United States Patent (19) 11) 4,296,104 Herschler 45) Oct. 20, 1981

(54) THERAPEUTIC DIMETHYL SULFOXIDE 3,743,727 7/973 Herschler ...... 424/181 COMPOSITION AND METHODS OF USE 4,112,946 9/1978 Herschler ...... 128/253 4,177,267 12/1979 Herschler ...... 424/238 76) Inventor: Robert J. Herschler, 3080 N.W. 8th St., Camas, Wash. 98.607 OTHER PUBLICATIONS (21) Appl. No.: 71,072 Mallach, 141, Annals New York Academy of Sciences, pp. 457-462. 22 Filed: Aug. 30, 1979 Primary Examiner-Stanley J. Friedman 51) Int. Cl...... A61K 31/10; A61K 31/17; Attorney, Agent, or Firm-Blakely, Sokoloff, Taylor & A61K 33/14 Zafman 52 U.S. C...... 424/153; 424/322; 424/337 57 ABSTRACT 58 Field of Search ...... 424/322, 153, 337 The use of urea and/or ethanol with pharmaceutical 56) References Cited compositions containing DMSO is disclosed. The result is a reduction in the undesirable side-effects normally U.S. PATENT DOCUMENTS associated with the application of DMSO compositions, 3,334,012. 8/1967 Herschler ...... 424/337 enhancement of the desired physiological effects pro 3,361,555 l/1968 Herschler ...... 71/103 duced by DMSO compositions, and other benefits. Spe 3,499,961 3/1970 Dobson et al...... 424/68 cific compositions contain DMSO with urea and/or 3,549,770 12/1970 Herschler ...... 424/337 alcohol and may also contain NaCl, KCl and/or acet 3,549,771 12/1970 Herschler ...... 424/337 amide. 3,551,554 12/1970 Herschler ...... 424/7 3,558,434 1/1971 Herschler ...... 195/81 The use of DMSO to reduce the mortality rate of fish. 3,711,602 1/1973 Herschler ...... 424/45 treated by hyperosmotic delivery is also disclosed. 3,711,606 1/1973 Herschler ...... 424/45 3,740,420 1/1973 Herschler ...... 424/45 8 Claims, No Drawings 4,296,104 1 2 blood cell lysis which results from intravenous DMSO THERAPEUTC DIMETHYL SULFOXIDE administration can be injurious or even fatal to the sub COMPOSITION AND METHODS OF USE ject. Furthermore, it is generally advisable to minimize the BACKGROUND OF THE INVENTION 5 dosage of any pharmaceutical substance administered to The present invention relates to pharmaceutical com a human or other animal subject to the smallest effective positions containing DMSO for adminstration to human amount. Although DMSO is one of the most penetrat or other animal subjects. More specifically, it relates to ing of pharmaceutical substances and is known to be new DMSO formulations containing substances which effective in minute doses, it would be desirable to fur enhance the effectiveness of DMSO, reduce undesirable 10 ther reduce the minimum dosage of DMSO needed to side-effects sometimes created by the use of DMSO and achieve a desired physiological effect. make DMSO compositions more appealing to users. A related problem is observed when fish are treated Dimethyl sulfoxide (DMSO) is a versatile substance with hyperosmotic concentrations of a membrane per that has numerous pharmaceutical and nonpharmaceuti meability altering agent as discussed in U.S. Pat. No. cal uses. It is widely used throughout the world for 15 treating humans and other animal subjects. 4,112,946, incorporated herein by reference. Such As Described in U.S. Pat. No. 3,549,770, No. agents include urea, NaCl and acetamide. 3,740,420, and No. 3,790,682, incorporated herein by These substances, with the possible exception of acet the reference, DMSO is an active agent in relieving the amide, are substantially nontoxic when applied exter signs and symptoms of numerous body disorders, in 20 nally to mammalian subjects. But, when incorporated in cluding accelerating the healing of certain injured body a solution in contact with an epithelial membrane of tissues and in relieving the signs and symptoms of anxi fish, such substances can be lethal at moderate concen ety. w trations (as low as about 3 weight percent). U.S. Pat. No. 3,551,554, No. 3,711,606 and No. To optimize delivery of therapeutic agents to fish, 3,743,727, incorporated herein by reference, describe 25 solutions containing greater than 3 weight percent of how DMSO is effective to enhance tissue penetration of the above listed solutes are required. Fish mortality is other substances, especially other physiologically active thus a substantial barrier to the effective use of hyperos agents. DMSO can thus be added to a variety of phar motic treatments. maceutical compositions to accelerate assimilation into SUMMARY OF THE INVENTION body tissue. In some instances this means that smaller 30 doses can be administered when DMSO is used. Specific DMSO compositions and methods of appli Yet, despite their many benefits, DMSO composi cation have now been discovered. Use of such composi tions are sometimes passed over in favor of other phar tions expands the acceptability of DMSO by eliminating maceutical compositions even in instances where or reducing undesirable side-effects. And, the new DMSO would be the most effective pharmaceutical 35 DMSO compositions and methods of use are observed agent. This is because many subjects suffer from one or to provide new therapeutic effects and beneficial uses. more side-effects when treated with DMSO. In some More specifically, it is found that when DMSO and cases, the side-effects are so pronounced that subject or urea are both administered to epithelial regions of a physician will forego the use of DMSO in favor of a less human or other animal subject, expected adverse skin effective therapeutic agent. 40 reactions, malodorous breath and foul taste are substan A variety of undesirable side-effects have been ob tially reduced. In most cases they are entirely elimi served to result from administration of DMSO. The nated. most frequently occurring are adverse skin reactions, Furthermore, urea is discovered to potentiate DMSO malodorous breath and foul taste. in certain instances. It appears that a DMSO composi The adverse reactions caused by DMSO are well 45 documented. At page 356 of the standard reference tion containing urea and/or ethanol is more rapidly Contact Dermatitis by Alexander A. Fisher, M.D. (2nd absorbed into tissue than are similar compositions con Ed., 1973), dimethyl sulfoxide (DMSO) is listed as a taining no urea or ethanol. If the DMSO composition primary urticariogen. Volume 141 of Annals New York includes another pharmaceutical substance, such other Academy of Sciences includes several articles describ 50 substance is also absorbed more rapidly when urea and ing the undesirable side-effects attributed to DMSO. /or ethanol are present. The presence of NaCl, KCl These include articles by Goldman, et al. at pages 429, and/or acetamide reduces discomfort resulting from 433-35; Sulzberger, et al. at pages 439-40; Brown at topical application of DMSO compositions and appears pages 500-501; and several others. to further enhance penetration. The magnitude of the malodorous breath problem is 55 It has also been discovered that certain DMSO for so large that, in some instances, hospitals have had to mulations, which contain urea, are effective in treating isolate wards where DMSO is administered from the diseased finger and toe nails, for softening cuticle to be central air conditioning system. Skin irritations from removed from finger and toe nails, and for softening topically applied DMSO have been so great that a sub epidermal thickenings to ease removal. stantial number of patients refuse treatment. When DMSO is administered intravenously with Another, potentially more serious side-effect is some ethanol, red blood cell lysis is reduced and continues to times observed when DMSO is administered intrave decline with repeated administrations. nously. This is red blood cell lysis, Intravenous adminis When DMSO is added to solutions containing urea, tration of DMSO is crucial if the substance is to be used NaCl, and/or acetamide, such solutions are less toxic to for treating the brain and spinal cord, for cancer ther 65 fish than similar solutions without DMSO. apy or to treat organ hypoxia, heart attack and other It is therefore an object of this invention to provide internal conditions. Large intravenous doses of DMSO pharmaceutical compositions and methods to allow the can have a therapeutic effect for such purposes but red application of DMSO to a human or other animal sub 4,296,104 3 4. ject without creating adverse skin reactions, malodor ceutical agents so that, at a given weight percent of ous breath or foul taste. solute, compositions containing urea in combination A further object is to provide compositions and meth with DMSO, have an enhanced penetrating ability. The ods of application whereby the lysis of red blood cells, effect is even greater if the amount of solute is increased resulting from intravenous administration of DMSO, is by addition of urea to the DMSO composition. reduced. It appears that the only suitable substances for en A further object is to provide compositions and meth hancing the penetration of DMSO compositions are ods of treatments for diseases of the fingers and/or toe substances having a weight per volume percentage nails. concentration at isotonicity less than that of DMSO, but Another object is to provide formulations and meth 10 only a few of such substances produce the desired in ods of application to enhance penetration of DMSO and crease in penetration. DMSO based pharmaceutical compositions into animal Further criteria of substances which enhance DMSO tissue. penetration include water solubility at ambient tempera An additional object is to provide safe and effective tures and a molecular weight no greater than 78. Sub skin softening agents which can be used to soften cuticle 15 stances which tend to polymerize are unsuitable. If the for removal from finger and/or toe nails and for soften substance is to be used with DMSO-urea compositions ing epidermal thickenings to facilitate removal. applied to living animals, it must also be relatively non Yet a further object is to provide means for protect toxic to the intended animal subject. ing fish from the toxicity of solutions used for hyperos Specific substances of value in promoting penetration motic treatment of fish. are found to include urea, NaCl, KC, ethanol and acet These and other objects, advantages and features of amide. Of these substances urea is the most effective. It the present invention will be apparent from the follow appears to act by "opening' membranes to allow in ing detailed description. creased penetration. DESCRIPTION OF PREFERRED 25 The fact that urea, NaCl, KCI, acetamide and/or EMBODIMENTS ethanol may be combined to enhance DMSO penetra The side-effects which have long hindered the use of tion is important in several respects. Although urea and DMSO as a therapeutic agent are quite surprisingly ethanol reduce or eliminate the side-effects created by eliminated when urea is administered with formulations the administration of DMSO, some subjects will con tinue to have at least mild reactions to the administra containing DMSO. Even small amounts of urea are 30 beneficial in reducing histamine release, burning and tion of DMSO. By administering urea, NaCl, KCl, acet itching, localized dermatitis, drying, cracking and blis amide and/or ethanol, it should be possible to reduce tering of skin, upleasant breath odor, after-taste, head the amount of DMSO administered as an active thera ache and nausea in subjects receiving dermally adminis peutic agent since greater tissue penetration of the ap tered DMSO compositions. 35 plied DMSO can be obtained. The ability to reduce the Urea is an especially excellent inhibitor of the side-ef amount of DMSO administered, without reducing the fects, because it is a naturally occurring substance in amount which is absorbed, should further diminish ad human and other animal subjects. Urea is substantially verse side-effects of the DMSO. non-toxic and animal subjects are quite tolerant to its A similar benefit results when urea, NaCl, KCl, acet presence. It thus can be used with almost any pharma 40 amide and/or ethanol are added to compositions which ceutical composition containing DMSO, without fear of contain DMSO as a carrier and penetration enhancer toxic effects. for other pharmaceutical substances such as antineo The reasons why urea is effective from reducing and plastic agents, analgesics, anti-inflammatory agents, eliminating the above listed undesired side-effects is not anticoagulants, vasodilators, anti-microbial agents, ul fully understood. It appears, however, that the urea 45 tra-violet screening agents, diagnostic dyes, diagnostic inhibits or prevents the production of undesirable radiopaque agents, dietary supplements, nutrients, phys DMSO metabolites. iologically active steroids and protein modifying agents. In the case of the breath odor problem, it is known Protein modifying agents include those substances that dimethyl sulfide is a minor metabolite of DMSO which modify collagen and, possibly, other substances and that it is expired through the lungs causing mal 50 found in connective tissue such as hyaluronic acid, elas odorous breath and foul taste. Urea possibly prevents tin, and fibrinogen. One such protein modifying agent is the breakdown of DMSO to dimethyl sulfide. Support methylsulfonylmethane (MSM) as described in my si ing laboratory experiments show that DMSO will de multaneously filed U.S. patent application entitled compose to dimethylsulfide when heated in a test tube. Preparations Containing Methylsulfonylmethane and But, when urea is added and the experiment repeated, 55 Methods of Use and Purification. no dimethyl sulfide is detected. DMSO is a known penetration enhancer for chemical Similarly, the adverse skin reaction, such as histamine agents having a molecular weight less than about 8,000, wheal and flare may be the result of an attack by the administered to intact body membranes. It is also found dimethyl sulfide molecule against mast cells in the sub to enhance the penetration of higher molecular weight cutaneous layer of skin to which DMSO is applied 60 substances, such as enzymes, that are administered to topically. Urea's blocking of DMSO breakdown to stressed membranes including membranes that are in dimethyl sulfide would thus account for the observed flamed, are scarified or have been subjected to severe antiurticariogenic effectiveness in reducing adverse skin osmotic stress. In both situations, the inclusion of urea, reactions caused by DMSO. NaCl, KCl, ethanol and/or acetamide can further assist The enhanced penetration of DMSO solutions con 65 the penetration of DMSO and the pharmaceutical taining urea is possibly related to the fact that urea has agent. Increased penetration can improve pharmaceuti a weight per volume percentage concentration at isoto cal effectiveness and, in some instances, make it possible nicity less than that of DMSO and most other pharma to reduce the dosage administered. 4,296,104 5 6 Penetration enhancing substances, such as urea, are DMSO-urea compositions are also well suited for use particularly useful in DMSO solutions containing diag with physical therapy techniques, particularly the use of nostic dyes. In the staining of cells for diagnostic exami energy such as ultra sound, in treating musculoskeletal nation, it is desirable to use the least possible amount of disorders. dye and other foreign chemicals to minimize alterations 5 As illustrated by several examples below, DMSO in the cells being dyed. the enhanced cell penetration urea compositions soften and moisturize the skin of that results from the addition of urea, makes it possible subjects receiving dermal applications. Certain DMSO to reduce the amount of dye and DMSO in staining urea compositions accordingly make excellent cosmetic compositions. The urea added has no adverse effect on skin softening lotions or gels. Also DMSO-urea compo the structure of cells to be dyed. 10 sitions are excellent as vehicles for other skin treating An additional benefit discovered is that human sub cosmetic agents. When the phrase "pharmaceutical jects treated with the previously described composi compositions" is used herein, it thus includes cosmetic tions preferred those compositions, containing a salt, preparations. most notably NaCl, over similar compositions without Urea is known to have some beneficial effect on skin, salt. Subjects interviewed indicated that topically ap 15 but in most standard cosmetic compositions it "washes plied DMSO compositions containing salt are more off. When DMSO and urea are used together, skin comfortable. permeation of each is enhanced so the skin-softening Urea can be administered with DMSO to produce the benefits of urea are increased and sustained even after same physiological effects attributed to DMSO compo washings. sitions administered without urea. For example, urea 20 As will be described below, the use of ethanol with can be administered with DMSO to an area of tissue DMSO in intravenous administrations proved to be inflammation in an amount effective for relieving signs independently effective in reducing malodorous breath and symptoms of inflammation, to a subject suffering and red blood cell lysis. from pain in an amount effective to relieve pain, to a Formulations subject suffering from abnormal muscle contractions in 25 an amount effective to promote muscle relaxation, to a As with any multi-purpose pharmaceutical composi subject suffering from symptoms of vascular insuffi tion, some experimentation is necessary to determine ciency in the blood and lymph circulatory system in an the optimum dosage of DMSO and urea to be applied amount effective to relieve symptoms of vascular insuf for a particular purpose. For example, when it is a goal ficiency. 30 to reduce a side-effect produced by the application of DMSO, the amount of urea used should be an amount Treatment with effective amounts of DMSO and urea effective to obtain the desired reduction. Likewise, if can also relieve signs and symptoms of a burn, can pro the goal is to enhance penetration, the amount of urea or mote healing of a skin graft area following a transplant, other penetration enhancing substance should be an and can relieve signs and symptoms of respiratory dis 35 amount sufficient to enhance penetration. tress. When DMSO is given with urea to subjects hav As described in the prior patents listed above, DMSO ing joints with arthritic signs and symptoms in an compositions for topical application should contain at amount effective to relieve signs and symptoms of ar least 10 weight percent DMSO to have any beneficial thritis, to subjects suffering from tissue damage in an effect. Compositions for clinical use should have at least amount effective to promote the repair of tissue dam about 40 weight percent DMSO; and for greatest effect, age, or to mammalian subjects suffering from signs and a composition should contain at least about 50 weight symptoms of anxiety in an amount effective to relieve percent DMSO. To be effective in reducing DMSO signs and symptoms of anxiety, improvement in sub induced side-effects and/or to enhance the penetration jects' conditions are observed. of DMSO compositions, urea should be present in a It has also been found that certain compositions, con 45 weight ratio to DMSO of greater than 1:99. Most signif. taining both DMSO and urea, have medical benefits not icant results are achieved when the weight ratio of urea produced by compositions containing DMSO or urea to DMSO is greater than 1:9. Normally, DMSO-urea alone. As one example, it is found that DMSO, adminis compositions should contain no more than about 60 tered with urea, will repair or remove abnormal, dead, weight percent urea since larger concentrations could or diseased tissue. DMSO-urea compositions can thus 50 only be obtained at the cost of reduced effectiveness be used to treat interstitial cystitis or connective tissue due to diminished DMSO amounts. diseases such as progressive systemic sclerosis. In special circumstances the desired DMSO concen DMSO-urea, compositions can also be administered tration for a pharmaceutical composition might be sub to benefit diseased finger and/or toe nails of human or stantially below 10 weight percent or above 50 weight other animal subjects. Administering DMSO and urea 55 percent. The above urea ratios will still apply in such to the diseased portion of a nail will soften the diseased circumstances. For example, if a low concentration (3 portion. After several days of treatment, the diseased to 4 weight. percent) DMSO composition is prepared portion of the nail can be removed painlessly by gentle for application to the eye, a suitable urea amount would urging using forceps. In most instances, the undiseased be one weight percent. portion of the nail is not adversely affected by treatment Compositions containing one weight percent or less with a DMSO-urea composition. of DMSO are effective for treating membranes that When used to treat diseased or damaged tissue, have been stressed, e.g. membranes that are inflamed, DMSO-urea compositions are most effective when ap are scarified or have been submitted to severe osmotic plied at a temperature above 37 C., preferably as hot as stress. In such instances, the abovestated DMSO-urea the subject will tolerate. Warm applications are espe 65 ratios would still apply. cially beneficial when treating musculoskeletal disor Pharmaceutical compositions for treating the skin, ders such as arthritis, sprains, strains, soft tissue injury oral cavity and rectal epithelium may contain DMSO and the like. and urea as their sole components. In any such composi 4,296,104 7 8 tion, the weight ratio of DMSO to urea should not be forms an overwrap. Heat can be applied to the exterior less than about 100:35. Even at this ratio, the solution is of the bag to accelerate treatment. best applied while heated to prevent precipitation of the Similary, spinal injuries can be treated by saturating a ea. fabric with a DMSO-urea solution and then spreading When NaCl, KCl, ethanol and/or acetamide are pres the fabric along the spinal column. The fabric may be ent in DMSO-urea compositions, to effectively increase covered with a nonporous plastic sheet and hot water penetration or comfort, they should be present in a bottles applied to speed penetration of the DMSO-urea certain minimum amount. The combined weight of solution. these agents, in ratio to the weight of urea present in the In most instances, it is preferred that DMSO and urea composition, should be greater than 1:99. Significant 10 be combined in a common composition for administra benefits are observed when the ratio is greater than 1:9. tion together or otherwise be administered concur The most effective formulations, at least for topical rently. In the specific case of topically applied DMSO application, includes about six grams of urea, one gram composition, some reduction in side-effects is also ob of NaCl and at least six grams of water for every nine served if patients are treated with DMSO without urea teen grams of DMSO. An aqueous medium is usually 15 after a preceeding treatment at the same site, with both required for any DMSO-urea composition containing DMSO and urea. For topical administrations at least, it NaCl or KC. is thus possible to use treatment regimens such as alter When included in DMSO compositions, urea, NaCl, nating applications of DMSO compositions with and and KCl may complex with DMSO molecules. As a without urea. result, such DMSO compositions may include DMSO 20 If a DMSO composition, for topical application, in urea complexes, DMSO-salt complexes and/or DMSO cludes substances which would react adversely or be urea-salt complexes of various types. The exact effect deactivated when combined with urea or would react that complexing has on the activity of the compositions with urea to form macromolecules which would retard containing DMSO and urea is not known. It is believed, tissue penetration, a urea composition can first be ap however, that compositions containing the complexes 25 plied to the treatment site and allowed to dry. The listed above are effective in obtaining desirable reduc DMSO composition could thereafter be applied at the tions in side-effects and penetration increases same site with less chance that the urea-sensitive sub When DMSO is used in the same composition with stances would be adversely affected. As desired NaCl, urea for the removal of cuticle, for treating diseased 30 KCl, ethanol and/or acetamide can be included in the finger and toe nails, or for the remover of epidermal urea composition or, if compatible, in the DMSO com thickenings the composition should include a hydro position. phobic ointment base and at least 10 weight percent As a penetration enhancing agent, urea can be admin each of DMSO and urea to be effective. It is also advan istered with DMSO by topical administration, intrave tageous to apply a salt such as NaCl or Na2S. Use of 35 nous administration, subepidermal injection or oral in such a salt increases the rate at which the nail or thick gestion. It can also be administered intrathecally, in ened skin are softened for removal. Specifically it ap travesically, rectally, or by instillation into eye, ear, pears that DMSO-urea compositions containing NaCl nose or abnormal sinuses of the body. and/or Na2S penetrate more deeply into the area be The following examples describe a few of many clini tween the nail and plate than do compositions without cal tests which show that the presence of urea can bene such a salt component. fit pharmaceutical compositions containing DMSO. In Such solutions for treating diseased nails should in each of the listed examples, DMSO and urea were ap clude between 0.25 and 10 weight percent of a salt plied in a common composition. selected from NaCl, Na2S and mixtures thereof, along As previously mentioned, alternating applications of with effective amounts of urea and DMSO. Less than 5 45 a DMSO composition and a urea composition may be weight percent salt is found to be fully adequate in most advisable under some circumstances. For example, in instances. the unusual circumstance that a preferrred DMSO com To facilitate topical applications, any of the above position contains a chemical agent that is unstable in the compositions may include a pharmaceutically accept presence of urea, it would be preferable to alternate able thickening agent to increase the viscosity of a com 50 applications of the DMSO composition and a urea com position. Such thickeners may be used to form creams, position or to mix the two compositions immediately lotions, gels, pastes, ointments and suppositories. before administration. Sequential applications of DMSO and urea compositions or mixing immediately Methods of Application before application might also be preferred if the sepa Urea may be administered with DMSO compositions 55 rate DMSO and urea compositions have a substantially by any route previously known for DMSO administra longer shelf life than some combined formulation. tion. The most dramatic reductions in side-effects are Whether to combine urea with a particular DMSO observed when urea is added to DMSO compositions composition is best determined by experimentation. for topical application. Subjects using topically applied DMSO suffer from substantially less malodorous 60 1. Cutaneous Administrations breath, foul taste, and adverse skin reaction when urea a dermal administrations is used along with the DMSO. Topical applications of DMSO and urea may be by EXAMPLE 1. any standard technique. They may be painted or spread on and allowed to dry or applied with saturated pads. 65 Wt. Percent One advantageous method for treating limbs or digits DMSO 61.7 is to place an appropriate DMSO-urea liquid composi water 26.5 tion in a plastic bag and insert the limb so that the bag ea 8.8 4,296,104 10 -continued W. Percent Wt. Purce NatC 3.0 DMSO 47.6 Wt: 33.3 A control solution containing 62 wt. percent DMSO in NaCl 4.8 water was also prepared. Both formulations were der UC 4.3 mally applied to each of five human subjects known to be sensitive to compositions containing DMSO at a Topical applications of this composition provided concentration of 50 wt. percent. 10 excellent results. Irritations and itching of the skin In a first test, the two solutions were topically applied stopped promptly and were followed by rapid healing concurrently but at different sites on skin of the sub in all cases. Minor cuts were also treated with the above jects. Each subject complained of itching and burning at solution; and infection free healing resulted in each case. sites where the control solution was administered. And 15 EXAMPLE 4 each had bad breath odor. At sites where the solution DMSO compositions have been used successfully to containing urea was applied, there was no discomfort or treat a variety of athletic injuries such as sprains, muscle adverse skin reaction, even when the solution was ap cramps, and the like. In this example, the following plied to particularly sensitive areas of the neck, below formulation was used: the chin. 20 Several days later, the same subjects were treated with only the solution containing urea. There was no DMSO 200 g. discomfort or adverse skin reaction. Neither bad breath urea 100 g. ethanol (absolute) 50 g. nor foul taste were observed. Each of the subjects vol Water 25g. unteered that their skin was softened when the composi 25 methyl salicylate tions containing urea was applied. (as odorant) 10 g. After several more days, the subjects were again 3% Carbopol 934 treated with the control solution. The subjects experi (carbomer-934) 80 g. enced a return of malodorous breath, foul taste, and triethanolamine 2 g. adverse skin reactions. 30 Topical application of this composition has proved EXAMPLE 2 effective in test treatments for sprains, muscle cramps A test was conducted to determine whether it would and other discomforts. Athletes receiving the above be advantageous to use DMSO compositions containing composition had no abnormal irritations. relatively large amounts of pharmaceutically acceptable 35 One subject having very fair complexion and red hair agents having a weight per volume percentage concen was unable to tolerate a 50 wt. percent aqueous DMSO tration at isotonicity less than that of DMSO. Urea is composition without urea. When a gel containing the such an agent. So, for comparison purposes, the compo above listed ingredients was applied to the skin of the sition of Example 1 was tested against the following subject, good medical relief was observed. Further composition which contains more urea and less water: 40 more, there was no indication of the undesirable side-ef fects previously experienced from the application of DMSO without urea. Wt. Percent DMSO 6.7 EXAMPLE.5 Water 7.6 45 La 17.6 A methyl salicylate ointment was prepared for use for NaC 3.0 treating human subjects suffering from whiplash. The ointment included urea and ethanol to enhance penetra Human subjects, suffering from a variety of ailments tion of methyl salicylate and DMSO. Specifically, it treatable with DMSO compositions, received each of 50 included: the two formulations at separate times during the course of treatment. In every instance, the subject expressed a DMSO 200 g. preference for the reformulated composition containing C 100 g. water 100 g. a greater amount of urea. 55 ethanol 50 g. Specific benefits observed include prompter medici methyl salicylate 10 g. nal response and increased comfort (less tissue irrita Carbopol 940 tion). Effected skin area, of subjects suffering from (carbomer-940) 2.4 g. scleroderma were softened when the high urea comcen triethanolamine 2 g. tration composition was applied topically. Such sub O jects receive no substantial relief when urea composi This ointment was applied to the necks of whiplash tions without DMSO are applied. victims who previously had experienced discomfort upon treatment with other DMSO substances. Methyl EXAMPLE 3 salicylate in the ointment penetrated extremely rapidly The following composition was used to treat subjects 65 such that the subjects reported immediate relief from suffering from contact dermatitis, from poison oak, pain. The ointment was soothing when applied topically insect bites and other conditions characterized by subd to low sensitivity skin areas and no malodorous breath ermal histamine release and consequent discomfort: or foul taste resulted. 4,296,104 1. 12 and treatment was stopped except for occasional appli EXAMPLE 6 cations to relieve discomfort. The paste of Example 13 was used to impregnate polyolefin foam pads. The impregnated pads were ap EXAMPLE 10 plied to unpared calluses and corns on the foot of a DMSO and urea may be used effectively together, subject. After daily treatments for four to eight days, even at very low concentrations, when applied to ani the epidermal thickenings were sufficiently softened for mal membranes which have been stressed. easy removal. In one laboratory experiment hair was removed from both flanks of rabbits of about three kilogram body EXAMPLE 7 10 weight. The flanks were then blade shaved to fully Persons with more severe epidermal thickenings expose the epidermis. Under anesthesia, the both flanks were treated according to the procedure of Example 13 of five rabbits were severely scarified using a scalpel to using the composition of Example 6. Good success in expose subcutaneous tissues. The right flank of each rabbit was kept continuously the softening of tissue for removal was achieved. 15 moistened with a 0.75 weight percent aqueous solution EXAMPLE 8 of DMSO containing 1.0 mg. of superoxide dismutase The DMSO-urea composition of Example 1 was ad enzyme (m.w.. 30,000-40,000) per 10 ml. of solution. The ministered to patients suffering from a hardening of the left flanks were treated identically except that the solu skin as a result of incurable, progressive systemic sclero tion applied was free of DMSO. 20 It was observed that healing was greatly accelerated sis. Previously, these patients had each been treated for on the right flanks as compared to the left. Specifically, their condition with solutions of DMSO and water strong tension resistant healing of the right flanks oc without urea, over a period of years. curred in about half the time. After treatment with the DMSO-urea composition of The addition of urea to such low concentrated Example 1, each of the patients reported a strong pref. 25 DMSO compositions can further improve healing rate, erence for that composition. Specifically, they reported enhance enzyme penetration and eliminate side effects relief from bad breath and a soothing effect upon appli resulting from DMSO application. When applied to cation. Most significantly, skin of the subjects was softer stressed tissue, not more than about 1.0 weight percent and more flexible after several weeks of administration. urea is required. For use in preparing the DMSO formu Also, blood circulation of the skin was improved. lation of this example, it would be sufficient for the Systemic sclerosis typically impairs blood circulation to aqueous solution to contain about 0.75 weight percent the skin as evidenced by a minimum of color change when finger pressure is applied to the effected area. Tea. After the patients were treated with the DMSO-urea EXAMPLE 11 composition of Example 1, once daily over the entire 35 Compositions containing methylsulfonylmetane body for several weeks, vascular blanching of the skin (MSM) and urea improve the softness and pliability of was observed when finger pressure was applied. Blush skin even of persons suffering from adverse skin condi ing occurred upon release. tions. In one test, two human subjects suffering from EXAMPLE 9 "hide bound disease' or progressive systemic sclerosis 40 were treated with a solution containing 20 weight per The vascular effect of DMSO compositions contain cent MSM, 20 weight percent urea, 30 weight percent ing urea was observed in the dramatic recovery of one dimethyl sulfoxide and 30 weight percent water. subject with ischemic ulceration of a finger. At the The subjects were treated by placing 15 milliliters of beginning of treatment, the finger was cyanotic, ulcer the solution in a plastic bag, placing a hand or foot to be ation was progressing, and surgical amputation had 45 treated in the bag. The hand or foot with plastic bag been recommended. overwrap was then immersed in a heated water bath In an attempt to improve this condition, the finger maintained at a temperature as warm as the subject was treated by dipping in an aqueous DMSO solution. would tolerate, taking care not to dilute the solution. But the treatments were soon abandoned because the The hands and feet were thus immersed for 30 min pain of treatment was too great. utes, three times daily, for a period of two weeks. The In a second attempt at treatment, the finger was result was a reduction in discomfort and increased skin dipped in the following preparation over a period of softness and pliancy. several weeks: Dimethylsulfoxide and urea in the solution enhanced penetration of MSM into the effected tissue. No adverse 55 side-effects resulted from administration of the DMSO. Wt. Percent DMSO 50 EXAMPLE 12 Water 35 lea 10 DMSO is a useful substance in many dermally applied NaCl 5 cosmetic preparations. But, cosmetics containing 60 DMSO are yet to be widely marketed because many test users experience malodorous breath and skin irrita Twenty-five to fifty milligrams of indomethacin were tion. Such cosmetic preparations include the long-last added to each 500 grams of this solution to serve as an ing antiperspirants described in U.S. Pat. No. 3,499,961. analgesic and anti-inflammatory agent. To test whether such antiperspirants would be im After about 12 weeks of treatment, there was appar 65 proved by the addition of urea, a comparison test was ently full recovery of the finger with ulcer healing and conducted. excellent general appearance of the entire finger. At the A first paste was prepared containing, by weight, 10 end of the period amputation was counter-indicated; parts of a 6:1 DMSO-aluminum chloride complex, 10 4,296,104 13 14 parts urea, 10 parts ethanol, 2 parts NaCl. Sufficient although cuticle of both fingernails was removable by colloidal silica (Cab-O-Sil) was added to the other in gentle rubbing. gredients so that a proper paste consistency was achieved. EXAMPLE 16 When applied to the left axilla of human subjects no Another paste was formed having the composition: breath odor or skin irritation resulted. A second, similar paste was formulated without urea. This applied one hour later to the right axilla of the Wt. Percent same subjects. The subjects experienced burning and DMSO 45 10 rea 45 itching of the right axilla within 15 minutes after app sodium sulfide 5 plication of the second paste. Malodorous breath was lanolin 5 detected 30-35 minutes after application of the second paste. This paste was prepared by heating to 60° C. amd 15 milling to a smooth, uniform paste. This paste was ap EXAMPLE 13 plied to subjects having defective fingernails in the An anti-mosquito lotion was prepared by combining manner described in Example 15. the following ingredients, with heating, to form a solu The paste of this example softened the defective nail tion: and debrided the underlying nail plate more rapidly 20 than did the paste of Example 15. A slight etching of the DMSO 30 g. normal nail surface also occurred. ethanol 30 g. 1822 * 15 g. EXAMPLE 17 N,N-diethyl-toluamide 10 g. The following substances were mixed to form a gel: 25 This preparation was applied to the skin of three DMSO 20g. human subjects, known to be sensitive to DMSO com tea 40 g. positions. None of the subjects had adverse skin reac ethanol 18 g. tions, urticaria or bad breath odor. Water 9 g. 30 isopropyl palmitate 8 g. b. nail administrations Carbopol 940 (carbomer-940) 2 g. EXAMPLE 14 di(2-ethylhexyl)amine 2 g. A paste was prepared containing: griseofulvin 1 g. 35 Various subjects having defective fingernails with Wt. Percent undergrowing fungal infection were treated using this DMSO 50 gel composition. Gauze or polyolefin foam pads were urea 40 impregnated with the gel, applied to the defective fin lanolin 10 gernails and covered by an occlusive overwrap. After 40 10 days, the overwrap and pads were removed. The By heating the mixture to 60 and milled until a smooth, diseased nails were then easily separated from the plate uniform paste was formed. by forcep urging. After removal of the nail, a single This paste was applied to a trauma injured toenail and painting of the preparation on the plate achieved both underlying plate. After application, the toe was covered debridement of diseased and dead tissue and control of by a protective overwrap. At the seventh day after 45 the infection. application, the nail was easily removed with gentle forcep urging EXAMPLE 1.8 In a related test, the composition of Example 16 was heated and then painted on diseased nails of human EXAMPLE 15 50 subjects. Twenty to thirty minutes after application, the Another paste was formed according to the composi nails were treated with an appropriate pharmaceutical tion: composition, e.g. one containing an antimicrobial agent. After several days of treatment, the diseased nail portions were removable by forcep urging and infec Wt. Percent 55 DMSO 45 tions were under control. Tea 45 EXAMPLE 19 NaCl 5 lanolin 5 Two liquid preparations were prepared for compari son purposes. These compositions were as follows: A mixture of these ingredients was heated to 60° C. and milled until a smooth, uniform paste was formed. Preparation 1: Wt. Percent This paste was applied to a chronically defective Tea 50 fingernail of a human subject and also to a healthy nail Water 25 on an adjacent finger. Both the fingers were covered by 65 DMSO 25 a protective overwrap. By the fifth day after applica tion, the defective nail was easily removed by gentle The ingredients were mixed, heated to 50° C. and urging with forceps. The normal nail was uneffected thereafter milled to a uniform dispersion. 4,296,104 15 16 equally effective in reducing the signs and symptoms of prostatitis. Preparation 2: Wt. Percent 2 50 EXAMPLE 22 Water 50 The composition of Example 21 was administered to human subjects who suffered from interstitial cystitis, The ingredients were mixed, heated to 50° C. and and who previously showed little improvement when thereafter stirred with cooling to form a uniform solu treated with a 50 wt. percent solution of DMSO in tion/dispersion. water. In each case, thiry to one hundred milliliters of Each of the preparations was used to impregnate O the composition was administered daily through a cath cellulose base foam sheets. The sheets, in turn, were eter inserted to the bladder. used to treat subjects having two adjacent digits with As compared to treatment with the 50 wt. percent defective toe or fingernails. One digit of each subject DMSO solution, the DMSO-urea formulation produced was wrapped with a foam sheet containing Preparation less patient discomfort. Also, improved therapeutic 1 and the adjacent digit wrapped in a foam sheet con 5 results were observed. Specifically, frequency of urina taining Preparation 2. It was observed that Preparation tion, pain and discomfort with bladder palpitation were 1 was about one-third more effective in softening defec reduced. Visual observation with a a cystoscope and tive, diseased nails for removal. biopsies of bladder tissue confirmed that there was a reduction in inflammatory lesions. After changing from EXAMPLE 2.0 20 treatment with aqueous DMSO to treatment with the A nail conditioner was formulated from the following DMSO-urea formulation, subjects' bad breath was sig ingredients: nificantly reduced. EXAMPLE 23 25 Wt. Percent For some time, DMSO has been used in concentra Water 50 tions of up to 100 percent for treating hypersensitive dimethylsulfoxide 20 methylsulfonylmethane (MSM) 10 teeth recovering from extensive restorative oral sur lea O gery. Substantial pain and discomfort as well as trauma glycerine 5 30 and localized tissue injury to the teeth and guns fre glyoxal (30% aqueous) 5 quently results from such surgical procedures. DMSO is known to promote general tissue repair and reduce The formulation was applied with cotton pads to pain so that after about 2 weeks a patient can comfort healthy nails of human subjects and allowed to remain ably chew solid food again. for at least 15 minutes. At the end of that time, the nails 35 A young female subject recovering from oral surgery were toughened, i.e. less brittle, and the cuticle was was treated with the following composition: softened such that it could be removed by gentle rub bing. It is not fully understood how the brittleness of the Wt. Percent 40 DMSO 7.4 nail is reduced by application of the formulation. It t 23.8 appears, however, that the MSM is the principal active water 4.8 agent. The DMSO and urea appear to accelerate pene tration of the MSM into the tough nail material. Subjects using this formulation has no reddening or Cotton pledgets were saturated in the solution and ap 45 plied to the gums. After two days of treatment both the irritation of the skin surrounding the nail and did not relief of pain and the extent of healing were greater than suffer from malodorous breath. observed for typical subjects receiving two weeks of 2. Mucosal and Urogenital Administrations treatment with high concentrations of DMSO solutions without urea. EXAMPLE 21 Oral surgery patients treated with DMSO typically The composition of Example 9 was slightly reformu experience a sulfurous breath and bad aftertaste which lated to include: can last for a full day after treatment. These adverse side-effects were greatly reduced in the subject treated Wt. Pert with the above composition containing urea. DMSO SO EXAMPLE 24 wit 35 12 Another composition suitable for treating dental pa NC 3. tients includes: 60 This formulation was administered to patients suffer W. Prett ing from urological problems such as prostatitis, OMSO 70 through a catheter extending to the neck of the bladder. 20 Clinical tests showed that patients receiving the watt 8 DMSO-urea composition experienced significantly less 65 NC discomfort and less malodorous breath than when a 50 wt. percent solution of DMSO in water was adminis When applied in the manner described in Example 23. tered without urea. The DMSO-urea composition was this composition produces superior results in reducing 4,296,104 17 18 dental pain and swelling associated with procedural of red cell lysis was great and there was a pronounced tralia. evolution of dimethyl sulfide. The test was repeated with the DMSO solution re EXAMPLE:25 placed by a 50 volume percent aqueous solution of Elderly persons and subjects having connective tissue 5 ethanol. After the solution was combined with the diseases, frequently have a problem with drying, painful blood, no odor was evolved but there was some red gums. To treat this condition, the following composi blood cell lysis. tion was prepared: In a third test, the procedure was repeated again, only the additive solution contained 66.6 volume percent DMSO 200 g. 10 DMSO and 33.3 volume percent ethanol. After one lea 50 g. volume of solution was added to nine volumes of blood, Water 50 g. neither red blood cell lysis nor dimethyl sulfide odor 3% Carbopol 934 (carbomer-934) 10 g. was observed. A sample of the atmosphere over the triethanolamine 0.4g. blood was collected and analyzed with a gas chromato 15 graph. There was no detectable dimethyl sulfide peak. The Carbopol 934, a polymer of acrylic acid manu EXAMPLE 27 factured by B. F. Goodrich Chemical Co. of Cleveland, Dog blood was drawn and heparinized to prevent Ohio, in combination with triethanolamine as a neutral clotting. After tubing, samples of the blood were mixed izer, caused the composition to be a soft gel. This was 20 with a 20 wt. percent aqueous dimethyl sulfoxide solu flavored with small amounts of spirits of peppermint to tion in such an amount that the DMSO-blood mixture mask the somewhat bitter taste of DMSO. contained 12 grams of DMSO to every 100g of blood. When applied topically to the gums of subjects, pain Upon mixing, there was almost instant red blood cell was reduced and gum tissue was softened. lysis with a predominant odor of dimethyl sulfide. The 25 presence of dimethyl sulfide was confirmed by gas 3. Intravenous Administrations chromatography. In current practice, DMSO is administered intrave In a separate test, a sample of the heparinized blood nously for a variety of therapeutic purposes, at a rate of from the same dog was combined with an aqueous solu 0.1 to 2.0 grams per kilogram body weight. Typically, tion containing 20 wt. percent DMSO and 10 wt. per intravenously administered DMSO is in an aqueous 30 cent ethanol. The solution and blood were again com solution. bined so that 12 grams of DMSO were present for each Subjects receiving DMSO intravenously have suf 100 grams of blood in the resulting mixture. After mix fered from not only malodorous breath, but also from ing, only a small amount of red blood cell lysis oc red blood cell lysis which could lead to renal failure. It curred. The odor of dimethylsulfide was absent, but gas is now found that urea, intravenously administered at 35 chromatography analysis showed a small peak for di the same time as DMSO, substantially reduces both red methyl sulfide. blood cell lysis and malodorous breath. To effectively neutralize the hemolytic activity of EXAMPLE 28 DMSO and at the same time reduce expired dimethyl Two intravenous administration bottles were pre sulfide to acceptable levels, the amount of ethanol ad 40 pared. The first contained a 20 wt. percent aqueous ministered should be at least about 0.05 grams per kilo DMSO solution. The second contained an aqueous gram body weight, up to about 0.5 grams ethanol per solution having 20 wt. percent DMSO and 10 wt. per kilogram body weight. The ethanol should be adminis cent ethanol. Two mongrel dogs of about 15 kg. body tered in a weight ratio to DMSO of between about 1:40 weight were catheterized. Then, the two solutions were and 5:1. 45 delivered intravenously to the respective dogs so that Superior results are achieved when the DMSO and each dog received 15 g. of DMSO (1 g per kg body urea are administered together in a common composi weight). tion with the amount of urea being selected to be effec When the first solution (DMSO-water) was adminis tive in reducing the undesirable side-effects of malodor tered to one dog, the laboratory rapidly filled with ous breath and/or hemolysis which can be expected 50 dimethyl sulfide odor. This odor was detected within from administration of the DMSO. 30-45 seconds after administration. Urine collected To enhance penetrating activity of DMSO adminis from the dog during the first hours after administration tered intravenously, such solutions can contain sub was a deep red color, indicating severe red blood cell stances to enhance penetration of the DMSO. Such lysis. substances include urea, NaCl, KCl and/or acetamide. 55 When the second solution was administered to the In most instances, such solutions will contain water as a other dog, no dimethyl sulfide odor was detected by diluent. ------nose. Expired air from the animal was captured for a Although several possible mechanisms can be postu period of time in a toluene liquid trap. The trap was lated to explain the improvements resulting from intra operated for 0.25 hours and the toluene was then ana venously administered ethanol, the actual mechanism 60 lyzed by gas chromatography. There was a small peak whereby malodorous breath and red blood cell lysis are showing that a trace of dimethyl sulfide had been col reduced, is not understood. The following examples lected. illustrate the effectiveness of this treatment: Urine collected from the second dog was clean and on close observation was free of evidence of any red EXAMPLE 26 65 blood cell lysis. A unit of urine from the second dog was In a first test, nine volumes of whole, heparinized centrifuged and the sediment collected and observed. human blood were combined with one volume of a 50 There were traces of a red precipitate, presumably he volume percent aqueous solution of DMSO. The extent moglobin. This may have been due in part to traumati 4,296,104 19 20 zation during catheterization or might represent a very TABLE I-continued slight amount of red blood cell lysis. Test Solute Conc. (wt.%) Lethality 4. Hyperosmotic Administrations to Fish 3 NaCl 5 2 of 2 dead As mentioned above, urea, NaCl, and acetamide may 4. ( NaCl 3 be beneficial in compositions for treating mammals. But, DMSO 3 role these same substances may be toxic to fish at concentra 5 ( NaC 4 tions of as little as 3 wt. percent in an aqueous solution applied to an epithelial membrane of the fish. DMSO 3 O Because these substances are the driving force behind 10 6 ( NaCl 5 hyperosmotic treatment of fish, the toxicity places an DMSO 3 of 2 dead upper limit on the extent to which hyperosmotic treat ments can succeed. Clearly, lethality was reduced when DMSO was used in It is now found that the lethality of certain solutions combination with the NaCl. containing hyperosmotic concentrations of a membrane 15 It was also observed that the gills of all fish tested permeability altering agent can be reduced by adding turned whitish upon exposure to the hyperosmotic solu DMSO to the composition. Such reformulated compo tion. The degree of whiteness, a possible indicator of sitions can then be administered, in conjunction with osmotic stress, corresponded to the concentration of health and/or welfare modifying agents, to an epithelial NaCl. Less whiteness, at a given NaCl concentration, membrane, such as the gill membrane, of a water-living was observed in fish treated with the solutions contain animal. The risk of death from osmotic stress is greatly reduced when the DMSO is used. ing DMSO. Suitable procedures for administration are described EXAMPLE 30 in U.S. Pat. No. 4,112,946. Other procedures for admin The procedure of Example 29 was repeated, except istration to epithelial membranes, particularly gill mem 25 that after exposure to the hyperosmotic solutions, the brane, may also be used. A reduction in mortality rate goldfish were transfered to fresh water containing 0.5 will result in any instance where membranes is sub jected to a composition having a moderately high hy wt.% trypan blue dye. The results appear in Table II: perosmotic concentration of urea, NaCl and/or acetam ide. TABLE II That DMSO would have this beneficial effect is quite Test Solute Conc. (wt.%) Lethality surprising in view of the fact that DMSO is not greatly 7 NaCl 3 O effective as a membrane permeability altering agent for 8 NaC 4 2 of 2 dead treating the membranes of water living animals, particu 9 NaC 5 2 of 2 dead larly gilled animals. 35 O ( NaC 3 Other dipolar, aprotic solutes, such as dimethyl acet DMSO 3 Oe amide (DMAC) and dimethyl formamide (DMF) are ( NaC 4. also beneficial in reducing mortality rate. But, these are DMSO 3 ce less effective than DMSO, 12 ( NaCl 5 The amount of dipolar, aprotic solute to use with a particular solution containing a hyperosmotic concen EDMSO 3 de tration of urea, NaCl and/or acetamide, is best deter 13 la 8 de mined by experimentation. The amount should be suffi 14 rea 12 Oc cient to produce a desired reduction in mortality rate or 15 ( rea 8 cell damage, but should not be so high as to expose the 45 DMSO 5 de water-living animal subjects to toxic amounts of the 16 ( a 12 solute. Also, if the solution includes a health and/or DMSO 5 One welfare modifying agent along with the permeability altering agent, the amount of the dipolar, aprotic sub stance administered should not be so great as to inhibit 50 Again, decreased lethality was observed for fish ex transport of the health and/or welfare modifying agent posed to a NaCl solution, when DMSO was added. into the animals. The fish were observed for dye uptake. Trypan blue, The following examples illustrate how DMSO is being a vital exclusion type dye, only colors non-living effective in reducing mortality due to osmotic stress. cells. Fish exposed to the saline solutions without 55 DMSO demonstrated an increased dye uptake, pro EXAMPLE 29 nounced when the NaCl concentration was 5 wt. per cent. From the dye uptake patterns observed, it appears A series of tests were made using twelve small gold that NaCl without DMSO is highly lethal to epithelial fish (Carassius auratus). In each test, two fish were cells, especially of the gills, fins and tail. exposed, by imersion, to an aqueous test solution con 60 Similar results were observed in tests 13-16 where taining a hyperosmotic concentration of NaCl. The fish the fish were exposed to hyperosmotic concentrations were exposed for five minutes each and thereafter were of urea. Although none of the test animals died, dye transferred to fresh water and observed. The results are uptake was greater when DMSO was absent. summarized in Table I: These results are contrary to expectations because TABLE I 65 DMSO is known to facilitate tissue penetration in most Test Solute Conc. (wt.%) Lethality instances. If DMSO behaved in an expected manner, it 1 NaC 3 1 of 2 dead would enhance NaCl and urea penetration. Such in 2 NaC 4. 2 of 2 dead creased penetration should logically increase cell dam 4,296,104 21 22 age and, consequently, trypan blue uptake. But, the I claim: results of tests 7-16 show that the opposite is true. 1. In a method for administrating dimethyl sulfoxide DMSO protects cells from attack by hyperosmotic to a host, the improvement comprising reducing the NaCl and urea solutions, and does not aid in the attack. side effects of malodorous breath and skin reactions Furthermore, the results of tests 7-16 indicate that normally associated with such dimethyl sulfoxide ad DMSO not only reduces the lethality of hypersomotic ministration, comprising the step of: solutions, but also retards injury of cells exposed to administrating to said host a composition comprising extreme osmotic challenge. dimethylsulfoxide and a side-effect reducing agent taken from the group consisting of urea, or ethanol EXAMPLE 31 10 or mixtures thereof in an amount which is effective Seven fingerling salmonids of 4-5 inch length were for said purpose. netted in the Washougal River (State of Washington). 2. A method according to claim 1 wherein said con The species was not identified; but most likely the fin position further includes acetamide. gerlings were wild, coho salmon (Oncorhynchus ki 3. The method according to claim 1 wherein said sutch). 15 composition further includes sodium chloride. Four of the fingerlings were immersed in an aqueous 4. The method according to claim 1 wherein said solution containing 5 wt. percent NaCl and 3 wt. per composition further includes potassium chloride. cent DMSO. After five minutes, the fingerlings were 5. The method according to claim 1 wherein said returned, for observation, to a holding pond containing composition further includes water. river water. The procedure was repeated with the re 20 6. The method according to claim 1 wherein the maining three fingerlings being immersed in an aqueous weight ratio of urea to dimethyl sulfoxide is greater solution containing only 5 wt. percent NaCl. than 1:99. All of the fingerlings lost their friskiness when ex 7. The method according to claim 1 wherein the ratio posed to a hyperosmotic solution, but all were alive of urea to dimethyl sulfoxide is greater than 1:9. when transferred to the holding ponds. After three 25 8. The method according to claim 1 wherein said hours, the three fingerlings exposed to the NaCl solu composition has the following formulation: tion without DMSO had died. The other four finger lings were alive and were released into the river. dimethyl sulfoxide about 19 grams; While I have described and given examples of pre rea about 9 grams; ferred embodiments of my invention, it will be apparent 30 sodium chloride about 1 gram; and to those skilled in the art that changes and modifications water at least about 6 grams; may be made without departing from my invention in its broader aspects. I therefore intend the appended whereby the aforementioned composition formulation claims to cover all such changes and modifications as 35 is provided with the weight ratios noted. fall within the true spirit and scope of my invention. :k 3k k . k sk

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