JP3056881B2 - Virucidal agents and their use - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

FIELD OF THE INVENTION The present invention relates to a virucidal agent for IBDV and its use.

[0002]

BACKGROUND OF THE INVENTION Chicken infectious bursal disease is a viral infectious disease that first occurred in a poultry farm in the southeastern United States in 1957 and has the property of causing immunosuppression. Strong IBDV outbreak is one in America, Europe to Asia and Japan
It has happened since 990. This IBDV has a strong viral resistance, and it is difficult to expect a virucidal effect with currently marketed disinfectants and their known uses.
It is said that surfactants generally used widely (so-called inverted soaps and amphoteric soaps) have no effect on disinfecting and inactivating IBDV, and this has become common sense to some extent. I was It has been reported that the quaternary ammonium salt “dialkyldimethylammonium chloride” which is an inverse soap actually has no virucidal effect on IBDV.

[0003]

The present invention relates to a novel virucidal agent having excellent virucidal effect on IBDV and its use.

[0004]

The first aspect of the present invention is a general formula:

Embedded image Characterized in that it contains a dialkyldimethylammonium chloride represented by the formula (wherein R ¹ and R ² are both alkyl groups and may be the same or different), and an alkali agent. Disease virus (hereinafter I
BDV). It said C ₈ or higher alkyl group in a chloride dialkyldimethylammonium, particularly those of C ₈ -C ₁₂ are preferred. Specific compounds include, for example, dioctyldimethylammonium chloride, octyldecyldimethylammonium chloride, didecyldimethylammonium chloride, and among them, didecyldimethylammonium chloride is particularly preferred.

[0005]

【Example】

<Bacterial strain> PV1 strain: IBDV isolated from chickens of IBD that occurred in Italy and used for 62 generations in CAM and 23 generations in CK was used. This strain belongs to type IBDV1. <Hens and embryonated eggs> PDL maintained in an isolated chicken house
A 4-8 week old chick of line -1 was used for CK. C
A 10-day-old embryonated chicken egg of the PDL-1 line was used for AM. <CK culture method> To the growth medium of CK, a tryptose phosphate medium (TPB) at a ratio of 0.3% was added to the Eagle MEM medium “Nissui”, and further, antibiotics (penicillin 100 U / ml, kanamycin 100 μg / ml,
And fungizone (0.5 μg / ml), 0.15% sodium bicarbonate and 5% calf serum. For maintenance medium, Eagle MEM medium “Nissui”
To which an antibiotic and 0.15% sodium bicarbonate were added. The digestion solution was prepared by dissolving trypsin (Difco, 1: 250) in a citrate buffer at a ratio of 0.05%. For culture, kidneys of 4-8 week old SPF chickens were aseptically removed, digested, cells were thickened by centrifugation, and their volumes were measured and suspended at a rate of 0.25-0.3%. Each 5 ml of the cell suspension was dispensed into a petri dish (55 mm in diameter) and cultured in a 37 ° C. carbon dioxide egg containing 5% carbon dioxide and humidity. Two days after the start of the culture, the cells were used for virus inoculation and experiments. <Viral passage> The passage of the virus strain was performed using CK prepared as described above, and LE (0.5% lactalbumin hydrolyzate, 2 μg / ml amphotericin B, 200 U / m
1 ml of penicillin and 200 μg / ml of streptomycin in Earl's buffer solution), and inoculate a 0.4 ml petri dish with a virus solution diluted 10-fold, place in a carbon dioxide egg container for 1 hour, and withdraw the inoculated virus solution. 5 ml of a maintenance medium was added and cultured for 4 days. Cytopathic effect (C
The cells in which PE) appeared were collected together with the culture solution, frozen at −40 ° C., thawed, and sonicated. The same operation was repeated for 3 to
The virus was passaged four times. <Virus titer measurement> The virus titer was measured by the black method using the prepared CK. CK was inoculated with the virus solution serially diluted 10-fold with LE in the same manner as the virus passage. After removing the inoculated virus solution, 5 ml of Eagle's MEM medium containing 0.8% agar (Bacto agar), 0.3% TPB, antibiotics and 5% calf serum prepared at about 50 ° C. was overlaid. Cultured for days. On the third day of culture, at about 50 ° C.
Eagle ME containing 0.8% agar, 0.3% TPB, antibiotics and 1% neutral red prepared in
The M medium was overlaid with 5 ml, and the number of blacks was counted the next day. Black forming units (PFU) are given in log / ml.

<Implementation method> In Experiment 1, the equation

Embedded image The diluted solution of didecyldimethylammonium chloride and the virus solution were contacted at a ratio of 1: 1 by 0.4 ml each.
Contact was made at a ratio of 1: 1 for each ml. The concentrations of didecylmethylammonium chloride were three concentrations as shown in the table below, sodium hydroxide was used as an alkaline agent, and the concentration was 0.05%. The effects on IBDV are as follows.

[Table 1] * Residual virus titers were expressed as log ₁₀ PFU / ml. For ** virucidal index, × in residual virus titer 3.4 log ₁₀ or more of (no effect) △ those (almost no effect) of 2.4 to 3.4 log ₁₀ ○ is 1.4 2.4 log ₁₀ (having a ゝ effect) ◎ is 1.4 log ₁₀ or less (having a great effect). In order to confirm the effect of using an alkali agent,
A similar test was conducted with various concentrations of sodium hydroxide. The results are shown in the following table.

[Table 2]

<Comparative Example> In the same manner as in the above-mentioned Example, instead of didecyldimethylammonium chloride, a formula

Embedded image (R ³ is a C number of 9 to 15) [mono and bis (trimethylammonium chloride methylene)]-alkyl (C _{9 to} C ₁₅ ) toluene represented by the following formula is shown in the following table.

[Table 3]

[0008]

According to the present invention, a novel virucidal agent having an excellent bactericidal effect can be provided by using two components in combination, although the bactericidal effect is not obtained by itself.

──────────────────────────────────────────────────続 き Continuation of the front page (51) Int.Cl. ⁷ identification code FI A61P 31/12 171 A61P 31/12 171 (58) Investigated field (Int.Cl. ⁷ , DB name) A61K 33/00 A01N 33 / 12 A01N 59/00 A61K 31/14 A61L 2/18 A61P 31/12 CAPLUS (STN) REGISTRY (STN)

Claims (3)

(57) [Claims] 1. A compound of the general formula Characterized by containing a dialkyldimethylammonium chloride represented by the formula (wherein R ¹ and R ² are both alkyl groups and may be the same or different) and an alkali agent, Virus (hereinafter IBD)
V). 2. The virucidal agent according to claim 1, wherein the dialkyldimethylammonium chloride is didecyldimethylammonium chloride. 3. A method of disinfecting a barn, which comprises applying a solution containing a dialkyldimethylammonium chloride and an alkali agent to the barn.