Abstract:
Dihalonitromethylisoxazoles have been prepared from the corresponding nitronate salts. The compounds are useful as broad spectrum antibacterials, antifungals, antiprotozoals and aquatic biocides.

Description:
BACKGROUND OF THE INVENTION 
     The invention herein described was made in the course of or under a contract with the Office of Naval Research, Department of the Navy. 
    
    
     This invention concerns novel dihalonitromethyl heterocyclic ring compounds which are useful in the control of a variety of harmful microorganisms. 
     The nitration of active methylene compounds containing an activating group such as an ester, ketone or cyano group has been previously described. See Feuer et al., J. Am. Chem. Soc., 78, 4364 (1956), Ibid., 81, 5826 (1959), J. Org. Chem., 29, 939 (1964), Ibid., 31, 3152 (1966), Ibid., 34, 991 (1969); and Klager, Ibid., 20, 646 (1955). 
     In addition, p-anisylnitromethane and its ring nitro derivatives have been prepared by Zalukajevs et al., Latvijas P.S.R. Zinatnu Akad. Vestis, 109 (1956). The same auther also prepared α-naphthylnitromethane, J. Gen. Chem. U.S.S.R., 26, 657 (1956). 
     Primary or secondary nitro compounds form nitronate salts which react with bromine to form bromonitro compounds. In the case of α-nitrocyclic ketones, cleavage can occur upon bromination. See Feuer et al., J. Org. Chem., 29, 939 (1964), Ibid., 33, 3622 (1968), Ibid., 34, 991 (1969). The preparation of halo derivatives of various nitromethyl heterocycles was recently disclosed by Feuer et al., J. Org. Chem., 37, 3662 (1972). Zalukajevs et al. prepared the halo derivatives of 2-nitromethylquinoline, Zhur. Obshchei Khim., 28, 483 (1958). 
     There is no suggestion in any of the above-described publications that any of the compounds possess antimicrobial activity. 
     Belgian Patent No. 702,570 discloses 1-aryl-2-nitrohaloethanes useful in the control of bacteria, fungi and algae in water and aqueous compositions. Gum et al., U.S. Pats. Nos. 3,703,515 and 3,754,042, disclose dihalonitromethylsutstituted quinoxalines and cycloalkanes, respectively, which are said to have antimicrobial activity. 
     SUMMARY 
     This invention provides to the art a class of novel dihalonitromethyl heterocyclic ring compounds which possess strong activity against bacteria, fungi, and protozoa. Such compounds are those having the formula 
     
         R -- CX.sub.2 NO.sub.2 
    
     wherein X represents chloro or bromo; 
     R represents ##SPC1## 
     E represents oxygen or sulfur; 
     Ar combined with the two carbon atoms to which it is attached forms phenyl or naphthyl; 
     Each of the R 1  groups independently represents C 1  -C 3  alkyl or phenyl; 
     n represents 0-2 when R 1  represents C 1  -C 3  alkyl; 
     n represents 0 or 1 when R 1  represents phenyl; 
     R 2  represents hydrogen, C 1  -C 3  alkyl, or phenyl; 
     R 3  represents hydrogen or C 1  -C 3  alkyl; 
     R 4  represents C 1  -C 3  alkyl. 
     DESCRIPTION OF THE PREFERRED EMBODIMENT 
     The compounds of this invention are characterized by a dihalonitromethyl group attached to a heterocyclic ring, which may be substituted with lower alkyl or phenyl groups. 
     The term C 1  -C 3  alkyl is used in the formula above to refer to alkyl groups such as methyl, ethyl, n-propyl and isopropyl. 
     The term Ar is used in the structures above to indicate that the scope of this invention includes fused-ring compounds wherein a thiazole or oxazole ring is fused to a phenyl or naphthyl ring to form compounds wherein the dihalonitromethyl group is 2-substituted on rings such as benzoxazole and naphthothiazole. 
     While the generic formula above describes the compounds unambiguously, the following compounds are named to assure that those skilled in the art understand the invention. 
     2-dichloronitromethyl-1,3,4-oxadiazole 
     2-dibromonitromethyl-5-isopropyl-1,3,4-oxadiazole 
     2-dibromonitromethyl-5-phenyl-1,3,4-oxadiazole 
     2-dichloronitromethyl-1,3,4-thiadiazole 
     2-dichloronitromethyl-5-phenyl-1,3,4-thiadiazole 
     2-dichloronitromethyl-5-methyl-1,3,4-thiadiazole 
     2-dibromonitromethyl-2-oxazoline 
     2-dibromonitromethyl-4,5-diethyl-2-oxazoline 
     2-dibromonitromethyl-5-isopropyl-2-oxazoline 
     2-dichloronitromethyl-4-phenyl-2-oxazoline 
     2-dichloronitromethyl-5-ethyl-2-thiazoline 
     2-dichloronitromethyl-2-thiazoline 2-dibromonitromethyl-4,5-dimethyl-2-thiazoline 
     2-dichloronitromethyl-4-phenyl-2-thiazoline 
     2-dibromonitromethylthiazole 
     2-dibromonitromethyl-4,5-dimethylthiazole 
     2-dichloronitromethyl-4-ethylthiazole 
     2-dibromonitromethyl-5-phenylthiazole 
     2-dichloronitromethyloxazole 
     2-dichloronitromethyl-4-phenyloxazole 
     2-dichloronitromethyl-4-propyloxazole 
     2-dibromonitromethyl-4,5-diethyloxazole 3-dichloronitromethyl-5-phenylisoxazole 
     5-dichloronitromethyl-3,4-dimethylisoxazole 
     5-dibromonitromethyl-3-isopropylisoxazole 
     2-dibromonitromethylbenzothiazole 
     2-dibromonitromethyl-5-ethylbenzothiazole 
     2-dichloronitromethyl-4,6-dimethylbenzothiazole 
     2-dibromonitromethyl-6-phenylbenzothiazole 
     2-dibromonitromethyl-4-phenylbenzoxazole 
     2-dichloronitromethyl-7-ethylbenzoxazole 
     2-dibromonitromethylbenzoxazole 
     2-dibromonitromethyl-5,7-diethylbenzoxazole 
     2-dibromonitromethylnaphtho[1,2-d]thiazole 
     2-dichloronitromethyl-5-phenylnaphtho[2,1-d]thiazole 
     2-dichloronitromethyl-4,6-diethylnaphtho[2,1-d]thiazole 
     2-dichloronitromethyl-8-ethylnaphtho[1,2-d]thiazole 
     2-dichloronitromethyl18-methylnaphtho[1,2-d]oxazole 
     2-dichloronitromethyl-6,9-dipropylnaphtho[2,1d]oxazole 
     2-dichloronitromethylnaphtho[1,2-d]oxazole 
     2-dibromonitromethyl-5-phenylnaphtho[2,1-d]oxazole 
     A preferred subgenus of this invention comprises compounds wherein the R group contains a ring oxygen atom. The preferred individual compounds of the invention are 3-dibromonitromethyl-5-phenylisoxazole, 2-dibromonitromethylbenzoxazole, 3-dichloronitromethyl-5-phenylisoxazole, 5-dichloronitromethyl-3-methylisoxazole, and 2-dichloronitromethyl-2-oxazoline. 
     The compounds of this invention are easily prepared in a two-step synthesis from methyl-substituted heterocyclic ring compounds having the desired R 1 , R 2 , R 3 , or R 4  substituents. Such compounds are commercially available or can be prepared using known procedures in the chemical literature. 
     The first step of the synthesis is the formation of a nitronate salt, having the structure 
     
         R - CH = NO.sub.2 .sup.-M.sup.+ 
    
     In the formula, R represents the heterocyclic ring described above, and M represents alkali metal, preferably potassium. 
     Nitronate salts are formed by the procedure described by Feuer et al., J. Am. Chem Soc., 91, 1856 (1969). A methyl heterocycle is treated with an alkali metal amide in liquid ammonia, followed by n-propyl nitrate as the nitrating agent. The reaction is conducted at a temperature from -75° to -30°C. and is generally complete in less than one-half hour. For a discussion of possible variations in the formation of nitronate salts, including solvent effects and the different bases that may be employed, see Feuer et al., J. Am. Chem. Soc., 78, 4364 (1956). One skilled in the art will also recognize that other nitrating agents, such as amyl nitrate, can be used in the formation of nitronate salts. 
     The nitronate salt is converted to the dihalonitromethyl compounds of the present invention by treatment with a halogenating reagent. The dichloro compounds can be prepared from the nitronate salts by treating with an agent such as potassium hypochlorite at temperatures between -10°C. and 20°C. The corresponding dibromo compounds are best prepared by treatment with bromine in potassium hydroxide solution under similar conditions. The use of a two-fold excess of the aqueous halogenating reagent forms the dihalonitro compound, usually in less than one hour. 
     The preparative examples below illustrate the methods by which the compounds are made, and are not intended to limit the scope of the invention in any way. The first example shows the synthesis of a typical nitronate salt. 
    
    
     EXAMPLE 1 
     potassium 3-methyl-5-phenylisoxazolenitronate 
     Liquid ammonia, 250 ml., was placed in a flask equipped with a mechanical stirrer, thermometer, and Dry Ice condenser. With the temperature kept below -33°C. by external cooling using a Dry ice-acetone bath, a catalytic amount of ferric nitrate monohydrate was added, followed by 5.9 g. of metallic potassium. The mixture was stirred at temperatures below -33°C. until the mixture formed a gray suspension. To the ammonia solution was then added 15.8 g. of 3-methyl-5-phenylisoxazole in 30 ml. of anhydrous ethyl ether, and the mixture was stirred at reflux (approximately -33°C.) for 15 minutes more. the mixture was then cooled to -40°C. to -50°C., and 20.9 g. of n-propyl nitrate was added at such a rate as to control the temperature below -35°C. After stirring for 30 minutes at -33°C., the ammonia was allowed to evaporate and was replaced with 200 ml. of ethyl ether. The solid which formed was filtered, washed with ethyl ether, and dried under vacuum. The product, potassium 3-methyl-5-phenylisoxazolenitronate, was obtained in a sufficiently pure form to use in the following step without further purification. The yield was 20.9 g. 
     The intermediate nitronate salts are converted to the new dihalonitromethyl compounds by processes typified by the following example. 
     EXAMPLE 2 
     3-dibromonitromethyl-5-phenylisoxazole 
     A 5.9 g. portion of the nitronate salt made in Example 1 was dissolved in 45 ml. of water, and the solution was filtered. The solution was then added to a solution of 15.1 g. of KOH and 15.1 g. of bromine in 60 ml. of water at 0°-5°C. The reaction mixture was stirred for 1 hour at about 5°-10°C. during which time a precipitate formed. The solids were collected by filtration, washed with water and dried. The product was recrystallized from hexane, and identified by nuclear magnetic resonance analysis. The yield was 0.50 g. of 3-dibromonitromethyl-5-phenylisoxazole (I), m.p. 72°-74°C. 
     The following exemplary compounds were made by the processes of Examples 1 and 2 with small modifications which can readily be supplied by one skilled in the art. 
     2-dichloronitromethylnaphtho[2,1-d]thiazole (II), m.p. 114°-116°C. 
     2-dibromonitromethylbenzoxazole (III), m.p. 76°-80°C. 
     2-dibromonitromethylnaphtho[1,2-d]thiazole (IV), m.p. 116°-117°C. 
     2-dichloronitromethyl-5-methyl-1,3,4-thiadiazole (V), oil 
     2-dichloronitromethyl-4-methylthiazole (VI), oil 
     4-dibromonitromethyl-2-methylthiazole (VII), 95-95-98°c. 
     5-dibromonitromethyl-3-methylisoxazole (VIII), m.p. 59°-62°C. 
     5-dichloronitromethyl-3-methylisoxazole (IX), oil 
     3-dichloronitromethyl-5-phenylisoxazole (X), m.p. 65°-67°C. 
     2-dichloronitromethyl-2-oxazoline (XI), oil 
     2-dichloronitromethylnaphtho[1,2-d]thiazole (XII), m.p. 108°-112°C. 
     2-dichloronitromethylbenzothiazole (XIII), m.p. 38°38°43°C. 
     2-dichloronitromethylbenzoxazole (XIV), m.p. 41°-45°C. 
     2-dibromonitromethylbenzothiazole (XV), m.p. 71°-73°C. 
     2-dibromonitromethyl-5-methyl-1,3,4-thiadiazole (XVI), m.p. 76°-72°C. 
     2-dibromonitromethylnaphtho[2,1-d]thiazole (XVII), m.p. 129°-130°C. 
     2-dichloronitromethyl-2-thiazoline (XVIII), m.p. 58°-62°C. 
     2-dibromonitromethyl-2-oxazoline (XIX), m.p. 60°-62°C. 
     The compounds of the present invention are biocides in a broad sense. As the examples below indicate, the compounds kill or control microorganisms such as bacteria, fungi, protozoa, and algae, as well as some aquatic weeds. Those skilled in the art will appreciate that the biocidal properties of the compounds make them useful in many ways. 
     For example, the compounds can be added to bodies of water such as cooling towers and ponds, lagoons, lakes and the like for the control of aquatic weeds, algae, and slime-forming microorganisms. The control is brought about by adding an effective, economical amount such as from about 0.1 to about 100 ppm. of the compound to the water to be protected from aquatic organisms. It is often convenient to formulate the compound in an easily water-dispersible mixture before addition. Such formulations are known to the art, and comprise a finely divided powder, or a solution or suspension of the compound in a liquid such as water or an organic solvent, to which surfactants such as ethylene oxide adducts of nonylphenol and alkylbenzenesulfonates are often added to increase the dispersibiility of the mixture. 
     The compounds can also be added to such compositions as adhesives, inks, plasticizers, latices, polymers, resins, fuels, lubricants, soaps and detergents, cutting oils, and paints to prevent the growth of mold and the degradation of the products which results from attack by microorganisms. 
     The compounds can also be coated on or distributed through products such as textiles, paper and other cellulose products and may be impregnated into wood, wall paneling and plaster to protect such products from mold and decay caused by microbial infestation. The compounds are especially useful for the preservation of such products as cosmetic formulations. 
     The compounds are valuable disinfectants and sterilizing agents for surfaces such as floors, walls, hospital equipment, kitchen equipment and the like. 
     The compounds of this invention can be used to control microorganisms, especially fungi, growing on the skin and outer tissues of animals. The examples below show, for example, that the compounds are especially effective in the control of Candida tropicalis and Trichomonas vaginalis, both of which are important causes of vaginal infections. The compounds are used for the control of such infections by formulating them into ointments, creams and the like according to the usual pharmaceutical methods and applying them topically to the site of the infection. 
     The outstanding efficacy of the compounds against representative microorganisms is illustrated by the examples below. 
     EXAMPLE 3 
     in vitro agar dilution test 
     Bacteria and fungi in an agar medium were stamped on a plate to which one drop of a 100 μg./ml. or a 10 μg./ml. solution of the compound was surface applied. The agar plates were then incubated at 35°C. for 12 hours, at which time the antibacterial activities were evaluated. The fungi were incubated at 25°C. for an additional 60 hours before evaluation. The procedure was essentially that recommended in Acta Pathol. Microbiol. Scand. B., Suppl. 217, 11 (1971). A rating of 100 indicates that the compound prevented growth of the microbe at 100 μg./ml. and 10 indicates prevention at the 10 μg./ml. level. NT indicates the compound was not tested for activity against an organism, while a blank space indicates the compound was not active at the highest level tested (100 μg./ml.). 
     
         __________________________________________________________________________Com-    Staphylo-     Streptocc-            Proteus                 Salmonella                       Klebsiella-                               Klebsiella-                                       Escherichia                                              Pseudomonas                                                     C-476pound    coccus     us          typhosa                       aerobacter sp.                               aerobacter sp.                                       coli   aeruginosa    aureus     faecalis          KA14    KA17__________________________________________________________________________I   10    100    100  100   10      10      10     100    100II  10    10      10III 10    10     100   10   10      10      10      10     10VIIX   10    10      10   10   10      10      10      10    100XII 100   100XIII    100   100    100  100           100                   100XIV 10    100    100  100   100     100     10     100    100XVIII    100   100    100  100   100     100     100           100__________________________________________________________________________Com-    Salmon-    Pseudomonas           Erwinia                 Xanthomo-                       Candida                            Trichophyton                                   Botrytis                                        Cerato-                                             Fusarium                                                    Verticill-pound    ella solanacearum           amylovora                 nas   tropic-                            mentagrophy-                                   cinerea                                        cystis                                             oxysporum                                                    ium    typhi-            phaseoli                       alis tes         ulmi lycopersici                                                    albo-atrum    murium__________________________________________________________________________I   10   10     100    10   100  10     10    10  100    10II              100                     100  100         100III 10   10      10    10        10     10   100  100    10VII                                     100X   10   10     100    10    10  10     10    10  10     10XIIXIII    100         100   100        100    100  100  100    100XIV 100  100    100   100        10     10XVIII    100  100    100   100   100  10     100  100  100    100__________________________________________________________________________ 
    
     
              Staphyloccus        Staphyloccus                Streptoccus                        Proteus Salmonella                                        Klebsiella                                                 EnterobacterCompoundaureus 3055        aureus 3074                faecalis X66                        Morganii Pris                                typhosa SA12                                        pneumoniae                                                 Aerogenes__________________________________________________________________________                                                 EB17IV   100     100     100     100     100     100      100V                                            100VI   100     100     100             100VIII 100     100     100     100     100     100      100IX   100     100     100     100     100     100      100XI    10      10     100      10      10     100      100XV   100     100     100     100     100     100      100XVI  100     100     100     100     100     100      100XVII 100     100     100     100     100     100      100XIX  100     100     100     100     100     100      100__________________________________________________________________________     Serratia Escherichia                     Citrobacter                              Pseudomonas                                        Bordetella SalmonellaCompound  marcescens SE3              coli EC14                     freundii CF17                              aeruginosa X239                                        bronchiseptica                                                   typhimurium__________________________________________________________________________IV        100      100    100      100       100        100V                                            100VI                                           100VIII      100      100    100      100       100        100IX        100      100    100                100        100XI        100       10    100      100        10         10XV        100      100    100      100       100        100XVI       100      100    100      100       100        100XVII      100      100    100      100       100        100XIX       100      100    100      100       100        100__________________________________________________________________________       Psuedomonas                  Erwinia                        Candida  Trichophyton                                            Aspergillis                                                   Ceratocystis  Compound       solanacearum X185                  amylorora                        tropicalis A17                                 mentagrophyles 27                                            flavis                                                   ulmi__________________________________________________________________________  IV   100        100   100      100        100    100  V                     100                        100  VI              100   100       10         10     10  VIII 100        100   100      100        100    100  IX   100        100   100       10         10     10  XI   100         10    10      100        100     10  XV   100        100   100      100        100    100  XVI             100   100      100        100    100  XVII            100   100      100        100    100  XIX  100        100   100      100        100    100__________________________________________________________________________ 
    
     The example below reports representative results of testing the compounds in a slightly different in vitro test against a different group of microorganisms. 
     EXAMPLE 4 
     in vitro tube dilution test 
     The organisms against which the compounds were to be tested were grown in nutrient broth in test tubes under sterile conditions. One hundred μg./ml. of the compound to be tested was added to a tube of broth, and the treated broth was serially diluted with untreated broth in which the culture had been inoculated. The tubes were observed, and the results for each compound were recorded as the lowest concentration, in micrograms per milliliter, in which the compound prevented the growth of the microorganism. 
     
         __________________________________________________________________________Staphylococcus        Streptococcus  Mycoplasma                               Escherichia                                      Salmonella                                            PseudomonasCompoundsp. 1130        sp. 80  Vibrio coli                       gallisepticum                               coli   dublin                                            sp.__________________________________________________________________________I    50      50      50     25      &gt;50    &gt;50   &gt;50III  50      50      25     25      &gt;50    &gt;50    50IV   &gt;50     &gt;50     &gt;50    50      &gt;50    &gt;50   &gt;50V    &gt;50     50      50     25      &gt;50    &gt;50   &gt;50VI   12.5    12.5    25     6.25     50     25   &gt;50VII  &gt;50     &gt;50     50     &gt;50     &gt;50    &gt;50   &gt;50VIII &gt;50     &gt;50     &gt;50    50      &gt;50    &gt;50   &gt;50IX   25      25      &lt;.78   25       50     50   &gt;50X    12.5    6.25    25     12.5     25     25    50XI   12.5    25      6.25   6.25     25     25   &gt;50XII  25      12.5    3.12   25      &gt;50    &gt;50   &gt;50XIII 50      25      25     12.5     50     50    50XIV  25      25      12.5   50       50     50   &gt;50XV   &gt;50     &gt;50     &gt;50    &gt;50     &gt;50    &gt;50   &gt;50XVI  &gt;50     &gt;50     50     50      &gt;50    &gt;50   &gt;50XVII &gt;50     &gt;50     50     50      &gt;50    &gt;50   &gt;50XVIII12.5    6.25    6.25   6.25     12.5   12.5  50XIX  &gt;50     &gt;50     50     25      &gt;50    &gt;50    50__________________________________________________________________________         Pasteurella                P.  multocida Mycoplasma    Compound         multocida                turkey isolate                        Bordetella                              syroviae                                    M.  hyorhinis                                           M.  granularum__________________________________________________________________________    I    25     25      50    50    50     25    III  50     50      50    25    50     &gt;50    IV   50     &gt;50     50    25    25     50    V    50     50      50    25    &gt;50    50    VI   12.5   6.25    50    12.5  25     12.5    VII  &gt;50    50      &gt;50   50    50     50    VIII &gt;50    &gt;50     &gt;50   50    50     &gt;50    IX   12.5   3.12    50          50     25    X    3.12   3.12     12.5 12.5  50     6.25    XI   6.25   12.5     12.5 12.5  50     50    XII  12.5   25      25    12.5  25     12.5    XIII 25     25      50    25    50     25    XIV  12.5   12.5    25    12.5  50     25    XV   &gt;50    &gt;50     &gt;50   &gt;50   &gt;50    &gt;50    XVI  50     50      &gt;50   50    50     &gt;50    XVII 50     &gt;50     &gt;50   50    50     &gt;50    XVIII         3.12   6.25     12.5  12.5 25     6.25    XIX  &gt;50    &gt;50     &gt;50   50    50     50__________________________________________________________________________ 
    
     A generally similar test was performed to evaluate the compounds against additional species of fungi. 
     EXAMPLE 5 
     in vitro antifungal test 
     
         Compound   Trichophyton    CandidaNo.        mentagrophytes  albicans______________________________________VI         0.2          μg./ml.                          2.0   μg./ml.VIII       &gt;2.0                20XI         0.2                 20XIII       0.02                0.2XIV        0.02                2.0XV         0.002               20XVI        &gt;2.0                20XVIII      0.2                 2.0XIX        2.0                 20______________________________________ 
    
     In a different antifungal test, the compounds to be tested were absorbed on paper discs, and the discs were laid on plates of fungus-infected agar medium. The results were reported as the last amount of compound per disc which produced a measurable inhibition of the fungus. 
     
         ______________________________________Compound  Trichophyton    CandidaNo.       mentagrophytes  albicans______________________________________X         0.625 μg./ml.                     0.078 μg./ml.______________________________________ 
    
     Compounds of this invention were tested in vitro to determine their ability to control protozoa and algae. The examples below report representative results of such tests. 
     EXAMPLE 6 
     in vitro protozoa inhibition tests 
     The tests were run against four representative protozoa, Tetrahymena pyriformis (T), Ochromonas malhamensis (O), Euglena gracilis (E), and Trichomonas vaginalis (TV). The protozoa were grown in the laboratory in nutritive media. When a test was to be run, nutritive medium containing the test protozoa was mixed with sterile agar medium, and the mixed medium poured into plates. 
     Absorbent discs were treated with 0.02 ml. of a solution containing a known concentration of the compound to be tested. The concentrations used are indicated in the table below. The discs were placed on the surfaces of the protozoa-containing agar plates. 
     The plates were incubated for a time, and were then inspected to determine if the test compounds absorbed on the discs had inhibited the growth of the protozoa. Inhibition, if present, was measured as the diameter in millimeters of the zones of inhibition surrounding the discs. 
     The table below reports the zones of inhibition produced by representative compounds of this invention. The notation TR indicates that the compound gives a trace of inhibition. NT indicates that the compound was not tested against a given organism, and a blank space indicates that the compound was inactive. 
     
         ______________________________________CompoundNo.     Conc.     T        O      E      TV______________________________________I       2000               17     28     11   500                12     17     TR   125                TR     10III     2000      46       25     50     15   500       25       15     20     10   125       14       10     11IV      2000      25       25     23     11   500       12       14     10   125       TRV       2000      26       30     38   500       16       17     16   125       11       11     11VI      2000      17       19     22   500       TR       12     11X       2000      16       18     13     36   500       13       12     12     23   125       TR       TR     TR     15XI      2000                             14XII     2000               10     17   500                       TRXIII    2000      17       25     24   500       15       18     17   125       13       14     15XIV     2000      24       18     20     12   500       18       17     17   125       14       TR     TRXV      2000      20       24     25     TR   500       12       11     10XVI     2000      16       18     28     11   500       12       TR     12     TRXVII    2000               26     38     14   500                12     18     TR   125                TRXVIII   2000      20       17     20     10   500       13       12     10     TR   125       12       10     TRXIX     2000      25       17     24     15   500       13       14     12     TR______________________________________ 
    
     EXAMPLE 7 
     in vitro algae inhibition test 
     The activity of the compounds against a typical alga was determined by conducting a test, substantially identical to the test described above, against Chlorella vulgaris. Representative results are reported below. 
     
         ______________________________________CompoundNo.            Conc.           Chlorella______________________________________I              2000            72          500             32          125             10III            2000            77          500             40          125             12IV             2000            39          500             19          125V              2000            26          500             13          125VI             2000            10          500X              2000            16          500             13          125             11XI             2000XII            2000            TR          500XIII           2000            21          500             15          125             TRXIV            2000            30          500             17          125             TRXV             2000            38          500             16XVI            2000            42          500             13XVII           2000            58          500             20          125XVIII          2000            42          500             23          125             10XIX            2000            18          500             11______________________________________ 
    
     The tests reported below show the ability of the compounds of the invention to eradicate microorganisms established in a culture. 
     EXAMPLE 8 
     in vitro eradication test 
     Aqueous dispersions of 3-dibromonitromethyl-5-phenylisoxazole were inoculated to contain known concentrations of the organisms shown below. Viable organism counts, in organisms per milliliter, were made initially and after times of 25°C. incubation shown in the table below. A count of less than 100 organisms per milliliter is regarded as equivalent to complete kill of the organism. 
     
         ______________________________________0.5% of Compound       Pseudomonas      AspergillusDays        aeruginosa       niger______________________________________ 0          3.4 × 10.sup.6                        6 × 10.sup.3 1           &lt;100             &lt;100 7           &lt;100             &lt;10028           &lt;100             &lt;100______________________________________ 
    
     
         0.1% of Compound       Pseudomonas      AspergillusDays        aeruginosa       niger______________________________________ 0          6.4 × 10.sup.6                        5 × 10.sup.3 1           &lt;100            4 × 10.sup.3 7           &lt;100             &lt;10014            NT              &lt;10028           &lt;100             &lt;100______________________________________ 
    
     Similar tests were performed with 3-dichloronitromethyl5-phenylisoxazole. 
     
         ______________________________________0.5% of Compound       Pseudomonas      AspergillusDays        aeruginosa       niger______________________________________ 0          5.9 × 10.sup.6                        1 × 10.sup.4 1          1.9 × 10.sup.5                        2.7 × 10.sup.3 7          &lt;100             &lt;10014          &lt;100             &lt;10028          &lt;100             &lt;100______________________________________ 
    
     A similar test, against a broader range of microorganisms, was performed with 3-dibromonitromethyl-5-phenylisoxazole as the test compound. In this test, the fungi were inoculated in Sabouraud dextrose agar, and the bacteria in soybean-casein medium. The test was otherwise similar to the test described immediately above. 
     
         __________________________________________________________________________0.05% of Compound   Staphylococcus      Escherichia             Pseudomonas                    Candida                           Aspergillus                                  StreptococcusDays   aureus  coli   aeruginosa                    albicans                           niger  faecalis__________________________________________________________________________ 0 4.3 × 10.sup.6      6.9 × 10.sup.6             4.5 × 10.sup.6                    3.2 × 10.sup.6                           3.2 × 10.sup.5                                  2.5 × 10.sup.6 1 &lt;100    &lt;100   &lt;100   3.6 × 10.sup.5                           3.9 × 10.sup.5                                  &lt;100 7 &lt;100    &lt;100   &lt;100   3.7 × 10.sup.4                           4.4 × 10.sup.5                                  &lt;10014 NT      NT     NT     &lt;100   1.7 × 10.sup.5                                  NT21 NT      NT     NT     &lt;100   7.5 × 10.sup.2                                  NT28 &lt;100    &lt;100   &lt;100   &lt;100   &lt;100   &lt;100__________________________________________________________________________ 
    
     The compounds also kill or control many aquatic weeds. For example, hydrilla, coontail, and duckweed were controlled by adding 10 ppm. of 2-dibromonitromethyl-2-oxazoline to the water in which the weeds were growing. Similar weed control was produced by, for example, 3-dichloronitromethyl-5-phenylisoxazole and 2-dibromonitromethylbenzothiazole.