The present invention provides a novel purine 4'-thioarabinonucleoside represented by the following formula 1!: ##STR1## wherein B represents a purine base other than adenine. Also disclosed are a method for preparing the purine 4'-thioarabinonucleoside and pharmaceutical compositions containing the purine 4'-thioarabinonucleoside.

BACKGROUND OF THE INVENTION 
1. Field of the Invention 
The present invention relates to purine 4'-thioarabinonucleosides. 
2. Related Art 
The only purine 4'-thioarabinonucleoside reported heretofore is 
9-(4-thio-.beta.-D-arabinofuranosyl)adenine disclosed in J. Org. Chem., 
33(1), pp. 189-192, 1968. However, that journal does not describe the 
biological activities of this compound. 
Thus, it can be seen that purine 4'-thioarabino -nucleosides have scarcely 
been studied thus far. Therefore, it was considered that purine 
4'-thioarabinonucleosides having biological activities superior to those 
possessed by previously known 4'-thioarabinonucleoside might be 
discovered. 
SUMMARY OF THE INVENTION 
Under the aforementioned circumstances, we conducted careful research to 
achieve the above object and found that a variety of purine 
4'-thioarabinonucleosides can be easily obtained in a reduced number of 
reaction steps by the use of the novel synthesis method we developed, and 
that the resultant compounds have antiviral activities. The present 
invention was achieved based on these findings. 
Accordingly, an object of the present invention is to provide a novel class 
of purine 4'-thioarabinonucleosides and a method for synthesizing these 
compounds. 
In one aspect of the present invention, there is provided purine 
4'-thioarabinonucleoside represented by the following formula I!: 
##STR2## 
wherein B represents a purine base other than adenine. 
In another aspect of the present invention, there is provided a method for 
preparing purine 4'-thioarabinonucleoside of formula I! comprising steps 
1 through 4 described below. 
Step 1: 
In step 1, a sulfonyl group is introduced to each of the 2- and 5- 
positions of a compound of formula II!, after which the compound is 
reacted with a sulfide to obtain a compound represented by formula III!: 
##STR3## 
wherein R.sub.1 represents an alkyl group and R.sub.2 represents a 
protective group for a hydroxyl group. 
Step 2: 
The furanose ring of the compound represented by formula III! is 
hydrolyzed and then reduced to obtain a compound represented by formula 
IV!: 
##STR4## 
wherein R.sub.1 and R.sub.2 have the same meanings as defined above. Step 
3: 
The compound of formula IV!, while the hydroxyl groups at the 2- and 5- 
positions of the compound are protected, is reacted with an oxidizing 
agent to form a sulfoxide. The sulfoxide is converted into a compound of 
formula V! through Pummerer rearrangement: 
##STR5## 
wherein Ac represents an acetyl group and each of R.sub.2 and R.sub.3 
represents a protective group for a hydroxyl group. 
Step 4: 
The compound of formula V! is subjected to a glycosylation reaction so as 
to introduce a purine base represented by B to the 1- position of the 
saccharide moiety, after which the protective groups for the hydroxyl 
groups in the saccharide moiety are eliminated to obtain a compound of 
formula II!: 
##STR6## 
wherein Ac, R.sub.2, R.sub.3, and B have the same meanings as defined 
above. 
In still another aspect of the present invention, there is provided a 
pharmaceutical composition comprising purine 4'-thioarabinonucleoside 
represented by the above-described formula I! as an active ingredient. 
The other objects, features, and advantages of the present invention will 
become apparent from the following description.

DESCRIPTION OF PREFERRED EMBODIMENTS 
The present invention will next be described in detail. 
(1) Compounds 
The compounds of the present invention are represented by formula I! 
described above. Purine bases represented by B in the formula encompass, 
in addition to well-known bases of nucleic acid such as guanine and 
hypoxanthine excepting adenine, azapurine derivatives (8-azapurine, 
2-azapurine, etc.) and deazapurine derivatives (3-deazapurine, 
7-deazapurine, etc.). B may have one or a plurality of substituents (such 
as lower (C1-C5) alkyl, halogen, amino, alkoxy, etc.) as a result of 
introduction of these substituents to one or more arbitrary positions of 
the above-mentioned bases including adenine. Specific examples of bases 
having such substituents include, but are not limited to, 2-aminopurine, 
2,6-diaminopurine, 6-chloropurine, 6-chloro-2-aminopurine, 
6-methoxypurine, 6-methoxy-2-aminopurine, and 
6-cyclopropylmethylamino-2-aminopurine. 
The compounds of the present invention may take the forms of salts, 
hydrates, or solvates. Examples of the salts include acid addition salts 
formed in combination with inorganic acids (hydrochloric acid, sulfuric 
acid, phosphoric acid, etc.) or organic acids (fumaric acid, tartaric 
acid, succinic acid, etc.). 
The hydrates and solvates may be those in which 0.1-3.0 molecules of water 
or a solvent is added to 1 molecule of the compound of the present 
invention or a salt thereof. Also, the compounds of the present invention 
encompass a variety of isomers such as .alpha.-anomers, .beta.-anomers, 
and tautomers. 
Particularly preferred are 9-glycosylated compounds of the .beta.-anomer 
type. Specific examples of preferred compounds of the present invention 
include the following: 
9-(4-thio-.beta.-D-arabinofuranosyl)guanine 
9-(4-thio-.beta.-D-arabinofuranosyl)hypoxanthine 
9-(4-thio-.beta.-D-arabinofuranosyl)-2-aminopurine 
9-(4-thio-.beta.-D-arabinofuranosyl)-2,6-diaminopurine 
9-(4-thio-.beta.-D-arabinofuranosyl)-6-chloropurine 
9-(4-thio-.beta.-D-arabinofuranosyl)-6-chloro-2-aminopurine, 
and 
9-(4-thio-.beta.-D-arabinofuranosyl)-6-methoxy-2-aminopurine. 
(2) Process of Manufacture 
The compounds of the present invention are synthesized through the 
following 4 steps. 
Step 1: 
In Step 1 of the method of the present invention, a sulfonyl group is 
introduced to each of the 2- and 5- positions of a compound of formula 
II!, after which the compound is reacted with a sulfide to obtain a 
compound represented by formula III!: 
##STR7## 
wherein R.sub.1 represents an alkyl group and R.sub.2 represents a 
protective group for a hydroxyl group. 
The starting material used in the method of the present invention is a 
xylose derivative (hereinafter may be referred to as the starting 
compound) represented by formula II!. 
Examples of the alkyl group represented by R.sub.1 include C1-C3 lower 
alkyl groups such as methyl and ethyl, and substituted or unsubstituted 
benzyl groups such as benzyl and methoxy benzyl. 
The protective group for the hydroxyl group represented by R.sub.2 is not 
particularly limited so long as it is selected from those which are 
generally used. Specific examples of the protective group include alkyl 
groups, silyl groups, and acyl groups. More specifically, alkyl groups 
which may be used for the purpose of protection include those listed for 
R.sub.1. Examples of silyl groups include t-butyldimethylsilyl, 
t-butyldiphenylsilyl, etc., and examples of acyl groups include acetyl, 
benzoyl, pivaloyl, etc. 
The starting compound of the method of the present invention may be 
prepared using a well-known method such as the one described in 
Tetrahedron, 37, pp. 2379-2382 (1981), the content of which is 
incorporated herein by reference. 
Examples of the sulfonyl group which is introduced into the hydroxyl groups 
at the 2- and 5- positions of the compound of formula II! include mesyl 
and tosyl. 
A mesylation reaction and tosylation reaction may be performed using 
conventional methods. For example, mesylation reaction may be performed as 
follows. One mole of a starting compound is reacted with 2-10 mols, 
preferably 2-4 mols, of mesyl halide (e.g., mesyl chloride) at 
0-100.degree. C. for 0.5-5 hours while stirring in the presence of a base 
such as triethylamine in an organic solvent such as methylene chloride, 
acetonitrile, dimethylformamide, or pyridine (when pyridine is used as the 
organic solvent, a base such as triethylamine is not necessarily used). 
The reaction is preferably performed in an atmosphere of an inert gas such 
as argon or nitrogen. 
Subsequently, the thus-obtained compound is reacted with a sulfide to 
afford a compound of formula III!. 
The sulfide used in this reaction is not particularly limited as long as it 
is a metal sulfide (preferably, alkali metal sulfide) such as sodium 
sulfide, potassium sulfide, etc. 
The reaction may be performed by reacting 1 mol of a starting compound with 
1-20 mols of a sulfide at a temperature between room temperature and 
150.degree. C. for 0.5-5hours while stirring in an organic solvent such as 
dimethylformamide, dimethylsulfoxide, etc. When necessary, the reaction 
may be performed in an atmosphere of an inert gas such as argon or 
nitrogen. 
The thus-produced compound of formula III! may be separated and purified 
using conventional means for the separation and purification of protected 
saccharides. For example, the mixture may be partitioned using ethyl 
acetate and water, after which silica gel column chromatography may be 
performed using an organic solvent mixture for elution such as 
n-hexane-ethyl acetate, thereby separating and purifying the formula III! 
compound. 
Step 2: 
In Step 2 of the method of the present invention, the furanose ring of the 
compound represented by formula III! is hydrolyzed and then reduced to 
obtain a compound represented by formula IV!: 
##STR8## 
wherein R.sub.1 and R.sub.2 have the same meanings as defined above. 
The method of hydrolysis is not particularly limited so long as the 
furanose ring of the compound of formula III! can be hydrolyzed by the 
method. Methods using acid catalysts are particularly preferred. 
Examples of acid catalysts include inorganic acids such as hydrochloric 
acid, sulfuric acid, etc. and organic acids such as acetic acid and 
trifluoroacetic acid. 
The hydrolysis reaction may be performed in a water-soluble ether-derived 
solvent such as tetrahydrofuran, dioxane, etc. in the presence of any one 
of the above-mentioned acid catalysts between room temperature and 
100.degree. C. for 0.5-5 hours while stirring. 
When the thus-obtained compound is subjected to a reduction reaction, a 
compound of formula IV! is obtained. 
Examples of reducing agents include tetrahydroborates such as sodium 
tetrahydroborate (sodium borohydride), potassium tetrahydroborate, etc. 
The reduction reaction may be performed by reacting 1 mol of a compound of 
formula III! with 0.2-10 mols of a reducing agent in an alcoholic solvent 
such as methanol at a temperature between -80 and 100.degree. C. for 0.5-3 
hours while stirring. 
The thus-obtained formula IV! compound may be separated and purified using 
conventional means for the separation and purification of protected 
saccharides. For example, neutralization of the reaction mixture at the 
completion of reaction, evaporation of the organic solvent, extraction 
using chloroform, and silica gel column chromatography may serially be 
performed so as to obtain the target compound as a separated and purified 
product. 
Step 3: 
In Step 3 of the method of the present invention, the compound of formula 
IV! is reacted with an oxidizing agent while protecting the hydroxyl 
groups at the 2- and 5- positions of the compound to form a sulfoxide. 
Subsequently, the sulfoxide is converted into a compound of formula V! 
through Pummerer rearrangement. 
##STR9## 
wherein Ac represents an acetyl group and each of R.sub.2 and R.sub.3 
represents a protective group for a hydroxyl group. 
Examples of the protective groups represented by R.sub.3 and introduced to 
the 2- and 5- positions of the compound of formula IV! include lower 
alkyl groups such as methyl and ethyl; substituted or unsubstituted benzyl 
groups such as benzyl and dimethoxybenzyl; silyl groups such as 
t-butyldimethylsilyl and t-butyldiphenylsilyl; and acyl groups such as 
acetyl, benzoyl, and pivaloyl. 
Protective groups may be introduced by routine methods. For example, 
protective groups may be introduced by reacting 1 mol of a compound of 
formula IV! with 2-10 mols, preferably 3-8 mols, of an alkylating agent 
such as benzyl chloride, benzyl bromide, or p-methoxybenzyl chloride in a 
single organic solvent such as dimethylformamide, dimethylsulfoxide, etc. 
or in a solvent mixture such as tetrahydrofuran-dimethylsulfoxide in the 
presence of a base such as sodium hydride in an atmosphere of an inert gas 
such as argon, nitrogen, etc. at 0-50.degree. C. overnight while stirring. 
Examples of the oxidizing agent used in the oxidizing reaction include 
m-chloroperbenzoic acid and sodium metaperiodate. 
The oxidizing reaction may be performed by reacting 1 mol of a compound of 
formula IV! in which the hydroxyl groups at the 2- and 5- positions are 
protected with 0.2-5 mols of an oxidizing agent (such as 
m-chloroperbenzoic acid, sodium metaperiodate, etc.) in an organic solvent 
such as methylene chloride or alcohol (e.g., methanol) in a stream of an 
inert gas such as argon or nitrogen, if necessary, at a temperature 
between -100 and 0.degree. C. for 10 minutes to 2 hours. 
When the thus-obtained sulfoxide is subjected to a Pummerer rearrangement 
reaction, a compound of formula V! is obtained. 
The Pummerer rearrangement reaction may be performed by a conventional 
method. For example, the sulfoxide is stirred for 1-5 hours between 
60.degree. C. and the refluxing temperature in an acid anhydride such as 
acetic anhydride. 
The thus-obtained formula V! compound may be separated and purified using 
conventional separation and purification techniques. For example, 
neutralization, evaporation of the organic solvent, extraction from the 
aqueous layer using chloroform, and silica gel column chromatography may 
sequentially be performed so as to obtain the formula V! compound as a 
separated and purified product. 
If a purification step is required to be performed before oxidation 
reaction, the mixture may be partitioned using ethyl acetate and water, 
after which silica gel column chromatography may be performed using an 
organic solvent mixture for elution such as n-hexane-ethyl acetate, 
thereby separating and purifying the formula V! compound. 
Step 4: 
In Step 4 of the method of the present invention, the compound of formula 
V! is subjected to a glycosylation reaction so as to introduce a base 
represented by B to the 1- position of the saccharide moiety, after which 
the protective groups for the hydroxyl groups in the saccharide moiety are 
eliminated to obtain a compound of formula I!. 
##STR10## 
wherein Ac, R.sub.2, R.sub.3, and B have the same meanings as defined 
hereinbefore. 
Specific examples of Lewis acids used in the glycosylation reaction 
include, but are not limited to, trimethylsilyl trifluoromethane 
sulfonate, tin tetrachloride, titanium tetrachloride, zinc chloride, zinc 
iodide, and boron trifluoride. 
The glycosylation reaction may be performed by reacting 1 mol of a compound 
of formula IV! with 1-10 mols of a base of nucleic acid and 0.1-10 mols 
of any one of the aforementioned Lewis acids in an organic solvent such as 
methylene chloride, chloroform, dichloroethane, acetonitrile, or 
dimethylformamide in a stream of an inert gas such as argon or nitrogen at 
a temperature between -50 and 100.degree. C. for 1-3 hours. If a silylated 
base of nucleic acid is used, 7-glycosylated compounds can be synthesized 
with priority. 
Subsequently, when the protective group for the hydroxyl group in the 
saccharide moiety is eliminated, a compound of formula I! is obtained. 
The elimination of the groups protecting the hydroxyl groups may be 
suitably performed by hydrolysis, catalytic hydrogenation, or any other 
conventional process in accordance with the protective groups used. For 
example, when the protective groups are benzyl groups or benzyl-derived 
groups, they are eliminated through reaction with boron trichloride for 
between 10 minutes and 6 hours at a temperature between -100 and 
50.degree. C. in methylene chloride in a stream of an inert gas such as 
argon or nitrogen. 
The thus-obtained compound I! may be separated and purified by a suitable 
combination of conventional separation and purification methods 
(recrystallization, a variety of column chromatography procedures, etc.) 
for nucleosides. 
(3) Use 
Since the compounds of the present invention exhibit excellent antiviral 
activities, pharmaceutical compositions containing the compounds as active 
ingredients are useful for the prevention or the treatment of subjects who 
have been infected with a virus or who run the risk of infection with a 
virus. 
Examples of target viruses include herpes simplex virus type 1 (hereinafter 
referred to as HSV-1), herpes simplex virus type 2 (hereinafter referred 
to as HSV-2), human cytomegalovirus (hereinafter referred to as HCMV), and 
varicella zoster virus (hereinafter referred to as VZV), all of which 
belong to the herpes virus family. 
The dosage of the compound of formula I!, an active ingredient of the 
pharmaceutical composition of the present invention, varies depending on 
the patient's age and body weight, identity of disease, severity of 
disease, tolerance to the drug, manner of administration, etc. Therefore, 
the dose is determined considering these factors as a whole so as to be 
suited to the patient. Generally, the dose is between 0.001 and 1,000 
mg/kg body weight, and preferably between 0.1 and 100 mg/kg body weight, 
per day, and is administered at a single treatment or in plural doses. 
The manner of administration is not limited, and may be peroral, 
parenteral, enteral, or topical administration. 
When pharmaceutical compositions containing the compound of the present 
invention are formulated, it is a general practice to incorporate 
ordinarily employed carriers, vehicles, and other additives. Carriers may 
be either solid or liquid. Examples of solid carriers include lactose, 
kaolin, sucrose, crystalline cellulose, cornstarch, talc, agar, pectin, 
stearic acid, magnesium stearate, lecithin, and sodium chloride; and 
examples of liquid carriers include glycerol, peanut oil, polyvinyl 
pyrrolidone, olive oil, ethanol, benzyl alcohol, propylene glycol, and 
water. 
The compositions may take arbitrary forms. For example, if a solid carrier 
is used, tablets, powders, granules, capsules, suppositories, lozenges, 
etc. may be formed, and if a liquid carrier is used, syrups, emulsions, 
soft gelatin capsules, creams, gels, pastes, sprays, injections, etc. may 
be formed. 
The compounds of the present invention are expected to be developed and 
used as medicinal agents due to their remarkable antiviral activities. 
Moreover, the method of the present invention is particularly useful for 
the manufacture of purine 4'-thioarabinonucleoside because firstly it 
employs an inexpensive substance as a starting material, secondly it 
requires a reduced number of steps, and thirdly its procedure is simple 
and easy. 
EXAMPLES: 
The present invention will next be described by way of example. However, 
the invention should not be construed as being limited by any of the 
examples. 
Example 1: 
Synthesis of 
Ia-.alpha.!9-(4-thio-.alpha.-D-arabinofuranosyl)-2,6-diaminopurine and 
Ia-.beta.!9-(4-thio-.beta.-D-arabinofuranosyl)-2,6-diaminopurine (in 
formula I!, B=2,6-diaminopurine): 
1) Synthesis of 
2,5-anhydro-3-O-benzyl-1-O-methyl-2-thio-.beta.-D-arabinofuranose (formula 
III!, R.sub.1 =Me, R.sub.2 =Bn) 
While cooling on ice, methanesulfonyl chloride (6.33 ml) was added to 80 ml 
of pyridine in which 3-O-benzyl-1-O-methyl-.beta.-D-xylofuranose (6.93 g, 
formula II!, R.sub.1 =Me, R.sub.2 =Bn) had been dissolved. The mixture 
was stirred for 1 hour at room temperature under a flow of argon. Reaction 
was stopped by adding ice-water, after which the solvent was evaporated. 
The residue was partitioned using ethyl acetate-water, and the organic 
layer was dried. The solvent was evaporated, and the residue was dissolved 
in dimethylformamide (DMF, 100 ml). Sodium sulfide (9.84 g) was added, and 
the mixture was stirred for 1 hour at 100.degree. C. under a flow of 
argon. The solvent was evaporated, and the residue was partitioned using 
ethyl acetate-water. The organic layer was washed using water and then 
dried. The solvent was evaporated and the residue was purified by silica 
gel column chromatography. The fraction eluted with 5-10% ethyl 
acetate-n-hexane was collected and concentrated to obtain 5.05 g of the 
target compound (yield 73%). 
.sup.1 H--NMR (CDCl.sub.3) .delta. 7.36-7.29 (5H, m), 4.89 (1H, s), 4.62 
(1H, d, J=11.7 Hz), 4.52-4.48 (2 H, m), 4.37-4.36 (1H, m), 3.34 (4H, s), 
3. 04 (1H, dd, J=10.3, 2.0 Hz), 2.77 (1H, dd, J=10.3, 1.5 Hz) 
2) Synthesis of 2,5-anhydro-3-O-benzyl-1-O-methyl-2- 
thio-.alpha.-D-arabinofuranose (formula III!, R.sub.1 =Me, R.sub.2 =Bn) 
The procedure of 1) was repeated using 
3-O-benzyl-1-O-methyl-.alpha.-D-xylofuranose (6.13 g, formula II!, 
R.sub.1 =Me, R.sub.2 =Bn) instead of 
3-O-benzyl-1-O-methyl-.beta.-D-xylofuranose , thereby obtaining 4.75 g of 
the target compound (yield 42%). 
.sup.1 H--NMR (CDCl.sub.3) .delta.7.39-7.30 (5H, m), 5.13 (1H, d, J=2.4 
Hz), 4.66 (1H, d, J=11.7 Hz), 4.5 3 (1H, d), 4.36-4.35 (1H, brm), 4.29 
(1H, t, J=2.4 Hz), 3.51 (1H, t, J=2.4 Hz), 3.47 (3H, s), 3.04 (1H, dd, 
J=10.5, 2.2 Hz), 2.95 (1H, dd, J=10.5, 1.2 Hz) 
3) Synthesis of 3-O-benzyl-1-deoxy-4-thio-D- arabinofuranose (formula IV ! 
, R.sub.2 =Bn) 
2,5-Anhydro-3-O-benzyl-1-O-methyl-2-thio-D- arabinofuranose (9.50 g, 
.alpha.:.beta.=1:1) was dissolved in tetrahydrofuran (THF, 200 ml). To the 
solution was added 4N--HCl (100 ml), and the mixture was stirred for 1 
hour at room temperature. The reaction mixture was neutralized using solid 
sodium hydrogencarbonate. Insoluble matter was removed by filtration, 
after which THF was evaporated under reduced pressure. Extraction was 
performed three times using chloroform, and the organic layer was dried. 
The solvent was evaporated, and the residue was dissolved in methanol (150 
ml). While cooling on ice, a methanol solution containing 1.43 g of sodium 
borohydride was added dropwise. After completion of addition, the mixture 
was stirred for 45 minutes while being cooled on ice. The reaction mixture 
was neutralized using acetic acid, after which the solvent was evaporated 
and the residue was partitioned using chloroform-water. The aqueous layer 
was extracted twice using chloroform, and the organic layer was dried. The 
solvent was evaporated, and the residue was subjected to silica gel column 
chromatography. The fraction eluted with 33-50% ethyl acetate-n-hexane was 
collected and concentrated to obtain 8.18 g of 3-O-benzyl-1-deoxy 
-4-thio-D-arabinofuranose (yield: 90%). 
.sup.1 H--NMR (CDCl.sub.3 --D.sub.2 O) .delta.7.38-7.27 (5H, m) , 4.6 4 
(2H, s) , 4.38 (1H, dt, J=2.9, 4.4 Hz), 3.96 (1H, t, J=2.9 Hz), 3.78 (1H, 
dd, J=2.9, 11.7 Hz), 3.66 (1H, dd, J=3.9, 11.7 Hz), 3.60 (1H, dt, J=2.9, 
3.9 Hz), 3.21 (1H, dd, J=4.4, 11. 2 Hz), 2.90 (1H, dd, J=2.9, 11.2 Hz) 
4) Synthesis of 1-O-acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose 
(formula V!, R.sub.2 =R.sub.3 =Bn) 
3-O-Benzyl-1-deoxy-4-thio-D-arabinofuranose (5.0 g, 20.8 mmol) was 
dissolved in dimethylformamide (100 ml). To the solution was added 60% 
sodium hydride (4.16 g, 104 mmol) under a flow of argon, and the mixture 
was stirred for 1 hour at 0.degree. C. After the one hour of stirring, 
benzyl chloride (16.8 ml, 146 mmol) in dimethylformamide (52 ml) was added 
dropwise. The resultant mixture was stirred overnight at room temperature 
and subsequently poured into ice-water so as to stop the reaction. 
The mixture was partitioned using ethyl acetate. The organic layer was 
washed with saturated brine and then dried over sodium sulfate. The 
solution was concentrated and purified by silica gel column chromatography 
(AcOEt: Hex =1:6), thereby obtaining 5.54 g of 1-O-deoxy-2,3,5-tri-O 
-benzyl-4-thio-D-arabinofuranose (yield: 63.3%). 
Elementary analysis: for C.sub.26 H.sub.28 O.sub.3 S 
Calculated C: 74.25, H: 6.71 
Found C: 74.28, H: 6.82 
.sup.1 H-NMR (CDCl.sub.3) .delta.7.35-7.25 (15H, m) , 4.90 (1H m), 
4.72-4.45 (6H, m), 4.11 (1H, m), 3. 69 (1H, dd, J=7.3, 8.8 Hz), 3.56 (1H, 
ddd, J=3.4, 6.4, 7.3 Hz), 3.50 (1H, dd, J=6.4, 8.8 H z), 3.08 (1H, dd, 
J=4.9, 11.2 Hz), 2.90 (1H, dd, J=4.4, 11.2 Hz) 
The resultant tribenzyl derivative (2.88 g, 6.85 mmol) was dissolved in 
distilled methylene chloride (40 ml). To the solution was added dropwise 
80% m-chloroperbenzoic acid (1.48 g, 6.85 mmol) dissolved in distilled 
methylene chloride (40 ml) while maintaining the temperature at 
-78.degree. C. under a flow of argon. The mixture was stirred for 30 
minutes, and then the reaction was stopped using a saturated aqueous 
sodium hydrogen carbonate solution. 
Subsequently, the mixture was extracted with methylene chloride, and the 
organic layer was washed once with a 10% sodium thiosulfate solution, 
twice with a saturated aqueous sodium hydrogen carbonate solution, and 
then once with saturated brine, followed by drying over sodium sulfate. 
The solution was concentrated to quantitatively obtain a sulfoxide. 
To the resultant sulfoxide (6.85 mmol) was added acetic anhydride (34.2 
ml), and the mixture was heated while stirring for 3 hours at 100.degree. 
C. The mixture was allowed to cool, brought to dryness under reduced 
pressure, and purified by silica gel column chromatography 
(AcOEt:Hex=1:10), thereby obtaining 1.79 of 1-O-acetyl-2,3,5-tri-O 
-benzyl-4-thio-D-arabinofuranose (yield: 56.5%). 
Elementary analysis: for C.sub.28 H.sub.30 O.sub.4 S.0.75H.sub.2 O 
Calculated C: 70.63, H: 6.67 
Found C: 70.37, H: 6.24 
.sup.1 H--NMR (CDCl.sub.3) .delta. 7.35-7.24 (15H, m), 6.07 (1H, d, J=3.9 
Hz), 5.98 (1H, d, J=2.9 Hz), 4. 83-4.48 (6H, m), 4.26 (1H, dd, J=2.9, 
4.9Hz), 4.18 (1 H, dd, J=3.9, 8.8 Hz), 4.12 (1 H, dd, J =6.8, 8.8 Hz), 
4.03 (1H, dd, J=4.9, 6.4 Hz), 3.76 (1H, m), 3.73-3.44 (2H, m), 3.40 (1H, 
m), 2.04 (3H, s) 
5) Synthesis of Ia-.alpha.!9-(4-thio-.alpha.-D-arabinofuranosyl) 
-2,6-diaminopurine and Ia.beta.!9-(4-thio-.beta.-D-arabinofuranosyl) 
-2,6-diaminopurine (in formula I!, B=2,6-diaminopurine) 
1-O-Acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose (800 mg, 1.67 mmol) 
was dissolved in distilled acetonitrile (7 ml). 2,6-Diaminopurine (452 mg) 
and molecular sieve 4A (897 mg) were added thereto. To the resultant 
mixture was added dropwise trimethylsilyl triflate (0.75 ml) at room 
temperature, and the mixture was stirred for 1 hour. Subsequently, a 
saturated aqueous sodium hydrogen carbonate solution was added, followed 
by stirring for 30 minutes to stop the reaction. The mixture was extracted 
with methylene chloride, and the organic layer was washed with a saturated 
aqueous sodium hydrogen carbonate solution, followed by drying over sodium 
sulfate. The residue was concentrated and purified by silica gel column 
chromatography (5% MeOH in CHCl.sub.3). The resultant purified product 
(378 mg, 0.70 mmol) was dissolved in distilled methylene chloride (5 ml). 
1.0 M Boron trichloride (4.2 ml) was added dropwise at -78.degree. C. and 
the mixture was stirred for 1 hour. The mixture was further allowed to 
react for 2 hours at -20.degree. C. The reaction was stopped using a 
saturated aqueous sodium hydrogen carbonate solution (1.05 g). The mixture 
was separated using methylene chloride, after which the aqueous layer was 
concentrated and desalted by silica gel column chromatography (CHCl.sub.3 
:MeOH=5:1). Subsequently, the title compound was obtained via reverse 
phase HPLC. 
Ia-.alpha.! Melting point: 241-247 C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 281 nm (.epsilon.10300) 
UV .lambda..sub.max (H.sub.2 O) 259 nm (.epsilon.9000) 
Elementary analysis for C.sub.10 H.sub.14 N.sub.6 O.sub.3 S.1H.sub.2 O 
Calculated C: 37.97, H: 5.10, N: 26.57 
Found C: 37.90, H: 5.12, N: 26.39 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.00 (1H, s), 6.67 (2H, b r, D.sub.2 O 
exchangeable), 5.78 (2H, br, D.sub.2 O exchangeable), 5.75 (1H, br, 
D.sub.2 O exchangeable ),5.58 (1 H, br, D.sub.2 O exchangeable), 5.56 (1H, 
d, J=7.3 Hz), 4.90 (1H, br, D.sub.2 exchangeable), 4.45 (1H, t, J=7.3 Hz), 
3.86 (1H, dt, J=1.0, 11.0 Hz), 3.70 (1H, t, J=7.8 Hz), 3.6 3 (1H, m), 3.45 
(1H, dt, J=8.1, 11.0 Hz) 
Ia-.beta.! Melting point: 292-295.degree. C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 282 nm (.epsilon.0400) 
UV .lambda..sub.max (H.sub.2 O) 258 nm (.epsilon.8900) 
Elementary analysis for C.sub.1 H.sub.14 N.sub.6 O.sub.3 S.0.75H.sub.2 O 
Calculated C: 38.52, H: 5.01, N: 26.95 
Found C: 38.82, H: 4.97, N: 26.89 
.sup.1 H--NMR (DMSOd.sub.6) .delta.7.93 (1H, s), 6.67 (2H, b r, D.sub.2 O 
exchangeable), 5.93 (1H, d, J=5.4 Hz) 5.74 (1H, d, J=4.9 Hz, D.sub.2 O 
exchangeable), 5. 51 (1H, d, J=4.9 Hz, D.sub.2 O exchangeable), 5.1 9 (1H, 
br, D.sub.2 O exchangeable), 4.12 (1H, dt, J=5.9, 6.8 Hz), 4.04 (1H, dt, 
J=5.4, 6.8 Hz), 3.83 (1H, dd, J=4.9, 10.7 Hz), 3.68 (1H, dd, J=6.8, 10.7 
Hz), 3.22 (1H, ddd, J=4.9, 5.9, 6.8 Hz,) 
Example 2: 
Synthesis of Ib-.beta.!9-(4-thio-.delta.-D-arabinofuranosyl)guanine (in 
formula I!, B=guanine): 
The procedure of Example 1-5) was repeated using 
1-O-acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose and guanine, 
thereby obtaining the title compound. The target compound was also able to 
be obtained by treating the compound prepared in Example 1 with deaminase. 
Ib-.beta.! Melting point: 260-264.degree. C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 273 nm (.epsilon.9900) 
UV .lambda..sub.max (H.sub.2 O) 256 nm (.epsilon.13200) 
Elementary analysis for C.sub.10 H.sub.13 N.sub.5 O.sub.4 S.1H.sub.2 O 
Calculated C: 37.85, H: 4.76, N: 22.07 
Found C: 37.84, H: 4.76, N: 21.71 
.sup.1 H--NMR (DMSOd.sub.6) .delta.10.56 (1H, br), 7.92 (1H, s), 6.44 (2H, 
br, D.sub.2 O exchangeable), 5.86 (1H, d, J=5.4 Hz), 5.71 (1H, d, J=5.4 
Hz, D.sub.2 O exchangeable), 5.49 (1H, d, J=4.9 Hz, D.sub.2 O 
exchangeable), 5.14 (1H, t, J=5.4 Hz, D.sub.2 O echangeable), 4.07 (1H, 
dd, J=5.4, 11.0 Hz), 4.03 (1H, dd, J=6.6, 11.0 Hz) 3.83 (1H, dt, J =5.4, 
5.9 Hz), 3.67 (1H, dt, J=5.4, 5.9 Hz), 3.21 (1H, dt, J=5.4, 6.6 Hz) 
Example 3: 
Synthesis of Ic-.alpha.!9-(4-thio-.alpha.-D-arabinofuranosyl)adenine and 
Ic-.beta.!9-(4-thio-.beta.-D-arabinofuranosyl)adenine (in formula I!, 
B=adenine): 
The procedure of Example 1-5) was repeated using 
1-O-acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose and adenine, 
thereby obtaining the title compound. 
Ic-.alpha.! Melting point: 250.degree. C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 261 nm (.epsilon.13600) 
Elementary analysis for C.sub.10 H.sub.13 N.sub.5 O.sub.3 S.sub.1 
Calculated C: 42.40, H: 4.63, N: 24.72 
Found C: 42.32, H: 4.60, N: 24.44 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.41 (1H, s), 8.15 (1H, s) 7.24 (2H, br, 
D.sub.2 O exchangeable), 5.79 (1H, d, J=4.9 Hz, D.sub.2 O exchangeable), 
5.73 (1H, d, J=7.3 Hz), 5.61 (1H, d, J=4.4 Hz, D.sub.2 O exchangeale), 
4.93 (1H, t, J=4.6 Hz, D.sub.2 O exchangeable), 4.56 (1H, dt, J=4.4, 7.3 
Hz), 3.89 (1H, dt, J=3.9, 10.7 Hz), 3.75 (1H, dt, J=4. 4, 7.8 Hz), 3.66 
(1H, ddd, J=3.9, 7.8, 8.1 Hz), 3.50 (1H, dt, J=8.1, 10.7 Hz) 
Ic-.beta.! Melting point: 138-140.degree. C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 261 nm (.epsilon.11800) 
Elementary analysis for C.sub.10 H.sub.13 N.sub.5 O.sub.3 S.sub.1.2H.sub.2 
O 
Calculated C: 37.61, H: 5.37, N: 21.93 
Found C: 37.66, H: 5.37, N: 21.93 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.36 (1H, s), 8.13 (1H, s), 7.22 (2H, 
br, D.sub.2 O exchangeable), 6.05 (1H, d, J=5.4 Hz), 5.72 (1H, br, D.sub.2 
O exchangeable ), 5.51 (1H, d, J=2.9 Hz, D.sub.2 O exchangeable), 5.19 
(1H, br, D.sub.2 O exchangeable), 4.18-4.1 1 (2H, m), 3.87 (1H, dd, J=3.9, 
11.2 Hz), 3.7 8 (1H, dd, J=6.6, 11.2 Hz), 3.25 (1H, ddd, J=3.9, 5.9, 6.6 
Hz) 
Example 4: 
Synthesis of Id-.alpha.!9-(4-thio-a-D-arabinofuranosyl)-2-aminopurine and 
Id-.beta.!9-(4-thio-p-D-arabinofuranosyl)-2-aminopurine (in formula I!, 
B=2-aminopurine): 
The procedure of Example 1-5) was repeated using 
1-O-acetyl-2,3,5-tri-0-benzyl-4-thio-D-arabinofuranose and 2-aminopurine, 
thereby obtaining the title compound. 
Id-.alpha.! 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.56 (1H, s), 8.37 (1H, s), 6.54 (1H, 
s), 5.81 (1H, d, J=5.9 Hz), 5.65 (1H, d, J=7.3 Hz), 5.62 (1H, d, J=4.9 
Hz), 4.93 (1H, t, J=5.1 Hz), 4.47 (1H, dt, J=7.3 Hz), 3. 89 (1H, dt, 
J=3.4, 11.2 Hz), 3.72 (1H, dt, J=8.1 Hz), 3.64 (1H, dt, J=3.4, 8.1, 8.3 
Hz), 3. 43 (1H, dt, J=8.3, 11.2 Hz) 
Id-.beta.! 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.56 (1H, s), 8.29 (1H, s), 6.51 (2H, 
s), 6.00 (1H, d, J=4.9 Hz), 5.74 (1H, d, J=4.9 Hz), 5.50 (1H, d, J=4.4 
Hz), 5.16 (1H, t, J=4.9 Hz), 4.10 (1H, dt, J=4.9, 5.9 H z), 3.92 (1H, dt, 
J=3.4, 11.2 Hz), 3.75 (1H, dt, J=5.9, 8.3 Hz), 3.70 (1H, dt, J=7.8, 8.3, 
11.2 Hz), 3.51 (1H, dt, J=7.8, 11.2 Hz) 
Example 5: 
Synthesis of Ie-.alpha.!7-(4-thio-.alpha.-D-arabinofuranosyl)adenine and 
Ie-.beta.!7-(4-thio-.beta.-D-arabinofuranosyl)adenine (in formula I!, 
B=adenine): 
The procedure of Example 1-5) was repeated using 
1-O-acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose and silylated 
adenine (prepared via subjecting adenine and a catalytic amount of 
ammonium sulfate to refluxing in hexamethyldisilazane overnight), thereby 
obtaining the title compound. 
Ie-.alpha.! Melting point: 247-249.degree. C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 275 nm (.epsilon.9000) 
UV .lambda..sub.max (H.sub.2 O) 251 nm (sh, .epsilon.6000) 
Elementary analysis for C.sub.10 H.sub.13 N.sub.5 O.sub.3 
S.sub.1.0.55H.sub.2 O 
Calculated C: 40.96, H: 4.85, N: 23.88 
Found C: 41.27, H: 5.22, N: 23.99 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.62 (1H, s), 8.22 (1H, s), 6.96 (2H, 
br, D.sub.2 O exchangeable), 6.01 (1H, br, D.sub.2 O exchangeable), 5.93 
(1H, d, J=7.3 H z), 5.63 (1H, d, J=4.4 Hz, D.sub.2 O exchangeable), 5.03 
(1H, t, J=5.1 Hz, D.sub.2 O exchangeable), 4. 16 (1H, dt, J=7.3, 8.3 Hz), 
3.91 (1H, ddd, J=3.4, 5.1, 10.7 H z), 3.80 (1H, t, J=8.3 Hz), 3. 63 (1H, 
ddd, J=3.4, 7.8, 8.3 Hz), 3.53 (1H, d dd, J=5.1, 7.8, 10.7 Hz) 
Ie-.beta.! Melting point: 163-167.degree. C. (H.sub.2 O) 
UV .lambda..sub.max (H.sub.2 O) 273 nm (.epsilon.8900) 
UV .lambda..sub.max (H.sub.2 O) 249 nm (sh, .epsilon.6000) 
Elementary analysis for C.sub.10 H.sub.13 N.sub.5 O.sub.3 
S.sub.1.0.75H.sub.2 O 
Calculated C: 40.47, H: 4.92, N: 23.59 
Found C: 40.83, H: 4.87, N: 23.64 
.sup.1 H--NMR (DMSOd.sub.6) .delta.68.89 (1H, s), 8.15 (1H, s) 6.80 (2H, 
br, D.sub.2 O exchangeable), 6.08 (1H d, J=5.9 Hz), 5.78 (1H, d, J=5.9 Hz, 
D.sub.2 O exchangeable), 5.46 (1H, d, J=5.9 Hz, D.sub.2 O exchangeable), 
5.33 (1H, t, J=4.6 Hz, D.sub.2 O exchangeable), 4.14-4.10 (1H, m), 
3.87-3.83 (1H, m), 3.81-3.79 (2H, m), 3.16 (1H, ddd, J=4.4, 7.8, 8.3 Hz) 
Example 6: 
Synthesis of 
If-.alpha.!7-(4-thio-.alpha.-D-arabinofuranosyl)-2,6-diaminopurine and 
If-.beta.!7-(4-thio-.beta.-D-arabinofuranosyl)-2,6-diaminopurine (in 
formula I!, B=2,6-diaminopurine): 
The procedure of Example 1-5) was repeated using 
1-O-acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose and silylated 
2,6-diaminopurine (prepared via subjecting 2,6-diaminopurine and a 
catalytic amount of ammonium sulfate to refluxing in hexamethyldisilazane 
overnight), thereby obtaining the title compound. 
If-.alpha.! 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.21 (1H, s), 6.49 (2H, s), 5.97 (1H, 
bs), 5.76 (1H, d, J=7.8 Hz), 5.62 (1H, bs), 5.59 (2H, s), 5.02 (1H, bs), 
4.12 (1H, dt, J=8.3 Hz), 3.89 (1H, m), 3.58-3.49 (2H, m), 3.46 (1H, dt) 
If-.beta.! 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.51 (1H, s), 6.30 (2H, s), 5.95 (1H, d, 
J=5.4 Hz), 5.44 (2H, s), 4.08 (1 H, dd, J=5.4 Hz), 3.86 (1H, dd, J=8.3 
Hz),3. 78-3.72 (2H, dd.times.2, J=3.4, 4.9, 11.2 Hz),3. 13 (1H, dt, J=3.4, 
4.9, 8.3 Hz) 
Example 7: 
Synthesis of 
Ig-.alpha.!7-(4-thio-.alpha.-D-arabinofuranosyl)-2-aminopurine and 
Ig-.beta.!7-(4-thio-.beta.-D-arabinofuranosyl)-2-aminopurine (in formula 
I!, B=2-aminopurine): 
The procedure of Example 1-5) was repeated using 
1-O-acetyl-2,3,5-tri-O-benzyl-4-thio-D-arabinofuranose and silylated 
2-aminopurine (prepared via subjecting 2-aminopurine and a catalytic 
amount of ammonium sulfate to refluxing in hexamethyldisilazane 
overnight), thereby obtaining the title compound. 
Ig-.alpha.! 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.79 (1H, s), 8.42 (1H, s), 6.28 (2H, 
s), 5.93 (1H, bs), 5.70 (1H, d, J=7. 3 Hz), 5.65 (1H, bs), 4.99 (1H, bs), 
4.23 (1H, dt, J=7.3, 7.8 Hz), 3.91 (1H, dt, J=3.4, 11. 2 Hz), 3.70 (1H, 
dt, J=7.8, 8.3 Hz), 3.66 (1H, m, J=3.4, 7.8, 8.3 Hz), 3.51 (1H, dt, J=7.8, 
11.2 Hz) 
Ig-.beta.! 
.sup.1 H--NMR (DMSOd.sub.6) .delta.8.76 (1H, s), 8.59 (1H, s), 6.14 (1H, 
s), 6.00 (1H, d, J=5.9 Hz), 5.69 (1H, d, J=5.4 Hz),5.45 (1H, d, J=5.4 
Hz),5.26 (1H, t, J=4.9 Hz), 4.09 (1H, m), 3.93 (1H, m, J=5.9 Hz), 3.81 
(1H, m, J=4.4 Hz), 3.77 (1H, m), 3.24 (1H, m) 
Formulation Example 1: Tablets 
______________________________________ 
Compound of the invention 
30.0 mg 
Microcrystalline cellulose 
25.0 mg 
Lactose 39.5 mg 
Starch 40.0 mg 
Talc 5.0 mg 
Magnesium stearate 0.5 mg 
______________________________________ 
Using the above components, tablets were prepared via a routine method. 
Formulation Example 2: Capsules 
______________________________________ 
Compound of the invention 
30.0 mg 
Lactose 40.0 mg 
Starch 15.0 mg 
Talc 5.0 mg 
______________________________________ 
Using the above components, capsules were prepared via a routine method. 
Formulation Example 3: Injection preparation 
______________________________________ 
Compound of the invention 
30.0 mg 
Glucose 100.0 mg 
______________________________________ 
The above components were dissolved in purified water for injection 
preparations, thereby obtaining an injection liquid. 
Test Examples 
Test Method 
(1) Anti-HSV-1 Activity and Anti-HSV-2 Activity 
1. Human fibroblasts derived from fetal lungs were subjected to 
subculturing in Eagle MEM supplemented with 10% semi-fetal calf serum 
(Mitsubishi Chemical Corporation) at a 1:2-4 split every 4 days. 
2. A suspension of cells obtained by splitting their parent cells (1:2) was 
seeded in a 12-well multi-plate (2 ml/well), followed by culturing for 4-5 
days at 37.degree. C. in a CO.sub.2 -incubator. 
3. The culture liquid was discarded, and Hanks' MEM (250 .mu.l) containing 
50-150 PFU of VR-3 strain of HSV-1 or MS strain of HSV-2 was inoculated, 
and the virus was allowed to be adsorbed for 30 minutes at 37.degree. C. 
Thereafter, the viral liquid was discarded. 
4. A 2.5% serum-added Eagle MEM containing a test compound and 0.8% 
methylcellulose was added and the resultant mixture was incubated in a 
CO.sub.2 -incubator for 2-3 days at 37.degree. C. Generally, a test 
compound is diluted in serial 1/2 log.sub.10, and the maximal 
concentration is 10 .mu.g/ml. 
5. The culture liquid was discarded, and the cells were stained with a 0.5% 
crystal violet solution. Under a stereoscopic light microscope, the number 
of plaques in each well was counted. Using the equation below, the plaque 
formation inhibitory ratio (percent inhibition) was obtained. 
EQU Percent Inhibition=(1 -N.sub.1 /N.sub.2).times.100 
wherein N.sub.1 represents the number of plaques in wells containing the 
test compound and N.sub.2 represents the number of plaques containing in 
the control well (which contains no test compound). 
6. The plaque formation inhibitory ratio was plotted on a graph with 
respect to the concentration of the test compound (logarithmic 
representation). From this doseplaque inhibition curve, the concentration 
of the test compound exhibiting 50% inhibition was obtained (ED.sub.50). 
(2) Anti-Varicella Zoster Virus (VZV) Activity 
1. Human fibroblasts derived from fetal lungs were subjected to 
subculturing in Eagle MEM supplemented with 10% semi-fetal calf serum 
(Mitsubishi Chemical Corporation) at a 1:2-4 split every 4 days. 
2. A suspension of cells obtained by splitting their parent cells (1:2) was 
seeded in a 12-well multi-plate (2 ml/well), followed by culturing for 4-5 
days at 37.degree. C. in a CO.sub.2 -incubator. 
3. The culture liquid was discarded, and 750 .mu.l of a 5% serum-added 
Eagle MEM containing 50-100 PFU of Oka strain of VZV was inoculated, and 
the virus was allowed to be adsorbed for 1 hour at 37.degree. C. 
4. Without removing the virus, 750 .mu.l of Hanks' MEM containing the test 
compound was added, and the resultant mixture was incubated in a CO.sub.2 
-incubator at 37.degree. C. Generally, a test compound is diluted in 
serial 1/2 log.sub.10, and the maximal concentration is 10 .mu.g/ml. 
5. After culturing for 4-5 days, the culture liquid was discarded, and the 
cells were stained with a 0.5% crystal violet solution. Under a 
stereoscopic light microscope, the number of plaques in each well was 
counted. Using the equation used in (1) above, the plaque formation 
inhibitory ratio was obtained. 
6. The plaque formation inhibitory ratio was plotted on a graph with 
respect to the concentration of the test compound (logarithmic 
representation). From this doseplaque inhibition curve, the concentration 
of the test compound exhibiting 50% inhibition was obtained (ED.sub.50). 
(3) Anti-Human Cytomegalovirus Activity 
1. Human fibroblasts derived from fetal lungs were subjected to 
subculturing in Eagle MEM supplemented with 10% semi-fetal calf serum 
(Mitsubishi Chemical Corporation) at a 1:2-4 split every 4 days. 
2. A suspension of cells obtained by splitting their parent cells (1:2) was 
seeded in a 12-well multi-plate (2 ml/well), followed by culturing for 4 
days at 37.degree. C. in a CO.sub.2 -incubator. 
3. The culture liquid was discarded, and 750 .mu.l of a 5% serum-added 
Eagle MEM containing 50-100 PFU of AD-169 strain of HCMV was inoculated, 
and the virus was allowed to be adsorbed for 1 hour at 37.degree. C. 
4. Without removing the virus, 750 .mu.l of Hanks' MEM containing the test 
compound was added, and the resultant mixture was incubated in a CO.sub.2 
-incubator at 37.degree. C. for 4 days. Generally, a test compound is 
diluted in serial 1/2 log.sub.10, and the maximal concentration is 10 
.mu.g/ml. 
5. The medium was changed to a fresh 2.5% serum-added Eagle MEM containing 
0.8% methylcellulose and the test compound having the same concentration, 
followed by culturing further for 4-5 days. 
6. The culture liquid was discarded, and the cells were stained with 
May-Gruenwald's-Giemsa (.times.10). Under a stereoscopic light microscope, 
the number of plaques in each well was counted. Using the equation used in 
(1) above, the plaque formation inhibitory ratio was obtained. 
7. The plaque formation inhibitory ratio was plotted on a graph with 
respect to the concentration of the test compound (logarithmic 
representation). From this doseplaque inhibition curve, the concentration 
of the test compound exhibiting 50% inhibition was obtained (ED.sub.50). 
The results of these tests are shown in Table 1 below. 
TABLE 1 
______________________________________ 
Compound ED.sub.50 (.mu.g/ml) 
No. HSV-1 HSV-2 VZV HCMV 
______________________________________ 
Ia-.beta. 0.52 0.40 0.11 0.022 
Ib-.beta. 0.49 0.59 0.11 0.011 
______________________________________