New antidote

A NEW ANTIDOTE FOR CYANIDE POISONING ‎ ‎‎ By Dr. Mohammad Ali kavianpour Kermanshah medical university Iran May,2008 ‎

INTRODUCTION This paper describes a study of the antidotal actions of nickel nitrate against cyanide ‎poisoning. ‎ Hydrocyanic acid acts as a poison because it combines with, and stops the function of, ‎the iron atom in cytochrome oxidase, so blocking the electron transfer through the ‎cytochrome system, and checking the final oxidations involving oxygen uptake of the ‎tricarboxylic acid cycle. It also inhibits many other enzymes.Attempts to find an antidote ‎for cyanide have usually been in the direction of breaking down the cyanide-cytochrome ‎complex, and have looked for substances with a stronger affinity for cyanide ions than ‎has the oxidase.One such method is to convert some of the blood haemoglobin into ‎methaemoglobin,for example,by injection of a nitrite; the methaemoglobin combines ‎with the cyanide ion to form the very stable cyanmethaemoglobin. This has the drawback ‎that it involves the loss of oxygen-carrying power of the blood, so that, for example, to ‎antidote twice the LD50 of cyanide would be equivalent to the loss of about 12% (or in ‎an adult man 550 ml.) of the blood.‎ METHODS

The LD50 of hydrocyanic acid, by various routes of administration, was determined for ‎mice and rabbits. A solution of sodium cyanide was neutralized with acetic acid to about ‎pH 7,the content of hydrocyanic acid was estimated by silver titration, and the subsequent ‎dilution to the required concentration was made with 0.9% saline.At pH 7 the ‎hydrocyanic acid is about 1% ionized. In some experiments pure hydrocyanic acid was ‎used, but this had no advantage and was more difficult to handle. Owing to the volatility ‎of hydrocyanic acid, the solutions were kept in special vessels and, when any doubt ‎existed as to the final concentration, were again titrated at the end of the day’s ‎experiments. It was usually found that after 6 hr the concentration had fallen, often by ‎‎5%, and sometimes in hot weather by as much as 12%. ‎ Intravenous injections were made into the tail layer in mice and an ear layer in rabbits; ‎intramuscular injections were into the thigh muscles. Doses are expressed usually in ‎terms of umoles/kg of body weight, and for hydrocyanic acid sometimes also in terms of ‎LD50. Owing to the rapidity of action of hydrocyanic acid when large doses are given, ‎and to the difficulty of giving intravenous injections when the animal is convulsing, it ‎was more usual to give the antidote first, intravenously, and then to follow this in a matter ‎of seconds by the hydrocyanic acid, usually by intramuscular injection, but in some ‎experiments that order was reversed; the antidotal effects were at least as great, and often ‎greater, when that was done, provided the antidote was given soon enough. In order to ‎indicate the molar relationship between the hydrocyanic acid and the antidote in each ‎experiment, the ratio between them was usually expressed (moles of hydrocyanic acid per ‎kg divided by moles of nickel compound per kg) and is called the cyanide/nickel ratio.‎ RESULTS The effectiveness of a nickel nitrate, against hydrocyanic acid was examined mainly on ‎mice and rabbits.for rabbits (4 x LD50), was effective at a molar cyanide/nickel ratio of ‎‎4. and for mice (3 x LD50 at a molar ratio of 2), with no apparent side effects. At a molar ‎ratio of 1, it was slightly more effective than on rabbits.‎ Nickel nitrate by mouth was effective against orally administered hydrocyanic acid. The ‎oxygen uptake of the body, reduced by cyanide, improved when nickel antidotes has been ‎successfully administered. Relatively small amounts of nickel salts would be needed for ‎the neutralization of several fatal doses of cyanide.‎ By intravenous injection.The LD50 for mice was found to be about 42 umoles/ kg, and ‎for rabbits 33 umoles/kg. Mice given two to three times the LD50 convulse in a few ‎seconds and die in 50 seconds.By intraperitoneal injection,the results showed a rather ‎large spread, the mean LD50 for rabbits being 52 ,umoles/kg, and for mice 107 umoles/ ‎kg, so that by this route mice were only about half as sensitive as rabbits.This is probably ‎due to a more rapid detoxification by the mouse liver.It will be expected that toxicity ‎would be maximal when the cyanide is given quickly into a vein which drains into the ‎inferior vena cava, so that it is quickly delivered to the central nervous system, less toxic ‎when it enters more slowly, as by intramuscular injection, and least toxic when it enters ‎the circulation slowly, as when given intraperitoneally, subcutaneously or orally. the ‎descending order of toxicity proved to be intravenous intramuscular-intraperitoneal-‎subcutaneous-oral, The results of inhalation are more vague, but are probably comparable ‎with those by intravenous injection.A method of administration often used, but not ‎employed in the present investigation, is to anaesthetize the animal and to inject the ‎cyanide intravenously at a constant slow rate of about 0.1 mg/kg/min.The results so ‎obtained do not seem to differ greatly from those of the usual procedure for intravenous ‎injection. ‎ The nickel ion is known to be toxic to some micro-organisms, to depress metabolism in ‎tissue slices and, under certain conditions, metallic nickel can be carcinogenic, though ‎this last action has not been demonstrated for the nickel ion.‎ In acute experiments, nickel salts given intravenously cause a fall, followed by a rise and ‎a later fall, of arterial pressure, an increase in breathing, cramps, vomiting and acute ‎diarrhea with intestinal inflammation.‎ These effects have been analyzed and shown to be partly due to central action and partly ‎to peripheral effects. Death is due to respiratory failure when the administration is rapid, ‎to cardiac failure when more slowly given.‎ In more chronic administrations the principal effects are polycythaemia, porphyrinuria, ‎increase in the size of the adrenals, and goitre.Nickel salts have been given orally to ‎human subjects in doses up to 140 mg daily without clearly pronounced harmful effects. ‎Orally administered nickel is very slightly absorbed, and mainly excreted in the faeces; ‎injected intravenously it is mainly, and rapidly, excreted in the urine; according to ‎experiments what remains in the body is to be found mainly in liver, kidneys, pancreas ‎and spleen.In the present experiments, no evidence could be found that nickel was ‎present in any tissues or organs 14 hr after intravenous injection. Post mortem ‎examination showed intense congestion of the mucosa of the intestinal tract.On the rabbit ‎isolated ileum preparation the effect was relaxation, with diminution of the rhythmic ‎contractions on the rectum in situ intravenous injection caused powerful rhythmic ‎contractions, and these probably account for the colic and diarrhea which often follow ‎administration of nickel salts.‎ The toxicity of nickel compounds is related to the ease with which they yield nickel ions, ‎and hence is greatest in salts of nickel. Nickel nitrate yields ions readily; and the ‎compound would be expected to be somewhat toxic. ‎ Exactly what occurs when nickel and cyanide ions are introduced into the body is ‎uncertain; while being at the same time only slightly toxic (LD50=1,000 mg/kg). nickel ‎ions can reach the brain, that is they can penetrate the blood-brain barrier, have no ‎precipitating action on proteins, and, when introduced into the blood stream, are ‎eliminated by the kidney, which is not damaged.The opposition between nickel and ‎cyanide ions can be illustrated by experiments with isolated tissues. ‎ Some experiments were made to find whether nickel ions were able to reverse the ‎inhibition of the cytochrome oxidase system produced by cyanide.using p-‎phenylenediamine as the oxidation substrate.Only about 10% of the initial oxidase ‎activity was restored, Since the lethal dose of nickel salts is around 110 umoles/kg, the ‎amount given as an antidote has mostly been lower than that; but that dose should be able ‎to antidote 550 umoles of hydrocyanic acid, if the theoretical ratio holds good, and the ‎mutual antidotal action should in theory reduce any risk of nickel poisoning.‎ This would mean that, for a safe dose of a nickel salt, some 10 x LD50 of hydrocyanic‎ acid intraperitoneally for rabbits, and 5 x LD50 intraperitoneally for mice,should be the ‎maximum amounts that could be antidoted.In experiments done the maximum dose ‎antidoted was 10 x LD50, in a goat.The present series, using mice and rabbits, gave ‎results which are satis- factory for rabbits, but for mice fall short of the theoretical ‎predictions mentioned above. ‎ When the molecular ratios of cyanide and nickel are calculated, it is seen that, in rabbits, ‎the ratios are spread over the range 1.5 to 13. then in all cases there should be no ‎hydrocyanic acid left free. In the instances where an excess would be left it would be ‎only a fraction of an LD50, and this should have been nonlethal, but in one instance ‎‎(cyanide/nickel ratio of 12) was not, which indicates that the case is not so simple as it ‎first seemed.The results show that the nickel salt was more effective as an antidote for ‎rabbits than for mice.‎ For rabbits, provided the molar cyanide/ nickel ratio does not exceed 6, it seems that at ‎least 6 x LD50 can be neutralized, while at a ratio of 12 even twice the LD50 is not ‎antidoted. It should be noted, however, that, in the rabbit series, hydrocyanic acid was ‎usually given intravenously, whether before or after the nickel.‎ With mice that are less sensitive to hydrocyanic acid, and about as sensitive to nickel, as ‎are rabbits, and the results are less regular. It appears from the results that, under the most ‎favorable conditions, only upwards of twice the LD50 can be antidoted, and then only ‎when the cyanide/nickel ratio is below 5. The hydrocyanic acid was given ‎intraperitoneally and the nickel solution intravenously in this species, which might have ‎affected the results.‎ Thiosulphate alone had antidotal action which varied; for mice the value was around 2 x ‎LD50 at the best, but was irregular, and in rabbits it was also around twice the LD50. The ‎results are rather irregular, but show that, for mice, 3 x LD50 can be antidoted when the ‎nickel compound is given with glucose, and when thiosulphate is also given. ‎ Some of the cyanide/nickel ratios were very high, as much as 20, but, as thiosulphate was ‎also given, much of the beneficial effect must have been due to that. For rabbits the ‎results were similar, but were also improved by giving thiosulphate.‎ The most favorable condition for the action of the nickel antidotes would be expected to ‎be if they were mixed with the hydrocyanic acid before administration.It would be ‎expected that, if the resulting compound were not toxic and held the cyanide firmly ‎enough in competition with cytochrome oxidase, the effect of the hydrocyanic acid would ‎be entirely eliminated if enough antidote were present, and that, for instance, nickel salts ‎would be able to fix more molar equivalents of hydrocyanic acid, ‎ A simple experiment shows that the complex formed by nickel salts though not harmless ‎is not very toxic.Solutions of nickel nitrate and sodium cyanide were mixed in such ‎proportions that the mixture contained 260 ,umoles of cyanide and 45 ,umoles of nickel ‎salt per ml. This was injected into an ear vein of a rabbit in a dose of 1 ml./kg. This ‎would be, in terms of hydrocyanic acid, about 8.5-times the LD50. For some minutes ‎after injection there was panting and convulsions, followed by paresis of the hind limbs, ‎and loss of consciousness, as would have been the effects of, say, half a lethal dose of ‎hydrocyanic acid; but after 5 min, apart from a little panting, the animal was apparently ‎normal and made an uninterrupted recovery, with no subsequent diarrhea. We might ‎suppose, since the cyanide/nickel ratio was less than 6 that these transitory symptoms of ‎cyanide poisoning were to be attributed to slight dissociation of the nickelocyanide, but in ‎any case the experiment shows that there are grounds for the belief that the nickel ion ‎can, under the most favorable conditions, antidote the theoretical amount of cyanide.‎ These expectations were only in part borne out by experiment on mice, These results ‎show that when the dose of hydrocyanic acid is low (3 x LD50 or less) and the ‎cyanide/nickel ratio is 6 or less, the expectations from theory hold approximately, but ‎with four or more times the LD50 and cyanide/ nickel ratios above 4 the results are less ‎good.‎ In the experiments the oxygen utilization of rats anaesthetized with a mixture of ‎allobarbitone and urethane was measured by the use of a small-scale respirometer, and ‎the effect of an intraperitoneal dose of hydrocyanic acid, as large as possible without ‎causing arrest of respiration, was observed; nickel derivatives of various kinds were then ‎injected intravenously, and the subsequent oxygen usage measured over a sufficient ‎period. In several experiments, the nickel injection was given too late, but when the ‎animal survived the nickel injection restoration of oxygen usage was prompt and ‎complete and was often followed by increased usage, presumably the expression of ‎oxygen lack.Interaction between nickel salt and hydrocyanic acid given orally when ‎hydrocyanic acid or a cyanide is taken by mouth, one obvious line of treatment would be ‎to give a nickel salt by mouth immediately, or as soon as possible after any appropriate ‎intravenous therapy had been applied.‎ An experiment was carried out by giving hydrocyanic acid by stomach tube to mice, and ‎then following this up after an interval of time by administering by the same route a ‎dilute nickel nitrate solution. ‎ It would seem that nickel nitrate solution, given by mouth within a short time of ingestion ‎of cyanide, might be helpful; it would normally be followed by a wash-out of the ‎stomach, or by chelating any excess of nickel and there would then be little danger from ‎toxic effects due to absorption of nickel into the blood stream ‎ DISCUSSION The consequences of administration of cyanide depend mainly on the rate at which it ‎enters the blood stream, maximum effect resulting when the dose is high and the rate of ‎entry rapid, as for example, when a high concentration of hydrocyanic acid vapor is ‎breathed or when a large dose is injected into a systemic vein, in either of which cases ‎serious symptoms ensue in a matter of seconds and death within minutes. Even if ‎attempts at recovery appear to be successful, permanent damage to the central nervous ‎system may remain if the period of tissue anoxia has been more than a few minutes.‎ As the rate of entry of cyanide into the circulation is slowed, by slower administration ‎, another factor enters into consideration, namely, the rate of destruction of cyanide in the ‎body by conversion into thiocyanate, a change which takes place mainly in the liver by ‎the action of the enzyme rhodanese. ‎ The use of thiosulphate depends on the provision by it of available sulphur for this ‎reaction; the change seems to be somewhat slow, so it is better as a prophylactic than as ‎an antidote. Since cyanide when given orally or intraperitoneally enters initially into the ‎portal circulation, it will go direct to the liver, so that destruction is facilitated and larger ‎doses can therefore be tolerated. If the rate of eventual entry into the systemic circulation ‎is slow enough, as on subcutaneous, oral or respiratory administration.of small doses, the ‎effects are less serious, or may even be absent altogether. The mode of administration ‎therefore has a definite effect on the LD50, and as shown, the LD50 in increasing order is ‎intravenous-intramuscular-intraperitoneal -subcutaneous-oral, with inhalation, which is a ‎common industrial risk, comparable with administration by the intravenous route. ‎Another factor which influences the outcome is whether the antidote precedes or follows ‎the administration of the cyanide and, if the latter, after what interval of time. It would be ‎expected that the maximum antidotal effect would be seen when poison and antidote ‎were mixed before administration,and next to this when the two were injected ‎simultaneously into the circulation, or when antidote preceded the cyanide by a very short ‎time; the minimum effect would be seen when the antidote was given slowly, for ‎example subcutaneously, and at a time when the animal was near to death, when it would ‎be all but useless. In nearly every instance in the present experiments, the antidote was ‎administered intravenously, but intraperitoneal injection has also been found effective. ‎For practical reasons the nickel antidote was usually given to mice intravenously a few ‎seconds before the dose of hydrocyanic acid, intraperitoneally or intramuscularly. The ‎toxic action of nickel ions is the probable reason for this, but this is not great and would ‎largely be offset by the mutual detoxification of the two reactants. The number of mean ‎lethal doses of hydrocyanic acid which can be antidoted by nickel ions is greater for ‎rabbits than for mice, namely five to six for rabbits and two for mice, with maximal ‎cyanide/nickel ratios of 5 and 4 respectively. One reason for this is that mice are less ‎sensitive to hydrocyanic acid than are rabbits, while to nickel ions they are about equal in ‎sensitivity. Doses of nickel nitrate greater than 50 umoles/kg, though lowering the ‎cyanide/nickel ratio, were unsatisfactory for mice. Another reason in most of the present ‎experiments is that in rabbits both substances were given intravenously, and so had more ‎opportunity of reacting.‎ The fact that the cyanide/nickel ratio can be, with rabbits, effective at ratios as high as 5, ‎indicates that the changes involved in the antidotal reaction are much as expected from ‎theoretical considerations; and the fact that, with high doses of hydrocyanic acid the ‎effective cyanide/nickel ratio is usually lower, is probably to be explained on the basis ‎that the difference between the affinities of the cyanide ions for cytochrome and for ‎nickel are not very large.‎ ‎ The results of the present experiments indicate that in that respect there is not much to ‎choose between the two.The more favorable results which have been obtained with the ‎compound by present a disagreement which must be explained; one explanation being ‎that in those experiments cyanide and antidote were given almost simultaneously by ‎intravenous injection, which, gives very favorable conditions for reaction.One fact is ‎against the view that the compound is to be regarded as an ionizable salt of nickel, ‎despite the similarity of their respective toxicities when expressed in terms of molarity; ‎ ‎ Ratio at effectivity when no thiosulphate is given is much lower than with nickel. This is ‎also shown for the experiments with previous mixing, and bears out the findings of that ‎the reaction is complete at a molar ratio of about 2, , then the compound is not an ‎ordinary ionizable nickel salt, or it reacts with some blood constituent, or else it was not ‎obtainable in as pure a state.‎ The object of this investigation was to form an opinion as to the best antidote and method ‎of treatment for cyanide poisoning. First, as to the amounts of the various antidotes which ‎would be required for the treatment of a man who had received one LD50 of cyanide, say ‎‎50 mg of hydrocyanic acid, ‎ The question is, whether a dose of nickel nitrate of the order of 100 mg would be safe to ‎give intravenously, taking into account the probability that most of its toxicity would be ‎neutralized by the cyanide. The balance of evidence is that it would be a reasonably safe ‎dose under the circumstances. And then to give an intravenous injection of hypertonic ‎glucose.The use of oxygen administration has been advised,although the only explanation ‎of its value which is offered is that it helps to overcome the large oxygen debt which is ‎seen on recovery from the immediate effects of the poison; at all events, it should be ‎tried. So also should the administration of nickel nitrate by mouth, in cases where cyanide ‎has been ingested. ‎

ABSTRACT The effectiveness of a nickel nitrate, against hydrocyanic acid was examined mainly on ‎mice and rabbits.For rabbits (4 xLD50),was effective at a molar cyanide/nickel ratio of 4. ‎For mice (3 x LD50 at a molar ratio of 2), with no apparent side effects.‎ At a molar ratio of 1, it was slightly more effective than on rabbits.Nickel nitrate by ‎mouth was effective against orally administered hydrocyanic acid. The oxygen uptake of ‎the body, reduced by cyanide, improved when nickel antidotes has been successfully ‎administered.Relatively small amounts of nickel salts would be needed for the ‎neutralization of several fatal doses of cyanide.‎

KEY WORDS ‎ Antidote / Nickel nitrate / Cyanide poisoning ‎

REFFERANCES ‎Ambrose AM, Larson PS, Borzelleca JF, and Hennigar GR, Jr. 1976. Long term ‎toxicologic assessment of nickel in rats and dogs. J. Food Sci. Technol. 13: 181-187.‎ Broder I, Smith JW, Corey P, Holness L. Health status and sulfur dioxide exposure of ‎nickel smelter workers and civic laborers. J. Occup. Med. 31(4):347-353.‎ CARB. 1999. Air toxics emissions data collected in the Air Toxics Hot Spots Program CEIDARS Database as of January 29, 1999.‎ CDTSC. 1985. California Department of Toxic Substances Control. Review: Nickel and compounds. 11/15/85.‎ Cirla AM, Bernabeo F, Ottoboni F, and Ratti R. 1985. Nickel induced occupational ‎asthma:‎ Immunological and clinical aspects. In: Brown SS, and Sunderman FW. (eds.) Progress ‎in Nickel Toxicology. Boston, MA: Blackwell Sci. Pub.‎ Davies JE. 1986. Occupational asthma caused by nickel salts. J. Soc. Occup. Med. 36:29-‎‎31.‎ Dunnick JK, Elwell MR, Benson JM, Hobbs CH, Hahn FF, Haly PJ, Cheng YS, and ‎Eidson AF.‎ ‎1989. Lung toxicity after 13-week inhalation exposure to nickel oxide, nickel subsulfide, ‎or nickel sulfate hexahydrate in F344/N rats and B6C3F1 mice. Fundam. Appl. Toxicol. ‎‎12:584-‎ ‎594.‎ Glaser U, Hochrainer D, Oldiges H, and Takenaka S. 1986. Long-term inhalation studies ‎with NiO and As2O3 aerosols in Wistar rats. Int. Congr. Ser. Excerpta Med. 676: 325-328.‎ Haley PJ, Shopp GM, Benson JM, Cheng YS, Bice DE, Luster MI, Dunnick JK, and ‎Hobbs CH.‎ ‎1990. The immunotoxicity of three nickel compounds following 13-week inhalation ‎exposure in the mouse. Fundam. Appl. Toxicol. 15:476-487.‎ HSDB. 1995. Hazardous Substances Data Bank. National Library of Medicine, Bethesda, ‎MD ‎(TOMESWP:NOR version) Denver, CO: Micromedex, Inc. (Edition expires 4/30/95).‎ Hueper WC. 1958. Experimental studies in metal cancerigenesis. Arch. Pathol. 65:600-‎‎607.‎ Lundborg M, and Camner P. 1984. Lysozyme levels in rabbit lung after inhalation of ‎nickel,‎ cadmium, cobalt, and copper chlorides. Environ. Res. 34:335-342.‎ McConnell LH, Fink JN, Schlueter DP, and Schmidt MG. 1973. Asthma caused by nickel sensitivity. Ann. Int. Med. 78:888-890.‎ Muir DC, Julian J, Jadon N, Roberts R, Roos J, Chan J, Maehle W, Morgan WK. ‎Prevalence of small opacities in chest radiographs of nickel sinter plant workers. Br. J. Ind. Med. ‎‎50(5):428-‎ ‎431.‎ NTP. 1994a. National Toxicology Program. Technical Report on the Toxicology and Carcinogenesis Studies of Nickel Oxide in F344/N Rats and B6C3F1 Mice. NTP TR 451, ‎NIH Publication No. 94-3363. U.S. Department of Health and Human Services.‎ NTP. 1994b. National Toxicology Program. NTP Technical Report on the Toxicology ‎and Carcinogenesis Studies of Nickel Subsulfide in F344/N Rats and B6C3F1 Mice. NTP TR ‎‎453,‎ NIH Publication No. 94-3369. U.S. Department of Health and Human Services.‎ NTP. 1994c. National Toxicology Program. NTP Technical Report on the Toxicology ‎and Carcinogenesis Studies of Nickel Sulfate Hexahydrate in F344/N Rats and B6C3F1 Mice. ‎NTP TR 454, NIH Publication No. 94-3370. U.S. Department of Health and Human Services.‎ Novey HS, Habib M, and Wells ID. 1983. Asthma and IgE antibodies induced by ‎chromium and nickel salts. J. Allergy 72(4):407-412.‎ Ottolenghi AD, Haseman JK, Payne WW, Falk HL, and MacFarland HN. 1974. ‎Inhalation studies of nickel sulfide in pulmonary carcinogenesis of rats. J. Natl. Cancer Inst. ‎‎54(5):1165-‎ ‎1172.‎ U.S. EPA. 1985. United States Environmental Protection Agency. Health Assessment ‎Document for Nickel. EPA/600/8-83/012F, pp. 3-3.‎