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I N D U S T R I A L A N D E N G I N E E R I N G CHEMISTRY
plasticized with a fatty acid, preferably rather highly unsaturated, such as soybean fatty acid. It can be applied either as an alcoholic solution or as an aqueous dispersion made as previously described. Zein is outstanding in its resistance to penetration by greases or oils, and therefore has a large potential use in the making of food wrappings or containers where this property is of importance. This property is also useful in that a very thin layer on paper or other absorbent material will permit the application of a wax coating without penetration of the paper. It is also effective as a sealing coat against asphalt or phenolic resins which are very difficult to seal with other available materials. Fatty acids, or fatty acids and a resin, are the preferred nonzein constituents. Because of its cost zein can be used as an adhesive only in cases where its specific properties can justify its use. I n the application of high-grade wood veneers it is very useful in that the adhesive does not strike through the wood. A waterproof bond for this purpose, joining the veneer either to a wood or transite base, is made by using a solution of zein containing formaldehyde and an organic acid (9) and curing under pressure at an elevated temperature. A laminated board made from zein-impregnated paper is tough and fairly flexible, can be sheared and punched easily, or can be embossed by hot pressing. Uses for this material will be chiefly of a decorative nature where low cost, toughness, and ease of producing satisfactory colors are important, and where water resistance is not essential. Because of its high molecular weight, the impregnation of paper with zein is rather difficult, but a satisfactory product can be made by using an absorbent paper and applying the zein in a fairly concentrated solution a t a slightly elevated temperature. The use of rosin, ester gum, shellac, or resins of similar nature in amounts up to 50 per cent of the weight of the zein improves the flow in the laminating process, and in many cases reduces the cost without impairing the mechanical properties of the finished product. The impregnated paper is pressed dry at
Vol. 33, No. 3
about 100" C. under a pressure of 500 to 2000 pounds per square inch, depending on the plasticizers used. Zein is useful in solid color printing where the chief object is to coat a paper or cardboard surface as evenly as possible with a dye. It aids in the spread of the dye and brings out the true color of the dye in the solution rather than the metallic appearance due to dye crystals which is characteristic of many vehicles used for this purpose. I n many cases the fastness of the dye to light is improved. Bleeding of the dye may be decreased or eliminated. The production of films and fibers from zein is expected to provide outstanding uses. However, these developments are being deferred until more satisfactory methods of curing zein have been developed.
Acknowledgment The writer wishes to acknowledge the assistance of numerous members of the Research, Engineering, and Sales Departments of the Corn Products Refining Company in the development work described in this paper,
Literature Cited (1) Bizio, Giorn. di fisica, chimica, storia naturale, medicine ed arti Brugnatelli, [2] 5 , 127-35, 180 (1822). (2) Chittenden and Osborne. Am. Chem. J.. 13. 453-68. 629-62 (1891); 14, 20-44 (1892). (3) Cohn, C. J., Berggen, R., and Hendry, J., J. Cen. Physiol., 7,8198 (1924). (4) Gorharn, J., Quart. J. Sci., Literature & Arts, 11. 206-8 (1821). (6) Osborn, T. B..J. Am. Chem. SOC.,19, 524-32 (1897). (6) Osborn, T. B., and Clapp, S. H., A m . J . Physiol., 20, 477-93 (1908). (7) Ritthausen, J. prakt. Chem., 106, 471-89 (1869). (8) Showalter, M. F., and Carr, R. H., J. Am. Chem. SOC.,44. 2019-23 (1922). (9) Sturken, O., U. S. Patent 2,115,240 (April 26, 1938). (10) Watson, C. C., Arrhenius, S., and Williams, J. W., Nature, 137, 322 (1936).
.----,
PRESENTED before the Division of Industrial and Engineering Chemistry s t the 98th Meeting of the American Chemical Society, Boston, Mass.
Antiseptic and Germicidal SAMUEL S. EPSTEIN AND FOSTER DEE SNELL Foster D. Snell, Inc., Brooklyn, N. Y.
I
N RECENT years, medical and food-processing authorities have shown a growing interest in air-borne infections and in air-borne contaminations. The use of antiseptic and germicidal paints has been proposed as a helpful factor in reducing atmospheric microbial pollution. The purpose of this paper is t o review the literature on the subject of antiseptic and germicidal paints, particularly with respect to methods of testing, and to present some of our own test methods and results. Frequent washing of painted surfaces is recognized as a good sanitary practice. Robb ( 8 ) found that thorough scrubbing of painted walls effected a greater reduction in the number of air bacteria in the operating room than washing the floors with 0.1 per cent solution of mercuric bichloride or 5 per cent carbolic acid. In tracing the causes of occasional severe staphylococci infections originating in an operating room, Hart (6) always found Staphylococcus aureus on walls and ceilings despite frequent painting and daily washing.
Two approaches to the preparation of germicidal paints have been used. The first is simply the addition of antiseptic agents to otherwise standard paints. The second is modification of the vehicle by addition of halogen to the oil. Practically speaking, the latter means chlorination of linseed oil to a 4 per cent halogen content. Portier and Kling (7) found that the incorporation of chlorophenols rendered paints antiseptic, as based on tests with Escherichia coli and Staphylococcus aureus. One drop of a broth culture was deposited on a painted metal surface and allowed to dry in light and in darkness for 24 hours. A drop of sterile water was placed on the dried bacterial culture and allowed to remain for 15 minutes, after which the moistened area was swabbed with sterile cotton and the cotton inoculated into broth. Lack of growth indicated antiseptic power. Normal development of fresh inoculations of test cultures in such broth excluded the factor of inhibition. Using substantially the same technique, Troussaint (9) re-
March, 1941
INDUSTRIAL AND ENGINEERING CHEMISTRY
ported that oxyquinoline sulfate renders a paint germicidal. His modifications of technique included use of saline cultures and moistened cotton in place of broth cultures and the drop of sterile water. Additional organisms studied were Vibrio cholera, B. pestis, B. diphtheriae, B. typhosus, B. paratyphi A , and B. paratyphi B. More oxyquinoline sulfate was required for the last four cultures than for the others studied. As controls, scraping of drop saline cultures after contact with unpainted glass surfaces for 24 hours yielded growth in broth with six of the eight test organisms. The cholera and pestis bacilli apparently succumbed when exposed for 24 hours on ordinary glass.
AN ELABORATE two-year study by ValleB (11) involved more than three thousand bacteriological examinations. His antiseptic was a colloidal jelly compounded from halogenated compounds and aromatic aldehydes. This contained 40 per cent of antiseptic, and 0.5 per cent was added. He employed fifteen different bacteria, including staphylococci, streptococci, members of the colon-typhoid bacilli, tuberculosis bacillus, and the spore-producing anthrax bacillus. Painted pipets were inserted into freshly inoculated broth. Failure of growth of the test culture was attributed to the inhibitive action of the paint. Fragments of painted pipets were placed in freshly inoculated broth culture tubes, and after incubation for 2 and 7 days, subcultures were made into sterile media. Lack of growth in subcultures was regarded as indicative of antiseptic power. Two methods were used to determine destructive power. I n the first, dried cells of Staphglococcus aureus and dried spores of Bacillus anthracis were placed in Petri dishes and then covered with paint. After variable periods of contact, aseptically obtained scrapings were transferred to liquid and solid media and observed for growth. I n the second method, paint was applied to the interior of Petri dishes and then covered with large quantities of bacterial cultures of B. anthracis or Mycobacterium tuberculosis. After contact for 8, 16, 72, and 90 days, the dried organisms were injected into laboratory animals. As judged by the onset of the respective diseases in the animals, Valle6 observed that bacilli in contact with his bactericidal paint were destroyed, whereas bacilli dried in the absence of the paint for the same time periods were not destroyed. The use of ordinary paints as controls in the destructive experiments was not reported. Valle6 arrived a t the following general conclusions: 1. Ordinary paint, in all cases, is devoid of antiseptic power. 2. Oxyquinoline sulfate added to paint in doses up t o 3 per cent was effective against streptococci and staphylococci under
some conditions and not under all conditions. The other bacteria resisted the antiseptic agent. 3. Chloramines, thymol iodide, and tribromo-&naphtholwere found ineffective. 4. An antiseptic, compounded from halogenated substances and aromatic aldehydes, prepared in the form of a colloidal jelly, and added to paint at the rate of 50 grams of jelly per kg. of paint, rendered such paint bactericidal in all cases. All of these results must be considered as inconclusive since bacteriological tests on paints after protracted drying periods were not made. The use of primarily pathogenic bacteria is significant of the fact that formulation of these bactericidal paints through incorporation of antiseptics was designed for hospital and home use. As the result of an extensive study on bactericidal properties of paint, Wetchler, Lewis, and Battline (1.2) concluded that ordinary commercial paints are only temporarily bactericidal and become entirely devoid of such property after drying for 1 to 3 weeks, whereas a paint having a 4 per cent halogenated oil as a base has a bactericidal effect on Staphylococcus aureus and Bacillus typhosus even rafter drying for 3
399
to 4 years. The addition of phenol, cresol, iodoform, copper sulfate, hexylresorcinol, methyl salicylate, phenyl chlorides, silver nitrate, and sixty other chemicals to individual paints did not render them bactericidal. Wetchler et al. employed two modifications of the techniques of Portier and Kling and of ValleB. I n one, 0.1 cc. of a saline culture suspension containing 5,000,000 Bacillus typhosus or Staphylococcus aureus per cc. was spread over 1 square inch (6.5 sq. cm.) of a painted tin disk. The disks were then placed in Petri dishes containing a layer of filter
Addition of antiseptics in practicable amounts to paints has met with little success. Exceptions are chlorophenols and oxyquinoline sulfate which offer some promise. Halogenated vehicles, such as one containing 4 per cent of chlorine, added to linseed oil give the best results obtained to date. Techniques for examination are discussed in detail and modifications suggested. At best, however, antiseptic and germicidal paints are not actually so under all practical conditions of use, and further research is needed.
paper moistened with 1 cc. of water. After 24 hours the dried bacterial culture was rinsed with 1 cc. of sterile saline, and a definite amount transplanted to a tube of broth. I n a simpler form,.painted glass rods, 10 mm. in diameter and 130-140 mm. in length, were immersed in test tubes containing 4 cc. of saline solution and 500,000,000 bacteria. After 6 hours the rods were removed, and 0.2 cc. of the culture suspension was placed on the surface of nutrient agar plates. Valentine (10) studied similar halogenated paints by essentially the methods of Wetchler et al. With B. typhosus he found that the halogenated paints, after air-drying for 9 weeks and after accelerated drying equivalent to 6 years of air-drying, showed marked but not complete bactericidal action, whereas none of ten commercial paints bought in the open market possessed any appreciable bactericidal action after air-drying for only 7 to 10 days. I n tests against Staphylococcus aureus the halogenated paints were far less bactericidal than was reported by Wetchler et al. Difference in results with these two standard bacilli coincided with differences in resistance to phenol of the respective strains. The strain used by Valentine had the properties of strains used in phenol coefficient tests, whereas Wetchler et al. used a normal throat staphylococcus strain. PRACTICAL and laboratory tests by Gardner (4) t o determine the effect of small quantities of chemicals in preventing mold growth used 1 part of chemical to 300 parts of paint. The practical test consisted in applying three coats of paint to special wooden panels. The undersides of the panels were set on wet sand and kept under shade in the open. Observations for mold growth were made after exposure for 60 days. I n their first laboratory series a liquid suspension of mold spores from agar slants was stroked on the painted panels with small clean brushes just before the paint had become dry. I n the second series a thin layer of mold medium was
400
INDUSTRIAL AND ENGINEERING CHEMISTRY
alloaTed to solidify in Petri dishes, and the surface was stroked with a water suspension of pure mold culture. A small ring of the chemically treated paint was then dropped from a spatula upon the inoculated medium, and each dish was covered. Both dishes and panels were held a t 30" C. at a relative humidity of 85 per cent. Observations were made after 7 and J2 days. These tests showed that paint in the wet stage exerted only temporary inhibition with most chemicals, and only phenyl mercury acetate and arsenic trioxide (of fifty-six different chemicals tried) prevented mold growth entirely. Studies on a modified halogenated paint by Epstein (1, 2, 3) employed agar plate methods in which painted glass rods were embedded in seeded agar just prior to solidification. H e found that after air-drying for 60 days, the halogenated paint produced marked zones of inhibition when tested against more than h-enty species of microorganisms. Kone of fourteen ordinary commercial paints, after drying for 7 to 28 days, had an inhibitory effect on any of the test organisms. Spores of Aspergillus and Penicillium molds suspended in water solutions of flour and dextrose failed to develop on halogenated painted glass surfaces tested 104 days after application. Under similar conditions the molds grew well on all of six ordinary commercial control paints after air-drying for only 13 days. Death rates of bacteria and spores of yeasts and molds as influenced by paints were determined by immersing painted glass rods, 12 X 140 mm., into culture tubes of 13.5-mm. internal diameter containing 2 cc. of saline suspensions of 1000 to 3000 cells per cc. After various hour intervals, the painted rods were remoired and 1 cc. was plated directly or after serial dilution. Based on six tests with paints airdried for 27-44 days, 100 per cent reduction of sporulating yeasts was produced by the halogenated paint, whereas control paints were without effect. Mold spores were reduced by 86 per cent within 2 hours. SIMILAR tests have been conducted by the present authors against the F. D. A. strain of Staphylococcus aureus. Based on the average of six experiments, after air-drying for 108 to 148 days, the halogenated paint effected reductions of 85.2, 99.8, and 100.0 per cent after 2, 6, and 8 hours, respectively. The maximum reductions observed with any of five control paints after the same periods of time were 28.0, 53.8, and 67.6 per cent, respectively. These results indicated that from 2750 to 4150 cells were destroyed by the action of the paint under described conditions within 8 hours. Saline suspensions of old agar-slant growths of Bacillus mesentericus with initial concentrations of 1370 to 10,000 per cc. were reduced to the extent of 49.6, 80.2, 82.9, and 96.2 per cent after 2,6, 8, and 24 hours, while maximum reductions observed with any of five control paints after the same periods of time were 11.0, 19.1, 24.0, and 24.3 per cent, respectively. To determine the germicidal action of paint films on dried bacteria, glass rods painted and air-dried for 3 to 4 months were dipped and rinsed into saline suspensions of Staphylococcus aureus and then allowed to remain under cover for 16 to 17 hours, The concentration of the suspensions varied from 3000 to 300,000,000 bacteria, based on colony counts on agar after serial dilution. The rods, 10-11 mm. in diameter and 125 mm. in length, were dipped 50 to 60 mm. below the level of the liquid bacterial suspension and then placed in Petri dish bottoms which had been coated with a corresponding paint a t about the same time that the glass rods were painted. Each dish and rod were covered loosely with a sterile Petri dish cover and allowed to remain a t room temperature for 16 to 17 hours. A t the expiration of that time, some rods were placed in 5 cc. of sterile broth contained in standard cotton-plugged culture tubes, and other rods were rinsed in
VOl. 33, No. 3
broth and removed. Both sets of tubes were incubated a t 37" C. for 48 hours. In every instance where the rods had been merely rinsed in the broth and the tubes incubated, growth of Staphylococcus aureus was observed after 24 hours. I n every case where the rod was allowed to remain in the broth, there was no growth and the broth remained clear. In repeated experiments, tubes of broth which had contained painted rods for 48 hours were each inoculated with a loopful of a 24-hour broth culture of the test organism and reincubated for 6 days. I n every instance the culture failed to develop. These experiments showed that the halogenated paint could be considered antiseptic or germicidal under one set of conditions and ineffective under another set of conditions. Ingredients of definite inhibitive character were apparently extracted from the paint. With five commercial control paints air-dried for only 41 days, growth of Staphylococcus aureus mas observed on every instance in the presence of painted glass rods. The sanitary quality of paint surfaces under actual usage was evaluated by Epstein (1) by comparing colony counts obtained by swabbing definite areas under prescribed conditions. H e reported that the numbers of bacteria recovered from surfaces of halogenated paint in a hospital and several breweries were from 75 to 98 per cent less than the numbers recovered from general-use paints. I n places of excessive moisture, i t was observed that growth of molds on generaluse paint surfaces was common 4 to 6 weeks after application, while signs of mold growth on halogenated paints were first noted 16 to 17 weeks after application.
NOKE of the different methods employed by invescigators for determining the antiseptic, germicidal, and destructive power of paints under controlled laboratory conditions are M ithout some objections. Each method was devised to predict the reaction of paints in usage toward microorganisms. Only one method (5) provided means for determining the death rate of test cultures. I n every instance but one (11), the experimental conditions involved microorganisms in the wet stage. Where organisms were deposited on painted surfaces, the numbers were tremendous and the minimum exposure time was usually 24 hours.
TABLE I. GERMICIDAL ACTTOK OF PAINTS AGAINST Staphylococcus aureus AND Escherichia coli AS DETERMIKED BY AGAR DISK METHOD Distributor Drops per
Hypodermio needla
Pen point
130
..
Staph. aureus
E. coli
Staph. aureus
E . coli
8-13
9-11
10-12
..
50
CC.
Test culture Probable dropm
Capillary pipet
KO.per
No. colonies on nutrient or eosin methylene blue agar after exposure t o paints on wood for 4 hr. __*-
Total Control (wood) Halogenated paint Paint A Paint B Paint
C
7
Staph. aureus
Total
Staph. aurew
.. ..
.. ..
56 61
15
4 5
76 74 58 81
21 38 28 41
8
58
..
32
54 68 40
38 43
7 10
26
22
46 70 47 43
13 35 33 25 35
3 6 6
51 71
29 39
6
7
12
..
..
12
3
0
12
14
21
12 8 12
Determined b y direct colony counts on nutrient agar or eosin methylene blue agar. 0
INDUSTRIAL AND ENGINEERING CHEMISTRY
March, 1941
The agar disk method of Olson and Hammer (6)for testing the germicidal property of paints was felt to offer a means of determining the actual killing effect under the closest approximation to actual conditions of use. Briefly the method employed was as follows: 1. Small droplets of saline suspensions were deposited upon the surfaces of painted and unpainted wooden panels by means of a capillary pipet, hypodermic needle, or pen point. The droplets were so small that evaporation took place within a few minutes. The concentration of organisms was varied to duplicate different practical conditions. 2. The treated panels were placed in a sterile hood. 3. After various hour intervals, about 10 cc. of properly melted and cooled 2.5 per cent a ar medium were poured over an area of contaminated surface and allowed to solidify. 4. A sterile spatula was used to transfer the agar disk thus formed into a sterile Petri dish so that the disk portion which had been in contact with the surface examined was toward the top of the dish. 5. .Incubation was according t o the character of the test orgamsm.
Results of such tests on a halogenated paint and three control paints in which saline suspensions of Staphylococcus aureus and Escherichia Coli were used are shown in I. The halogenated paint exerted some germicidal action, as based On comparisons with Obtained with the paints. The action, however, was not marked. The halo-
401
genated paint had been air-dried for 75 days, while the control paints had been air-dried for only 20 days. Summarizing the subject of antiseptic and germicida1 paints, it can be stated the halogenated-oil paint may be the best thus far developed, but i t is far from being actually antiseptic or germicidal under all practical conditions of use. While i t is a step forward, further development work will be required before a true antiseptic or germicidal paint is obtained.
Literature Cited (1) Epstein, S. S., F o o d I n d u s t r i e s , 9, 386-7, 419-20 (1937). (2) Ibid., 9, 513, 541-3 (1937). (3) EPstein, 8. 8.9 Am. Brewer, 70,41-63 (1937). (4) Gardner, H.A., “Physical and Chemical Examination of Paints, Varnishes, Lacquers and Colors”, 6th ed., pp. 445-52, Washington, Inst. of Paint and Varnish Research, 1933. ( 5 ) Hart, J. Thoracic Surgery, 6, 45-81 (1936). (6) Olson, H. c., and Hammer, B. w.9 A@. Exp. Sta. Iowa College, Bull. 300 (1933). (7) Portier, P., and Kling, A., Bull. m a d . mdd., 106, 305-9 (1931); Peintures, pigments, vernis, 9, 14-16 (1932). (8) Robb, H., Am. J. Obstetrics, 40, 451 (1909). (9) Troussaint, M., Bull. m a d . mdd., 109, 448-52 (1933). (10) Valentine, E., Proc. SOC.Exptl. Bid. M e d . , 34, 166-70 (1936). (11) Valleb, M. J., Chintie et industrie, 31, numero special, 962-4 (1934). (12) Wetchler, S., Lewis, A., and Battline, F., P a i n t V a r n i s h Production Mgr., 14, 12-18 (1936).
*.,
PR~SENTED before the Division of Paint and Varnish Chemistry at the 100th Meeting of the American Chemical society, Detroit, Mich.
Line Coordinate Chart for Vapor Pressures of Organic Solvents D. S . DAVIS, Wayne University, Detroit, IlIich.
I
N CONNECTION with his study of evaporative index, Gardner [IND.ENG.CHEM.,32, 226 (1940)l presented
vapor pressure-temperature data for eight organic solvents. From the actual coordinates of the plotted points given in a private communication, it is possible to show that the relationship between p , the vapor pressure in mm. of mercury, and t , the centigrade temperature, is given by logp =
A t + 253 +
where A and B are constants specific to the compound. The linearity of log p and l/(t 253) enables construction of the accompanying line coordinate chart which presents the data in a compact and convenient form. The broken line indicates that the vapor pressure of n-butyl acetate is 41 mm. a t 50” C. The nature of the agreement between values read from the chart and the actual data is shown in the table which includes values of A and B for each material.
+
T Compound n-Amyl acetate n-Butyl acetate
Diacetone alcohol Diisoamylene
Diisobutylene Ethylene glycol monoethyl ether Isodecane
Isooctane
~
~ Pressure, ~ , , Mm. Hg Data Chart 56.9 23.9 24.0 106 6 204 200 49.2 43.2 39 7 93.5 274 270 17.8 17.6 66.5 139.3 324 324 58.0 21.6 23 0 110.7 188 188 35.8 72.3 72.0 72.0 303 304 49.9 23.2 23.0 94.0 180 180 59.8 24.5 25.0 120.3 254 258 64.0 31.0 65.1 63.5 243 240
C.
B
1000 A -2.077
8.078
-1.964
8.099
-2.188
8.091
-1.953
7.641
-1.623
7.474
-2.139
8.416
-1.948
7.620
-1.592
7.413
2 4
n-Amyl Acetate n-duty/ Acstafc Di-accfone Akoho/ D i - /soamy/cne
7
/so-octme
I
z
..
ti
