (ATP). The basal metabolic rate increases, placing increased demands on the cardiorespiratory system. Excess lactic add results from anaerobic glycolysis. 

Dinitrophenol uncouples oxidative phosphorylation by leaking protons Into the matrix... 

Dinitrophenol is a member of the aromatic family of pesticides, many of which exhibit insecticide and fungicide activity. DNP is considered to be highly toxic to humans, with a lethal oral dose of 14 to 43mg/kg. Environmental exposure to DNP occurs primarily from pesticide runoff to water. DNP is used as a pesticide, wood preservative, and in the manufacture of dyes. DNP is an uncoupler, or has the ability to separate the flow of electrons and the pumping of ions for ATP synthesis. This means that the energy from electron transfer cannot be used for ATP synthesis [75,77]. The mechanism of action of DNP is believed to inhibit the formation of ATP by uncoupling oxidative phosphorylation. 

Answer C. The toxic agent (example, 2,4-dinitrophenol) would uncouple oxidative phosphorylation, leading to a fall in ATP levels, increased respiration, and increased substrate utilization. 

Toxicology. 2,4-Dinitrophenol (2,4-DNP) uncouples oxidative phosphorylation from electron transport, resulting in diminished production of ATP, with the energy dissipated as heat, which can lead to fatal hyperthermia. ... 

Moreover, uncoupHng experiments using Escherichia coli cells were described. An addition of glucose to stationary E. coli cells leads to an increase of fluorescence intensity in the region of NAD(P)H, because it is formed in the glycolysis. This increase was stopped abruptly by addition of 2,4 dinitrophenol (DNP), which effects a decrease in the fluorescence signal according to the theory of uncoupled oxidative phosphorylation. The dynamics of this process. 

In an attempt to determine the best treatment regimen for mice given intraperitoneal doses of 4,6-dinitro-o-cresol (DNOC), which, like 2,4-DNP, uncouples oxidative phosphorylation and is hyperthermic, the effect of hypothermic dinitrophenols (i.e., 2,3-DNP, 2,5-DNP, and 3,4-DNP) on the lethality of DNOC was studied (Harvey 1959). At a dose of 10 mg/kg, DNOC itself resulted in 100% mortality. When the hypothermic DNPs were given immediately after DNOC, mortality was 60% after 2,3-DNP, 100% after 2,5-dinitrophenol, and 50% after 3,4-dinitrophenol. 2,4-DNP, which, like DNOC, is hyperthermic, afforded no protection of the mice. 

Dinitrophenol produces a prolonged choleresis (S39), but the BSP excretion rate remains the same or is slightly elevated (S40). This compound also causes a marked pyrexia, but choleresis is not due to hyperthermia since no change in bile flow occurs when animals are warmed to 42°C (S40). A wide range of related compounds have been tested (P12) phenols and mononitrophenols had no effect isomeric dinitro-phenols had less effect picric acid and 2,4-dinitrophenetole were as effective. 2,4-Dinitrophenol is known to uncouple oxidative phosphorylation, but it is difficult to correlate a decrease in ATP with an increased bile flow. Moreover, the 3,5 isomer which is a more potent uncoupler (B59) has a smaller choleretic effect. 

Because of the role of mitochondria in cellular respiration and energy production, efforts to elucidate the mechanism of thyroid hormone action in metabolism and calorigenesis have focused on mitochondrial studies. Thyroid hormones in vitro are known to uncouple oxidative phosphorylation in isolated mitochondria, but these effects occur at unphysiological doses of T4. In physiological concentrations, T4 increases adenosine triphosphate (ATP) formation and the number and inner membrane surface area of mitochondria (21), but T4 does not reduce the efficiency of oxidative phosphorylation. Furthermore, 2,4-dinitrophenol, a classic uncoupler of oxidative phosphorylation, can neither relieve hypothyroidism nor duplicate other physiological effects of thyroid hormones. 

The cyanophenol herbicides arrived as an intelligent replacement for Truffaut s dinitro-o-cresol. A typical example is ioxynil (6.71) 2,6-di-iodo-4-cyanophenol (Wain, 1963). It is a contact herbicide which powerfully uncouples oxidative phosphorylation (Section 4.5), in fact much more strongly than 2,4-dinitrophenol does (Kerr and Wain, 1964). It and the less expensive, if slightly slower acting, 2,6-dibromo-4-cyanophenol (bromoxynil), are used against young dicotyledonous weeds in cereal crops. They are also available as the octoate esters, for better penetration. 

Figure 14.1 Dinitrophenol and uncoupling protein uncouple oxidative phosphorylation from electron transport.

A pathway for fatty acid activation, involving a reaction with nonphosphorylated high-energy intermediates rather than the formation of acetyl-CoA derivatives has also been postulated. The supporting evidence includes the observations that (1) blocking the electron transport chain with cyanide or uncoupling oxidative phosphorylation with dinitrophenol interferes with the fatty acid oxidation (2) oligomycin, which blocks the biosynthesis of ATP but does not affect the formation... 

Dinitrophenol causes cataracts In humans, rabbits, and chicks however, this effect is not necessarily related to its ability to uncouple oxidative phosphorylation. 

Although there are several mechanisms by which respiratory electron transport can be inhibited, the compounds in Figure 5.2 are classed as inhibitory uncouplers. " This activity is not restricted to mitochondrial electron transport as photosystem II (PSII) electron transport and phosphorylation are also affected by dinitrophenols and hydroxybenzonitriles (Refs. 3-5 and Chapter 1). In isolated mitochondrial preparations, lower concentrations of herbicides uncouple oxidative phosphorylation, that is, promote electron transport, without ATP synthesis, whereas higher concentrations inhibit electron transport. Fedtke calculated that uncoupling is 2-10 times more sensitive for a range of phenolic/acidic compounds." A similar difference has also been reported for dinoterb (Figure 5.2), which uncouples oxidative phosphorylation at 0.05-0.15 mM and inhibits respiration at 0.5-1.0mM. ... 

Many inhibitors of substrate oxidations, substrate transport, electron transport, and ATP synthesis are known including many well-known toxins (see Sherratt, 1981 Harold, 1986 Nicholls and Ferguson, 1992). These are not discussed here except to mention specific uncouplers of oxidative phosphorylation. Classic uncouplers such as 2,4-dinitrophenol have protonated and unprotonated forms, both of which are lipid soluble and cross the inner mitochondrial membrane discharging the proton gradient. This prevents ATP synthesis and stimulates respiration. 

Figure 12-8. Principles of the chemiosmotic theory of oxidative phosphorylation. The main proton circuit is created by the coupling of oxidation in the respiratory chain to proton translocation from the inside to the outside of the membrane, driven by the respiratory chain complexes I, III, and IV, each of which acts as a protonpump. Q, ubiquinone C, cytochrome c F Fq, protein subunits which utilize energy from the proton gradient to promote phosphorylation. Uncoupling agents such as dinitrophenol allow leakage of H" across the membrane, thus collapsing the electrochemical proton gradient. Oligomycin specifically blocks conduction of H" through Fq.

Dinitrophenol (1 mM) and potassium cyanide (KCN) (5 mM) are used as uncouplers of oxidative phosphorylation from electron transport (105). 

Dinitrophenols had been used as insecticides since 1892 but it was not until the 1930s that their value as herbicides was discovered and 4,6-dinitro-o-cresol (DNOC) was introduced. The trouble with dinitrophenols was their toxicity to all living organisms that respire. Their mode of action is through the uncoupling of oxidative phosphorylation, an effect that leads to a rapid death of any organism that comes into contact with the chemical, including the operator. 

Depletion of ATP is caused by many toxic compounds, and this will result in a variety of biochemical changes. Although there are many ways for toxic compounds to cause a depletion of ATP in the cell, interference with mitochondrial oxidative phosphorylation is perhaps the most common. Thus, compounds, such as 2,4-dinitrophenol, which uncouple the production of ATP from the electron transport chain, will cause such an effect, but will also cause inhibition of electron transport or depletion of NADH. Excessive use of ATP or sequestration are other mechanisms, the latter being more fully described in relation to ethionine toxicity in chapter 7. Also, DNA damage, which causes the activation of poly(ADP-ribose) polymerase (PARP), may lead to ATP depletion (see below). A lack of ATP in the cell means that active transport into, out of, and within the cell is compromised or halted, with the result that the concentration of ions such as Na+, K+, and Ca2+ in particular compartments will change. Also, various synthetic biochemical processes such as protein synthesis, gluconeogenesis, and lipid synthesis will tend to be decreased. At the tissue level, this may mean that hepatocytes do not produce bile efficiently and proximal tubules do not actively reabsorb essential amino acids and glucose. 

Certain foreign compounds can cause changes in body temperature, which may become a toxic response if they are extreme. Substances such as 2,4-dinitrophenol and salicylic acid will raise body temperature, as they uncouple mitochondrial oxidative phosphorylation. Thus, the energy normally directed into ATP during oxidative phosphorylation is released as heat. Substances that cause vasodilation may cause a decrease in body temperature. 

In the mammal, complex polysaccharides which are susceptible to such treatment, are hydrolyzed by successive exposure to the amylase of the saliva, the acid of the stomach, and the disaccharidases (e.g., maltase, invertase, amylase, etc.) by exposure to juices of the small intestine. The last mechanism is very important. Absorption of the resulting monosaccharides occurs primarily in the upper part of the small intestine, from which the sugars are earned to the liver by the portal system. The absorption across die intestinal mucosa occurs by a combination of active transport and diffusion. For glucose, the aclive transport mechanism appears to involve phosphorylation The details are not yet fully understood. Agents which inhibit respiration (e.g., azide, fluoracetic acid, etc.) and phosphorylation (e.g., phlorizin), and those which uncouple oxidation from phosphorylation (e.g., dinitrophenol) interfere with the absorption of glucose. See also Phosphorylation (Oxidative). Once the various monosaccharides pass dirough the mucosa, interconversion of the other... 

Structures of two uncouplers of oxidative phosphorylation, 2,4-dinitrophenol (DNP) and carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP). DNP has a weakly dissociable proton on the oxygen atom FCCP has a similar proton on one of the nitrogens. 

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