JPH08165287A - 1,2-dioxetane derivative - Google Patents

Abstract

PURPOSE: To provide a 1,2-dioxetane derivative, useful as a substrate for immunoassay, excellent in thermal stability and capable of inducing the chemiluminescence. CONSTITUTION: This compound is represented by formula I [R<1> is H, an alkyl or Si(R<9> R<10> R<11> ); R<2> , R<3> , R<5> and R<6> are each independently H or an alkyl; R<4> is an alkyl, hydroxyl group, etc.; R<7> is an alkyl; Ar is an aryl, an alkoxyl- substituted aryl, etc.], e.g. 3-t-butyl-4-methoxy-3-[(2-methoxyethoxy)methyl]-4-(3'- phosphoryloxy)phenyl-1,2-dioxetane. The compound is obtained by reacting an ester of formula II (Ar' is an aryl, etc.) with an alkyl halide of the formula, R<7> X (X is a halogen) in the presence of a strong base, then reacting the prepared enol ether with an alkyl-lithium, etc., and subsequently reacting the resultant product with singlet oxygen. The compound of formula I has a functional group, OR<1> and can readily be bound to an amino acid, a peptide, etc.

Description

Detailed Description of the Invention

[0001]

FIELD OF THE INVENTION The present invention has the general formula

[Chemical 4] (In the formula, R ¹ is a hydrogen atom, an alkyl group or —Si (R ⁹ R
¹⁰ R ¹¹ ), R ² , R ³ , R ⁵ and R ⁶ are hydrogen atoms or alkyl groups, and R ⁴ is an alkyl group, a hydroxyl group, an alkoxyl group or —OSi (R ⁹ R ¹⁰ R ¹¹ ). . R ⁷ is an alkyl group. Ar is an aryl group substituted with an unsubstituted or -R ^8. R ⁸ is an alkoxyl group, -OSi
(R ⁹ R ¹⁰ R ¹¹ ) or a phosphate group. R ⁹ , R ¹⁰ and R ¹¹ are alkyl groups. ) Related to the 1,2-dioxetane derivative. The 1,2-dioxetane derivative represented by the general formula (I) is a compound capable of inducing chemiluminescence, and can be used, for example, as a substrate for immunoassay.

[0002]

2. Description of the Related Art Conventionally, various 1,2-dioxetane derivatives have been synthesized, but it has been known that a compound having a spiroadamantyl group bonded to the 3-position is particularly useful as a chemiluminescent substrate (for example, (See Japanese Patent Publication No. 5-21918 and Japanese Patent Publication No. 5-45590).

[0003]

However, the conventional compounds have low thermal stability and are not sufficient in the amount of light emission, and improvements thereof have been desired.

[0004]

The present inventor has found that the 1,2-dioxetane derivative represented by the general formula (I) can overcome the drawbacks of the conventional compounds and can complete the present invention. It is a thing. The 1,2-dioxetane derivative represented by the general formula (I) of the present invention is O
It is a very useful compound because it has a functional group R ¹ (or R ⁴ ), and has a molecular design so that it can be easily bonded to amino acids or peptides.

The present invention will be described below in accordance with the reaction formula.

[0006]

Embedded image (In the formula, R ² , R ³ , R ⁵ , R ⁶ and R ⁷ are the same as the above. R ⁴¹ is a hydrogen atom or an alkyl group. Ar ′ is unsubstituted or substituted with —R ^81. R ⁸¹ is an alkoxyl group or —OSi (R ⁹ R ¹⁰ R ¹¹ ). ⁹ , R ¹⁰ and R ¹¹ are the same as the above.)

In describing the present invention in detail below, the "alkyl group" in the present invention means a linear or branched alkyl group having 1 to 20 carbon atoms which may have a substituent. The alkyl group is a straight chain of methyl, ethyl, propyl, butyl, pentine, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, icodecyl. A group and a group in which the above alkyl group is appropriately branched to each other. The optionally substituted group is, for example, a hydroxyl group,
Examples thereof include an alkoxyl group, an aryl group and a heterocyclic group. Examples of the alkoxyl group include methoxy, ethoxy,
Propoxy, butoxy, pentyloxy, hexyloxy, methoxyethoxy, methoxypropoxy, ethoxyethoxy, ethoxypropoxy, methoxyethoxyethoxy groups and the like, and the aryl group thereof, for example, phenyl, naphthyl group, etc. Examples of the group include furyl, thienyl and pyridyl groups.

In the present invention, the "alkoxyl group" means
It is the same as the alkoxyl group which may be substituted on the above-mentioned alkyl group, and in the present invention, the "aryl group" means phenyl, aromatic hydrocarbon group such as naphthyl group and nitrogen in the ring,
It refers to a heteroaryl group having an oxygen or sulfur atom.

(Steps 1-1 and 1-2) In this step, the ester represented by the general formula (II) or (V) is converted into t
-In the presence of a strong base such as potassium butoxide, sodium hydride, lithium hydride, etc.

[Chemical 6] (In the formula, R ⁷ is the same as above, and X is a halogen atom), and an enol ether represented by the general formula (III) or (VI) is obtained by reacting with an alkyl halide or a dialkyl sulfuric acid represented by Is manufactured.

The compound represented by the general formula (II) which is a raw material in this step is, for example, a compound represented by the corresponding arylcarboxylic acid ester and the general formula

[Chemical 7] (Wherein R ⁴¹ , R ⁵ and R ⁶ are the same as defined above, and R is an alkyl group), and a compound that can be synthesized from an acetic acid ester compound.

Embedded image (In the formula, R ⁴¹ , R ⁵ and R ⁶ are the same as above.), Which can be easily produced from the chloride represented by the general formula (IX),
The compounds represented by formula (X) and formula (XI) are industrially easily available compounds.

(Steps 2-1 and 2-2) In this step, the compound represented by the general formula (III) or (VI) is added to a reducing agent such as diisobutylaluminum hydride or an alkyllithium or halogenated compound. By reacting with an organometallic reagent such as alkylmagnesium, the compound of the general formula (IV)
Alternatively, the compound represented by (VII) is produced.

When reducing with diisobutylaluminum hydride, an aromatic hydrocarbon solvent such as toluene is used,
The reaction is performed at -100 ° C to 0 ° C.

When alkyl lithium is used,
0 to 1 using an organic ether solvent such as tetrahydrofuran
The reaction is carried out at 00 ° C.

(Third step) In this step, the compound represented by the general formula (IV) or (VII) is represented by the general formula (IX) in the presence of a strong base such as sodium hydride or lithium hydride. With an alkyl halide, or in the presence of an organic amine such as triethylamine or imidazole,
Reaction with halogenated trialkylsilane

[Chemical 9] (In the formula, R ² , R ³ , R ⁵ , R ⁶ , R ⁷ and Ar ′ are the same as the above. R ²¹ is an alkyl group or —Si (R ⁹ R ¹⁰ R
¹¹ ) and R ⁴² is an alkyl group, an alkoxyl group or —OSi (R ⁹ R ¹⁰ R ¹¹ ). ) The compound represented by these is manufactured.

In both this step and the first step, the reaction is carried out at 0 to 180 ° C. using a solvent such as dimethylformamide, dimethylsulfoxide, tetrahydrofuran and the like.
To carry out the alkylation reaction in good yield, the above general formula (IV)
Alternatively, it is preferable that the raw material represented by (VII) is treated with a strong base and then reacted with an alkyl halide.

(Fourth Step) This step is carried out according to the general formula (VII)
The compound represented by I) is deprotected and then reacted with a corresponding halogenated trialkylsilane or halogenated phosphate to form a silyloxy group or a phosphoric acid group.

[Chemical 10] (In the formula, Ar ″ is —OSi (R ⁹ R ¹⁰ R ¹¹ ) or

[Chemical 11] It is an aryl group substituted with. R ⁴² is an alkyl group, an alkoxyl group or —OSi (R ⁹ R ¹⁰ R ¹¹ ). R ⁹ ,
R ¹⁰ and R ¹¹ are the same as defined above, and R ¹² and R ¹³ are alkyl groups or groups in which R ¹² and R ¹³ may be combined to form a ring. R ²¹ is the same as above. ) The compound represented by these is manufactured.

The deprotection reaction can be carried out by a method well known to those skilled in the art, that is, by reacting the anion of alkylthiol or by subjecting it to a hydrogenation reaction. It may be appropriately selected depending on the group to be protected.

Further, in this step, as described above, a halogenated trialkylsilane or a halogenated phosphate is reacted to produce the compound represented by the general formula (XII).

Further, in this step, when a phosphate group is introduced, it is preferable to subject it to the following fifth step as a phosphate, for example, when the desired phosphate is reacted with chloroethylene phosphate. Can be achieved by converting it into a sodium salt of cyanoethyl phosphate with sodium cyanide, further eliminating the cyanoethyl group, and converting into a sodium salt of ammonium. Further, the ammonium sodium salt can be easily converted into a disodium salt by reacting with sodium hydrogen carbonate, for example.

(Fifth Step) This step is carried out according to the general formula (I
V), the general formula (VII), the general formula (VIII) or the compound represented by the general formula (XII) is reacted with singlet oxygen to form the 1,2-type compound represented by the general formula (I). It is for producing a dioxetane derivative.

The reaction of singlet oxygen is carried out according to the general formula (I
V), the above general formula (VII), the above general formula (VIII) or the above general formula (XII), is dissolved in a solvent such as dichloromethane, dichloroethane, carbon tetrachloride, alcohol or the like to prepare methylene blue or rose. It is achieved by irradiating visible light under an oxygen atmosphere in the coexistence of a photosensitizer such as bengal or tetraphenylporphine (TPP). The reaction is carried out at -80 ° C to 0 ° C.

[0022]

EXAMPLES The present invention will be described in more detail with reference to Examples and Reference Examples.

(Reference Example 1)

[Chemical 12]

4.99 g of dried zinc chloride (37 mmo) in 30 ml of anhydrous dichloromethane at room temperature under a nitrogen atmosphere.
l), ethyl benzoyl acetate (Compound [1]) 14.6
g (76 mmol) and t-butyl chloride 16.
5 ml (152 mmol) was added, and the mixture was heated under reflux overnight.
The reaction mixture was poured into saturated saline and extracted with dichloromethane. The extract layer was dried over magnesium sulfate, concentrated, and distilled under reduced pressure to give ethyl 2-benzoyl-2-t-butylacetate (Compound [2]) 8.76 g, yield 46.5.
Obtained in%.

Mp45-47 ° C / bp 89-90 ° C
(0.4 mmHg) ¹ HNMR (400 MHz, CDCl ₃ ); δ1.16
(S, 9H), 1.17 (t, J = 7.2Hz,
3H), 4.13 (q, J = 7.2Hz, 2
H), 4.31 (s, 1H), 7.26 to 7.96.
(M, 5H) p pm IR (KBr); 3368, 3173, 3067, 16
62, 1624, 1404, 686 cm ^-1 Mass (m / z,%); 248 (M ⁺ , 1), 192.
(100), 146 (10), 105 (58), 77
(2)

(Reference example 2)

[Chemical 13]

Compound [2] 6.23 synthesized in Reference Example 1
g (25 mmol) was added to 50 ml of anhydrous DMSO, and 5.61 g (50 mmol) of potassium t-butoxide was added to the solution stirred at room temperature under a nitrogen atmosphere and stirred for 30 minutes. This solution was cooled to 0 ° C. and dimethylsulfate 4.75 m
After 1 (50 mmol) was added, the mixture was stirred at room temperature for 1 hour. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of dichloromethane and hexane 1: 2, and 2-t-butyl-3-methoxy-3-phenyl-
2.46 g of ethyl 2-propenoate (compound [3]),
The yield was 52.6%.

¹ HNMR (400 MHz, CDCl ₃ );
δ 0.86 (t, J = 6.8 Hz, 3H), 1.
30 (s, 9H), 3.30 (s, 3H), 3.8
1 (q, J = 6.8 Hz, 2H), 7.26 to 7.3
4 (m, 5H) pp m IR (liquid film); 2959,171.
8,1296,1072 cm ^-1 Mass (m / z,%); 262 (M ⁺ , 43), 24
7 (100), 187 (30), 105 (30), 87
(17), 77 (15).

(Reference Example 3)

Embedded image

Compound [3] 5.24 synthesized in Reference Example 2
g (20 mmol) was added to 10 ml of anhydrous toluene, and the mixture was stirred at -78 ° C under a nitrogen atmosphere. Diisobutylaluminum hydroxide (1.5 M toluene solution) was added to this solution.2.
92 ml (44 mmol) was added and stirred for 1 hour. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated to give 2-t-butyl-3-methoxy-3-phenyl-2-propen-1-ol (Compound [4]).
It was obtained as a colorless amorphous solid in a yield of 3.99 g and a yield of 90.7%.

¹ HNMR (400 MHz, C ₆ D ₆ ); δ
1.48 (s, 9H), 3.01 (s, 3H),
3.86 (d, J = 4.8 Hz, 2H), 7. 0
6-7.31 (m, 5H) ppm IR (KBr); 3285, 2957, 1633, 12
92, 1113, 1010, 696 cm ^-1 Mass (m / z,%); 220 (M ⁺ , 35), 20
5 (30), 187 (65), 163 (56), 105
(63), 77 (100).

(Example 1)

[Chemical 15]

210 m of compound [4] synthesized in Reference Example 3
g (0.95 mmol) and 5 mg of TPP were dissolved in 10 ml of dichloromethane, and the mixture was stirred at 0-5 ° C under an oxygen atmosphere. This solution was irradiated with a Na lamp (940 W) for 1 hour. The reaction mixture was concentrated, subjected to preparative TLC and developed with a 10: 1 mixture of hexane and diethyl ether to give 3-t-butyl-3-hydroxymethyl-4.
-Methoxy-4-phenyl-1,2-dioxetane (Compound [5]) was isolated in a yield of 86 mg and a yield of 35.8% as a yellow oil.

¹ HNMR (400 MHz, C ₆ D ₆ ); δ
0.59 (t, J = 7.3 Hz, 1H), 1.4
3 (s, 9H), 2.80 (s, 3H), 3.8
3 (q _AB , J = 7.3 Hz, 2H), 7.00-7.4
8 (m, 5H) ppm IR (liquid film); 3584, 296
6,1450,1249,1107,1041,706
cm ^-1 Mass (m / z,%); 253 (M ⁺ +1,0.
5), 220 (22), 205 (8), 187 (1
1), 163 (11), 136 (19), 117
(8), 105 (67), 85 (16), 77 (4
4), 55 (100)

(Reference Example 4)

Embedded image

0.24 g (6 mmol) of sodium hydride in 10 ml of anhydrous THF under a nitrogen stream at 0 ° C., Reference Example 3
1.16 g (5 mmol) of the compound [4] synthesized in 1. and 0.38 ml (6 mmol) of methyl iodide were sequentially added, followed by heating under reflux for 3 hours. The reaction mixture was poured into saturated aqueous ammonium chloride solution and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated to give 2-t-butyl-1,3-dimethoxy-1-.
1.15 g of phenyl-1-propene (compound [6])
Obtained as a yellow oil in a yield of 93.2%.

¹ HNMR (400 MHz, C ₆ D ₆ ); δ
1.07 (s, 9H), 2.98 (s, 3H),
3.05 (s, 3H), 3.65 (s, 2H),
7.07-7.39 (m, 5H) ppm IR (liquid film); 2955, 108
6,704 cm ^-1 Mass (m / z,%); 234 (M ⁺ , 17), 21
9 (14), 203 (27), 187 (32), 177
(35), 163 (31), 147 (32), 121
(76), 105 (95), 77 (100), 55 (7
1).

(Example 2)

[Chemical 17]

Compound [6] 100 m synthesized in Reference Example 4
g (0.43 mmol) and TPP 5 mg were dissolved in dichloromethane 10 ml, and the mixture was stirred at 0 to 5 ° C. under an oxygen atmosphere. This solution was irradiated with a Na lamp (940 W) for 1 hour. The reaction mixture was concentrated, subjected to preparative TLC, developed with a 10: 1 mixed solvent of hexane and diethyl ether to give 3-t-butyl-4-methoxy-3-methoxymethyl-4-phenyl-1,2-dioxetane. (Compound [7]) was isolated as a yellow oily substance in 77 mg in a yield of 67.7%.

¹ HNMR (400 MHz, C ₆ D ₆ ); δ
1.50 (s, 9H), 2.35 (s, 3H),
2.85 (s, 3H), 3.58 (q _AB , J =
6.0Hz, 2H), 7.05 to 7.59 (m, 5H)
ppm IR (liquid film); 2932, 145
0,1255,1099,975,704 cm ^-1 Mass (m / z,%); 234 (M ⁺ -32,3),
187 (2), 177 (4), 136 (25), 130
(10), 105 (72), 85 (14), 77 (4
6), 55 (100).

(Reference Example 5)

Embedded image

7.9 g of dimethoxy magnesium (92 m
mol) at room temperature under a nitrogen atmosphere, diethyl ether 50 m
30 ml of diethyl ether was added to the stirred solution.
Dimethyl malonate dissolved in 11.0 g (83 mmo
1) was added dropwise over 20 minutes, followed by stirring for 30 minutes.
The solution was cooled and 3-anisoyl chloride (compound [8]) 14.0 dissolved in 20 ml of diethyl ether.
g (82 mmol) was added dropwise over 20 minutes, followed by 30
After stirring for 1 minute, the mixture was refluxed for 2 hours. The reaction mixture was poured into saturated aqueous ammonium chloride solution and extracted with ethyl acetate.
The extract layer was washed with saturated saline. The extract was dried over magnesium sulfate and concentrated. Diethyl ether was added to the concentrate, the insoluble matter was filtered off, and the filtrate was concentrated. Dimethyl 3-anisoylmalonate (compound

[9]) 17.1 g, yield 78.3%
To give a yellow oil.

¹ HNMR (90 MHz, CDCl ₃ ); δ
3.81 (s, 6H), 3.86 (s, 3H),
7.12-7.49 (m, 4H) ppm IR (liquid film); 3480, 300
9,2950,1678,1637,1440,139
6,1281,1238,1095 cm ^-1 Mass (m / z,%); 266 (M ⁺ , 18), 23
4 (11), 135 (100), 107 (27).

(Reference Example 6)

[Chemical 19]

Compound synthesized in Reference Example 5

[9] 17.0
g (64 mmol) was added to 80 ml of DMF, and 13.3 g (96 mmo of potassium carbonate was added to the solution stirred under ice cooling.
1) was added and stirred for 1 hour. 12.9 g (102 mmol) of dimethylsulfate was added to this solution, and the mixture was heated under reflux for 2 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and dichloromethane 1: 4,
Dimethyl (3-methoxyphenyl) methoxymethylenemalonate (Compound [10]) was 8.1 g, yield 45.3%.
Was obtained as a pale yellow oil.

¹ HNMR (90 MHz, CDCl ₃ ); δ
3.53 (s, 6H), 3.82 (s, 3H),
3.84 (s, 3H), 6.87 to 7.37 (
m, 4H) ppm IR (liquid film); 2957,175
6,1739,1693,1593,1533,143
5, 1217, 1150, 1035 cm ^-1 Mass (m / z,%); 280 (M ⁺ , 62), 24
9 (100), 221 (86), 205 (28), 19
1 (33), 181 (72), 135 (37).

(Reference Example 7)

Embedded image

Compound [10] 7.0 synthesized in Reference Example 6
g (25 mmol) was added to 50 ml of THF and methylmagnesium bromide (3M was added to the solution stirred under ice cooling.
42 ml (126 mmol) of a THF solution) was added dropwise and stirred for 1 hour, followed by heating under reflux for 2 hours. The reaction mixture was poured into saturated aqueous ammonium chloride solution and extracted with ethyl acetate. The extract layer was washed successively with saturated brine, saturated aqueous ammonium chloride solution and saturated brine, dried over magnesium sulfate and concentrated. When the concentrate was washed with hexane, 1,
1-bis (2-hydroxypropan-2-yl) -2-
Methoxy-2- (3-methoxyphenyl) ethene (compound [11]) was obtained as a colorless amorphous solid in 2.6 g and a yield of 37.1%.

[0049]¹HNMR (90MHz, CDCl₃); Δ
1.21 (s, 6H), 1.67 (s, 6H),
3.07 (s, 3H), 3.81 (broad
s, 5H), 6.76 to 7.35 (m, 4H) ppm IR (KBr); 3230, 2979, 1595, 14
85, 1321, 1234, 1182, 1094, 10
72,944,859,772 cm^-1  Mass (m / z,%); 262 (M⁺-H₂O, 5),
247 (5), 230 (60), 215 (3
1), 199 (29), 187 (30), 173
(11), 159 (8), 146 (7), 135 (10
0), 10 7 (55).

(Reference Example 8)

[Chemical 21]

Compound [11] 1.0 synthesized in Reference Example 7
0 g (3.6 mmol) was added to 20 ml of THF, and a solution stirred at 0 ° C. under a nitrogen stream was added with sodium hydride 0.4
3 g (10.8 mmol) was gradually added, and the mixture was stirred for 30 minutes. 0.9 ml of methyl iodide (14.4 m) was added to this solution.
mol) was added and the mixture was heated under reflux for 1 hour. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated to give 1-methoxy-1- (3-methoxyphenyl) -2,2-bis (2-methoxypropan-2-yl) ethene (compound [ 1.24 g of a crude product of [12]) was obtained as a pale yellow oily substance.

¹ HNMR (90 MHz, CDCl ₃ ); δ
1.42 (s, 6H), 1.57 (s, 6H),
2.72 (s, 3H), 3.06 (s, 3H),
3.22 (s, 3H), 3.80 (s, 3H), 6.7
6 to 7.50 (m, 4H) ppm IR (liquid film); 2957, 175
6,1739,1698,1593,1533,143
5,1217,1150,1035 cm ^-1 Mass (m / z,%); 308 (M ⁺ , 6), 293.
(2), 277 (4), 261 (4), 245 (4),
229 (3), 189 (3), 181 (8), 173
(2), 135 (3), 107 (3), 89 (17),
73 (100).

(Reference Example 9)

[Chemical formula 22]

Compound [11] 1.0 synthesized in Reference Example 7
0 g (3.6 mmol) was added to 20 ml of THF, and a solution stirred at 0 ° C. under a nitrogen stream was added with 0.1 parts of sodium hydride.
7 g (4.3 mmol) was gradually added and stirred for 30 minutes. 0.34 ml of methyl iodide (5.4 mm
ol) was added and the mixture was stirred at room temperature for 1 hour. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated to give 1- (2
-Hydroxypropan-2-yl) -2-methoxy-2
1.10 g of a crude product of-(3-methoxyphenyl) -1- (2-methoxypropan-2-yl) ethene (compound [13]) was obtained as a pale yellow oil.

¹ HNMR (90 MHz, CDCl ₃ ); δ
1.21 (s, 6H), 1.61 (s, 6H),
3.08 (s, 3H), 3.24 (s, 3H),
3.82 (s, 3H), 5.89 (s, 1H), 6.7
2 to 7.35 (m, 4H) ppm IR (liquid film); 3426, 296
7,2934,1693,1482,1383,128
8, 1213, 1170, 1098, 1052, 94
9,762 cm ^-1 Mass (m / z,%); 294 (M ⁺ , 5), 262
(15), 247 (78), 231 (27), 203
(10), 189 (31), 173 (36), 149
(13), 135 (45), 107 (22), 96 (5)
3), 73 (100).

(Example 3)

[Chemical formula 23]

Compound [11] 50 m synthesized in Reference Example 7
g (0.18 mmol) and 50 mg of silica gel-supported methylene blue were added to 15 ml of carbon tetrachloride, and the mixture was stirred at 0 ° C. in an oxygen atmosphere. Add Na lamp (940
Light irradiation was performed for 30 minutes in (W). The reaction mixture is concentrated,
It was subjected to preparative TLC, developed with a mixed solvent of hexane and ethyl acetate 3: 1 to give 3,3-bis (2-hydroxypropan-2-yl) -4-methoxy-4- (3-methoxyphenyl) -1, 2-dioxetane (compound [14]) 8
Isolated as a colorless oil in mg, yield 14.4%.

¹ HNMR (90 MHz, CDCl ₃ ); δ
1.47 (s, 3H), 1.61 (s, 3H),
1.68 (s, 3H), 1.70 (s, 3H),
3.46 (s, 3H), 3.82 (s, 3H), 6.8
4 to 7.65 (m, 4H) ppm

(Example 4)

[Chemical formula 24]

Compound [12] 100 synthesized in Reference Example 8
mg (0.32 mmol) and methylene blue-100 mg supported on silica gel were added to 15 ml of carbon tetrachloride, and the mixture was stirred at 0 ° C. in an oxygen atmosphere. Add Na lamp (9
Light irradiation was performed at 40 W) for 1.5 hours. The reaction mixture was concentrated, subjected to preparative TLC and developed with a 1: 4 mixed solvent of hexane and dichloromethane to give 3-methoxy-3- (3-
Methoxyphenyl) -4,4-bis (2-methoxypropan-2-yl) -1,2-dioxetane (compound [1
5]) was isolated as a yellow oil in 40 mg, 36.2% yield.

¹ HNMR (400 MHz, CDCl ₃ );
δ1.28 (s, 3H), 1.31 (broad
s, 3H), 1.50 (s, 3H), 1.56
(S, 3H), 2.06 (s, 3H), 3.27 (s,
3H), 3.86 (m, 3H), 6.99-7.
55 (m, 4H) ppm IR (liquid film); 2979, 293.
7,1729,1664,1595,1484,127
8,1256,1072,1041,782 cm ^-1 Mass (m / z,%); 308 (M ⁺ -32,4),
280 (14), 262 (5), 247 (9), 235
(22), 208 (54), 175 (100), 135
(31), 73 (100).

(Reference Example 10)

[Chemical 25]

Diisopropylamine 13.0 ml (9
2.8 mmol) was added to 75 ml of anhydrous THF under an argon atmosphere at room temperature and stirred to obtain 55.0 ml of butyllithium (1.62M hexane solution) (89.1 m).
(mol) and stirred for 30 minutes. This solution is -78 ° C
Cooled to 15.0 ml of ethyl t-butyl acetate (89.
5 mmol) and stirred for 20 minutes, and subsequently 10.05 g (60.5 m) of methyl 3-methoxybenzoate [16] [16]
mol) was added. The solution was stirred at −78 ° C. for 2 hours and 40 minutes, then at 0 ° C. for 1 hour and 30 minutes. The reaction mixture was poured into 1N hydrochloric acid and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. When the concentrate was applied to a silica gel column and eluted with dichloromethane, ethyl 2-t-butyl-2- (3-methoxybenzoyl) acetate (compound [17]) was 1
Obtained as a colorless oily substance with a yield of 2.24 g and a yield of 72.7%.

¹ HNMR (300 MHz, CDCl ₃ );
δ1.15 (s, 9H), 1.18 (t, J =
7.2Hz, 3H), 3.86 (s, 3H), 4.1
3 (q, J = 7.2 Hz, 2H), 4.28 (s,
1H), 7.11 (d with fine co
upling, J = 8.3 Hz, 1H), 7.3
7 (dd, J = 8.3 and 7.6 Hz, 1H),
7.48 (with swine fine couple
ng, 1H), 7.54 (d, J = 7.6 Hz,
1H) ppm IR (liquid film); 2964,291
2,1736,1696,1598,1582 cm ^-1 Mass (m / z,%); 278 (M ⁺ , 10), 22
2 (26), 176 (18), 135 (100)

(Reference Example 11)

[Chemical formula 26]

Compound [17] synthesized in Reference Example 10
30 g (4.68 mmol) was added to 10 ml of anhydrous DMSO, and 1.02 g (9.09 mmol) of potassium t-butoxide was added to the solution stirred at room temperature under a nitrogen atmosphere.
Stir for minutes. This solution was cooled to 0 ° C., 0.80 ml (8.44 mmol) of dimethylsulfate was added dropwise, and the mixture was stirred for 50 minutes. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 15: 2. As a result, ethyl 2-t-butyl-3-methoxy-3- (3-methoxyphenyl) -2-propenoate (compound [1
8]) was obtained as a colorless oily substance in 1.15 g and a yield of 84.2%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.91 (t, J = 7.1 Hz, 3H), 1.
29 (s, 9H), 3.33 (s, 3H), 3.8
0 (s, 3H), 3.86 (q, J = 7.1Hz, 2
H), 6.81 to 6.96 (m, 3H), 7.2
2 (t, J = 7.8Hz, 1H) ppm IR (liquid film); 2960,172
0, 1634, 1598, 1580 cm ^-1 Mass (m / z,%); 292 (M ⁺ , 60), 27
8 (31), 277 (100), 247 (21), 23
1 (21), 135 (35).

(Reference Example 12)

[Chemical 27]

Compound [18] synthesized in Reference Example 11.2.
49 g (8.53 mmol) was added to 30 ml of anhydrous toluene, and the mixture was stirred at -78 ° C under an argon atmosphere. To this solution, 10.0 ml (17.6 mmol) of diisobutylaluminum hydride (25% toluene solution) was added and stirred for 1 hour and 20 minutes. Methanol was added to the reaction mixture until there was no foaming, then the mixture was poured into a mixed solution of water and ethyl acetate,
After filtration through Celite, the organic layer was separated. The organic layer was washed with saturated brine, dried over magnesium sulfate, and concentrated to give 2
-T-Butyl-3-methoxy-3- (3-methoxyphenyl) -2-propen-1-ol (Compound [19])
Was obtained as a colorless oily substance with a yield of 2.08 g and a yield of 97.6%.

¹ HNMR (300 MHz, CDCl ₃ );
δ1.32 (s, 9H), 3.24 (s, 3
H), 3.82 (s, 3H), 3.94 (d, J = 5.
5Hz, 2H), 6.85 to 6.95 (m, 3
H), 7.24 to 7.32 (m, 1H) ppm IR (liquid film); 3464, 295.
6, 1636, 1598, 1580 cm ^-1 Mass (m / z,%); 250 (M ⁺ , 67), 23
5 (89), 219 (35), 217 (73), 193
(100), 187 (21), 135 (28) 133
(27).

(Reference Example 13)

[Chemical 28]

Compound [19] synthesized in Reference Example 12.
50 g (10.0 mmol) was added to 15 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. 6 in this solution
420 mg (00.5 mmol) of 0% sodium hydride
Was added and the mixture was heated with stirring at 110 ° C. for 10 minutes. 1.50 ml of neopentyl bromide (11.9 mmo) was added to this solution.
1) was added, and the mixture was heated with stirring at 110 ° C. for 2 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. Apply the concentrate to a silica gel column and add 1 of hexane and ethyl acetate.
When it was poured out with a mixed solvent of 0: 1, 2-t-butyl-1-methoxy-1- (3-methoxyphenyl) -3-
Neopentyloxy-1-propene (Compound [20])
Was obtained as a colorless oily substance in a yield of 2.18 g, 68.1%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.91 (s, 9H), 1.27 (s, 9
H), 2.83 (s, 2H), 3.25 (s, 3H),
3.61 (s, 2H), 3.81 (s, 3H),
6.86 (ddd, J = 8.2, 2.6 and
1.0Hz, 1H), 6.92 (s with
fine coupling, 1H), 7.00 (d
with fine coupling, J =
7.5 Hz, 1H). 7.23 (dd, J = 8.
2 and 7.5 Hz, 1 H) ppm IR (liquid film); 2956,286
8, 1636, 1598, 1580 cm ^-1 Mass (m / z,%); 320 (M ⁺ , 31), 26
3 (73), 249 (19), 234 (19), 233
(43), 219 (42), 217 (41), 203
(21), 193 (100), 187 (21), 177
(29), 121 (29), 111 (25), 97 (3
3), 83 (28), 71 (37), 57 (55).

(Reference Example 14)

[Chemical 29]

60% sodium hydride 530 mg (1
1.0 mmol of ethanethiol was added to a solution prepared by suspending 3.3 mmol) in 20 ml of anhydrous DMF at 0 ° C. under an argon atmosphere.
ml (13.5 mmol) was added. After stirring this solution at room temperature for 30 minutes, the compound synthesized in Reference Example 13 [20]
2.16 g (6.75 mmol) of anhydrous DMF 15 ml
Was dissolved in and added under reflux for 3 hours. The reaction mixture was poured into saturated aqueous ammonium chloride solution and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 5: 1 to give 2-t-butyl-1- (3-hydroxyphenyl).
1.2-g of -1-methoxy-3-neopentyloxy-1-propene (Compound [21]) was obtained as a colorless oily substance with a yield of 59.5%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.92 (s, 9H), 1.26 (s, 9
H), 2.83 (s, 2H), 3.25 (s, 3H),
3.58 (s, 2H), 6.80 (ddd, J =
8.1, 2.6 and 1.0Hz, 1H),
6.90 (dd, J = 2.6 and 1.5H
z, 1H), 6.98 (d with fine co
upling, J = 7.6 Hz, 1H), 7.2
0 (dd, J = 8.1 and 7.6Hz, 1
H) ppm IR (liquid film); 3400,296
0,2908,2872,1652,1596,148
2 cm ^-1 Mass (m / z,%); 306 (M ⁺ , 25), 24
9 (56), 219 (60), 205 (39), 204
(62), 203 (34), 189 (36), 179
(47), 161 (29), 153 (29), 121
(100).

(Reference Example 15)

Embedded image

Compound [21] 41 synthesized in Reference Example 14
1 mg (1.34 mmol) was added to 5 ml of anhydrous toluene, and the mixture was stirred at 0 ° C under an argon atmosphere. 0.22 ml (1.58 mmol) of triethylamine was added to this solution, followed by 2-chloro-1,3,2-dioxaphosphorane-2.
-Oxide (0.125 ml, 1.35 mmol) was added, and the mixture was stirred at 0 ° C for 30 minutes and then at room temperature for 2 hours. The reaction mixture was concentrated, ether was added to filter the insoluble matter, and the filtrate was concentrated to give 3- (2-t-butyl-1-methoxy-3-neopentyloxy-1-propen-1-yl). Phenyl ethylene phosphate (compound [22])
570 mg of a crude product of was obtained as a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.93 (s, 9H), 1.26 (s, 9
H), 2.84 (s, 2H), 3.25 (s, 3H),
3.56 (s, 2H), 4.27 to 4.41
(M, 2H), 4.43 to 4.57 (m, 2H)
H), 7.15 to 7.35 (m, 4H) ppm

(Reference Example 16)

[Chemical 31]

Compound [22] 57 synthesized in Reference Example 15
0 mg (1.38 mmol) was added to 5 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. 69 mg (1.34 mmo) of sodium cyanide (95%) was added to this solution.
1) was added and stirred overnight. The reaction mixture was concentrated, the concentrate was dissolved in hexane, extracted with water, and the extract layer was lyophilized to give sodium 3- (2-t-butyl-1-methoxy-3-neopentyloxy-1-propene-1. A crude purified product of -yl) phenyl-2'-cyanoethyl phosphate (Compound [23]) was obtained in an amount of 589 mg as an amorphous solid.

[0082]¹HNMR (300MHz, CD₃OD);
δ 0.95 (s, 9H), 1.31 (s, 9
H), 2.81 (t, J = 6.3 Hz, 2H), 2.8
 6 (s, 2H), 3.29 (s, 3H), 3.6
8 (s, 2H), 4.1 5 (dt, J = 7.7)
and 6.3 Hz, 2H), 7.10 to 7.3.
5 (m, 4H) ppm IR (KBr); 2958, 2868, 2260, 16
01,1579,1482,1262,1104cm^-1 Mass (FAB-pos, m / z,%); 485
([M + H + Na]⁺ , 26), 484 ([M + N
a]⁺, 100), 382 (24), 125 (5
5).

(Reference Example 17)

Embedded image

Compound [23] 48 synthesized in Reference Example 16
5 mg (1.05 mmol) was added to 2 ml of THF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 28% aqueous ammonia (3.0 ml) and water (1.0 ml) were added, and the mixture was stirred for 3 days. The reaction mixture was concentrated, the concentrate was dissolved in hexane and extracted with water. When the extract layer was freeze-dried, ammonium sodium 3- (2-t-butyl-1-
Methoxy-3-neopentyloxy-1-propene-1
-Yl) phenyl phosphate (compound [24] is 46
Obtained as 0 mg, an amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.94 (s, 9H), 1.31 (s, 9
H), 2.85 (s, 2H), 3.29 (s, 3H),
3.69 (s, 2H), 7.08 (d, J = 7.
4 Hz, 1H), 7.20 (s, 1H), 7.2
5 (dd, J = 7.4 and 8.0 Hz, 1
H), 7.35 (broad d, J = 8.0 Hz, 1
H) ppm IR (KBr); 2958, 2866, 1598, 15
79, 1481, 1217, 1084 cm ^-1 Mass (FAB-pos, m / z,%); 431
([M + H-NH ₄ + Na] ^+, 48), 343 (3
0), 329 (35), 125 (100).

(Example 5)

[Chemical 33]

Compound [24] 69 synthesized in Reference Example 17
mg (0.162 mmol) and TPP 2 mg were dissolved in dichloromethane 15 ml, and the mixture was stirred at 0 ° C. under an oxygen atmosphere. This solution was irradiated with light from a Na lamp (180 W) for 2 hours. The reaction mixture was concentrated, applied to a silica gel column, and sequentially poured out using dichloromethane, a mixed solvent of dichloromethane and methanol in a ratio of 4: 1 and 2: 1, to give 3-t-butyl-4-methoxy-3-neopentyloxymethyl-4. 23 mg of-(3'-phosphoryloxy) phenyl-1,2-dioxetane ammonium sodium salt (Compound [25]) was obtained as an amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.73 (s, 9H), 1.33 (s, 9
H), 2.25 (d, J = 8.2 Hz, 1H), 2.6
4 (d, J = 8.2Hz, 1H), 3.07
(S, 3H), 3.47 (d, J = 10.2H
z, 1H), 3.84 (d, J = 10.2Hz, 1
H), 7.13 to 7.55 (m, 4H) ppm

(Example 6)

Embedded image

Compound [25] 13m synthesized in Example 5
g (0.028 mmol) was dissolved in 2.8 ml (0.028 mmol) of 0.01N sodium hydrogen carbonate aqueous solution and freeze-dried. As a result, 3-t-butyl-4-
Methoxy-3-neopentyloxymethyl-4- (3 '
13 mg of -phosphoryloxy) phenyl-1,2-dioxetane disodium salt (Compound [26]) was obtained as an amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.74 (s, 9H), 1.33 (s, 9
H), 2.28 (d, J = 8.2 Hz, 1H), 2.6
3 (d, J = 8.2Hz, 1H), 3.06
(S, 3H), 3.44 (d, J = 10.2H
z, 1H), 3.84 (d, J = 10.2Hz, 1
H), 7.00 to 7.60 (m, 4H) ppm IR (KBr); 2960, 2872, 1590, 14
84, 1296, 1272, 1108, 992 cm ^-1 Mass (FAB-pos, m / z,%); 485
([M + Na] ⁺ , 21), 463 ([M + H] ⁺ , 3
8), 401 (26), 379 (14), 299
(52), 277 (73), 125 (43), 115
(100).

(Reference Example 18)

Embedded image

Compound [21] 16 synthesized in Reference Example 14
8 mg (0.549 mmol) was dissolved in 2 ml of DMF, and the mixture was stirred at room temperature under an argon atmosphere. 66 mg (0.969 mmol) of imidazole and t-
Butyldimethylchlorosilane 100mg (0.663m
mol) was added and stirred for 2 hours. The reaction mixture was poured into water and extracted with ethyl acetate. Wash the extract layer with saturated saline,
The extract was dried over magnesium sulfate and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate of 20: 1, and 2-t-butyl-1- [3-
(T-Butyldimethylsiloxy) phenyl] -1-methoxy-3-neopentyloxy-1-propene (Compound [27]) was obtained as a colorless oily substance in a yield of 158 mg and a yield of 68.5%.

¹ HNMR (90 MHz, CDCl ₃ ); δ
0.19 (s, 6H), 0.90 (s, 9H),
0.98 (s, 9H), 1.26 (s, 9H),
2.81 (s, 2H), 2.33 (s, 3H), 3.6
2 (s, 2H), 6.73 to 6.87 (m, 2
H), 6.97 to 7.03 (m, 1H), 7.1
4 to 7.19 (m, 1H) ppm

(Example 7)

Embedded image

Compound [27] 50 synthesized in Reference Example 18
mg (0.119 mmol) and 5 mg of TPP were dissolved in 10 ml of dichloromethane, and under oxygen atmosphere, at -78 ° C.
It was stirred at. This solution was irradiated with a Na lamp (940 W) for 2 hours. The reaction mixture was concentrated, applied to a silica gel column, and poured out with a mixed solvent of hexane and ethyl acetate of 20: 1. 3-t-butyl-4- [3- (t
-Butyldimethylsiloxy) phenyl] -4-methoxy-3-neopentyloxymethyl-1,2-dioxetane (Compound [28]) was obtained as a colorless oily substance in 35 mg, yield 65%.

¹ HNMR (90 MHz, CDCl ₃ ); δ
0.20 (s, 6H), 0.68 (s, 9H),
0.99 (s, 9H), 1.29 (s, 9H),
2.38 (q _AB , J = 8.4 Hz, 2H), 3.04
(S, 3H), 3.63 (q _AB , J = 10.0H
z, 2H), 6.78-6.89 (m, 1H),
7.00 to 7.24 (m, 3H) ppm IR (liquid film); 2960, 160
0,1480,1280,1100,920,840c
m ^-1

(Reference Example 19)

Embedded image

25.0 ml of diisopropylamine (0.
178 mol) was added to 200 ml of anhydrous THF under an argon atmosphere at room temperature, and 110 ml (0.178 m) of butyllithium (1.62M hexane solution) was added to the stirred solution.
ol) was added and stirred for 1 hour. This solution was cooled to −78 ° C. and 30.0 ml (0.17
8 mol) and stirred for 30 minutes, and then 21.0 g of methyl 3-benzyloxybenzoate (Compound [29])
After adding (86.8 mmol), the mixture was stirred at room temperature for 3 hours. The reaction mixture was poured into 1N hydrochloric acid and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate (9: 1) to give 2- (3-benzyloxybenzoyl) -2-t-.
28.31 g of ethyl butyl acetate (compound [30]),
The yield was 92.2%.

Mp: 53.5-54.0 ° C. (colorless fine-grained crystal, recrystallized from methanol) ¹ HNMR (300 MHz, CDCl ₃ ); δ1.14
(S, 9H), 1.18 (t, J = 7.1Hz,
3H), 4.13 (q, J = 7.1Hz, 2
H), 4.26 (s, 1H), 5.11 (s, 2H),
7.18 (dwine fine couple)
ng, J = 8.2 Hz, 1H), 7.32 to 7.
48 (m, 6H), 7.52 to 7.59 (m, 2H) p
pm IR (KBr); 2964, 1728, 1696, 15
92 cm ^-1 Mass (m / z,%); 354 (M ⁺ , 19), 29
8 (18), 211 (39), 91 (100).

(Reference Example 20)

[Chemical 38]

Compound [30] 2 synthesized in Reference Example 19
8.1 g (79.4 mmol) of anhydrous DMSO 200 m
In addition to 1, 10.1 g (0.179 mol) of potassium t-butoxide was added to the solution stirred at room temperature under a nitrogen atmosphere and stirred for 15 minutes. This solution was cooled to 0 ° C., 15.0 ml (0.158 mol) of dimethyl sulfate was added dropwise over 15 minutes, and then the mixture was stirred at room temperature for 30 minutes. 0 this solution
Cooled to 7.degree. C., 7.30 g (65.
1 mmol) followed by 5.4 ml of dimethylsulfate (56.9)
(mmol) was added in two portions and the mixture was stirred for 4 hours and 30 minutes at room temperature. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 10: 1 to give ethyl 3- (3-benzyloxyphenyl) -2-t-butyl-3-methoxy-2-propenoate ( The compound [31]) was obtained as a colorless oily substance in a yield of 85.2% (24.9 g).

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.90 (t, J = 7.1 Hz, 3H), 1.
29 (s, 9H), 3.30 (s, 3H), 3.8
5 (q, J = 7.1 Hz, 2H), 5.06 (s, 2
H), 6.90 to 7.01 (m, 3H), 7.2
2 (t, J = 7.8Hz, 1H), 7.28 ~
7.47 (m, 5H) ppm IR (liquid film); 2960, 171
8, 1636, 1596, 1580 cm ^-1 Mass (m / z,%); 368 (M ⁺ , 59), 35
4 (25), 353 (100), 91 (83).

(Reference Example 21)

[Chemical Formula 39]

Compound [31] 1 synthesized in Reference Example 20
9.63 g (53.3 mmol) of anhydrous toluene 150
In addition to ml, the mixture was stirred at -78 ° C under an argon atmosphere.
To this solution, 70.0 ml (0.123 mol) of diisobutylaluminum hydride (25% toluene solution) was added and stirred for 45 minutes. Further, 7.0 ml (1% of 25% toluene solution) of diisobutylaluminum hydride was added to this solution.
2.3 mmol) was added and stirred for 2 hours. The reaction mixture was poured into 1N hydrochloric acid and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. Crystallization from the concentrate with a mixed solvent of hexane and ethyl acetate yielded 3- (3-benzyloxyphenyl) -2-t-butyl-3-methoxy-2-propen-1-ol (Compound [32]). Obtained in 8.70 g, yield 50.0%. The filtrate was concentrated, applied to a silica gel column, and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 4: 1 to obtain 6.80 g of a compound [32] in a yield of 39.1%.

Mp: 59.5-60.0 ° C. (colorless granular crystal, recrystallized from hexane and ethyl acetate) ¹ HNMR (300 MHz, CDCl ₃ ); δ 0.94
(T, J = 5.4 Hz, 1H), 1.31 (s,
9H), 3.23 (s, 3H), 3.92 (d,
J = 5.4 Hz, 2H), 5.09 (s, 2H), 6.
90 to 7.00 (m, 3H), 7.25 to 7.4
7 (m, 6H) ppm IR (KBr); 3464, 2956, 1634, 15
88 cm ^-1 Mass (m / z,%); 326 (M ⁺ , 46), 31
1 (43), 269 (39), 91 (100).

(Reference Example 22)

[Chemical 40]

Compound [32] 77 synthesized in Reference Example 21
2 mg (2.37 mmol) was added to 2.0 ml (8.96 mmol) of 1-bromoundecane, and the mixture was stirred at room temperature under an argon atmosphere. 2.0 ml (25.0 mmol) of 50% aqueous sodium hydroxide solution and 89 mg (0.276) of tetrabutylammonium bromide were added to this solution.
mmol) was added and the mixture was heated with stirring at 80 ° C. for 3 hours. Tetrabutylammonium bromide 99 was added to this solution.
mg (0.307 mmol) was added, and the mixture was heated with stirring for 3 hours and 30 minutes. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate of 10: 1, and 1- (3-benzyloxyphenyl) -2-t-butyl-1-methoxy-3-undecanoxy-1-propene was obtained. (Compound [33]) was obtained as a colorless oily substance in a yield of 39.4% (448 mg).

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.84 to 0.92 (m, 3H), 1.20
1.36 (m, 16H), 1.28 (s, 9H)
, 1.44 to 1.56 (m, 2H), 3.20 (t,
J = 6.6 Hz, 2 H), 3.23 (s, 3)
H), 3.67 (s, 2H), 5.07 (s, 2)
H), 6.95 (d with fine couple)
ing, J = 8. 2Hz, 1H), 6.99
(D, J = 7.6 Hz, 1H), 7.05 (s
with fine coupling, 1H), 7.
25 (dd, J = 8.2 and 7.6 Hz,
1H), 7.28 to 7.48 (m, 5H) ppm IR (liquid film); 2928, 285
6, 1636, 1596, 1580 cm ^-1 Mass (m / z,%); 480 (M ⁺ , 28), 42
4 (31), 423 (100), 333 (19), 10
9 (16), 91 (67).

(Reference Example 23)

Embedded image

Compound [33] 71 synthesized in Reference Example 22
8 mg (1.50 mmol) and 10% Pd-C
125 mg was added to a mixed solvent of 7 ml of ethyl acetate and 2 ml of methanol, and the mixture was stirred under a hydrogen atmosphere at room temperature for 2 hours.
The reaction mixture was filtered through Celite and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 5: 1 to give 2-t-butyl-1-
(3-hydroxyphenyl) -1-methoxy-3-undecanoxy-1-propene (compound [34]) was 528
It was obtained as a colorless oily substance with a yield of 90.5%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.84 to 0.92 (m, 3H), 1.18 to
1.35 (m, 16H), 1.27 (s, 9H)
, 1.43 to 1.59 (m, 2H), 3.21 (t,
J = 6.5 Hz, 2 H), 3.24 (s, 3
H), 3.66 (s, 2H), 6.80 (dd
d, J = 8.0, 2.6 and 0.8 Hz, 1
H), 6.86 (s with fine co
upling, 1H), 6.95 (d with h)
fine coupling, J = 7.6 Hz, 1
H), 7.21 (dd, J = 8.0 and
7.6 Hz, 1 H) ppm IR (liquid film); 3384,292
8, 2856, 1636, 1596, 1584 cm ^-1 Mass (m / z,%); 390 (M ⁺ , 16), 33
4 (23), 333 (100), 219 (17), 20
3 (32), 179 (22), 161 (20).

(Reference Example 24)

Embedded image

Compound [34] 84 synthesized in Reference Example 23
mg (0.215 mmol) was dissolved in DMF (1.5 ml), and the mixture was stirred at room temperature under an argon atmosphere. To this solution was added 30 mg (0.441 mmol) of imidazole and t-
Butyldimethylchlorosilane 60 mg (0.398 mm
ol) was added and the mixture was stirred for 2 hours. To this solution was further added 18 mg (0.264 mmol) of imidazole and 36 mg (0.239 mmo of t-butyldimethylchlorosilane.
1) was added and stirred for 1 hour. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a 20: 1 mixed solvent of hexane and ethyl acetate to give 2-t-butyl-1- [3- (t-butyldimethylsiloxy) phenyl] -1-methoxy-3.
-Undecanoxy-1-propene (Compound [35]) was obtained as a colorless oily substance in 100 mg, yield 92.1%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.19 (s, 6H), 0.84 to 0.92
(M, 3H), 0.99 (s, 9H), 1.18-
1.35 (m, 16H), 1.28 (s, 9H),
1.42-1.53 (m, 2H), 3.17
(T, J = 6.6 Hz, 2H), 3.23 (s,
3H), 3.68 (s, 2H), 6.80 (ddd, J
= 8.1, 2.5 and 0.9 Hz, 1
H), 6.84 (s with fine co)
upling, 1H), 6.96 (d with fi
ne coupling, J = 7.6Hz, 1
H), 7.18 (dd, J = 8.1 and d
7.6Hz, 1H) ppm

(Embodiment 8)

[Chemical 43]

Compound [35] 60 synthesized in Reference Example 24
mg (0.119 mmol) and TPP 5 mg were dissolved in dichloromethane 20 ml, and the mixture was stirred at 0 ° C. under an oxygen atmosphere. This solution was irradiated with a Na lamp (180 W) for 3 hours. The reaction mixture was concentrated, subjected to preparative TLC, developed with a 20: 1 mixed solvent of hexane and benzene to give 3-t-butyl-4- [3- (t-butyldimethylsiloxy) phenyl] -4-methoxy-. 3-Undecanoxymethyl-1,2-dioxetane (Compound [36])
Was isolated as a colorless oil in 38 mg, yield 59.6%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.20 (s, 6H), 0.84 to 0.92
(M, 3H), 0.99 (s, 9H), 1.02
1.35 (m, 18H), 1.28 (s, 9H),
2.47 (dt, J = 9.0 and 6.2H
z, 1H), 2.87 (dt, J = 9.0 an)
d6.4 Hz, 1H), 3.03 (s, 3H), 3.5
0 (d, J = 10.1 Hz, 1H), 3.72
(D, J = 10.1 Hz, 1H), 6. 84 (d
dd, J = 8.0, 2.4 and 1.0 Hz, 1
H), 6. 90 to 7.18 (m, 2H), 7.2
4 (t, J = 8.0 Hz, 1H) p pm

(Reference Example 25)

[Chemical 44]

Compound [19] synthesized in Reference Example 12
164 g (4.66 mmol) was added to 12 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution was added 340 mg of 60% sodium hydride (8.50 mmo
l) and benzyl bromide 0.83 ml (6.98)
mmol) was added and the mixture was stirred at room temperature for 1.5 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column, and hexane and ethyl acetate were added.
When it was poured out with a mixed solvent of 0: 1, 1.099 g of 3-benzyloxy-2-t-butyl-1-methoxy-1- (3-methoxyphenyl) -1-propene (Compound [37]), Obtained as a colorless oil in a yield of 69.4%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 1.30 (s, 9H), 3.25 (s, 3
H), 3.76 (s, 2H), 3.78 (s, 3H),
4.31 (s, 2H), 6.87 (ddd, J =
8.2, 2.6 and 1.1Hz, 1H),
6.93 to 7.00 (m, 2H), 7.19 to
7.39 (m, 6H) ppm IR (liquid film); 2956, 163
4,1596,1580 cm ^-1 Mass (m / z,%); 340 (M ⁺ , 36), 28
3 (30), 234 (75), 233 (29), 219
(32), 217 (46), 203 (23), 193
(100), 187 (24), 177 (67), 91
(88).

(Reference Example 26)

Embedded image

Compound [37] 48 synthesized in Reference Example 25
7 mg (1.43 mmol) and 60% sodium hydride 150 mg (3.75 mmol) were added to DMF6 ml, and it stirred at 0 degreeC under argon atmosphere. 0.23 ml (3.11 mmol) of ethanethiol was added to this solution, and the mixture was stirred for 10 minutes and then heated and stirred at 120 ° C. for 2 hours. The reaction solution was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 5: 1. 3-benzyloxy-2-t-butyl-1- (3-
Hydroxyphenyl) -1-methoxy-1-propene (Compound [38]) was obtained in a yield of 218 mg and a yield of 46.7%.

Mp: 93.0-94.0 ° C. (colorless granular crystal, recrystallized from hexane) ¹ HNMR (300 MHz, CDCl ₃ ); δ1.29
(S, 9H), 3.24 (s, 3H), 3.76
(S, 2H), 4.32 (s, 2H), 4.72
(S, 1H), 6.79 (ddd, J = 8.1, 2.5
and 0.7Hz, 1H), 6.85 (bro
ad s, 1H), 6.94 (dwith fi
ne coupling, J = 7.7Hz, 1H)
, 7.19 (dd, J = 8.1 and 7.7H
z, 1H), 7.20 to 7.35 (m, 5H) p
pm IR (KBr); 3272, 2956, 2908, 16
38,1594 cm ^-1 Mass (m / z,%); 326 (M ⁺ , 36), 26
9 (32), 220 (76), 219 (27), 205
(33), 203 (60), 179 (82), 163
(68), 161 (36), 91 (100).

(Reference Example 27)

Embedded image

Compound [38] 12 synthesized in Reference Example 26
7 mg (0.390 mmol) was dissolved in 2 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. 55 mg (0.808 mmol) of imidazole and t
-Butyldimethylchlorosilane 85 mg (0.564 m
mol) was added and the mixture was stirred overnight. The reaction solution was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate of 100: 1 to give 3-benzyloxy-2-t-.
Butyl-1- [3- (t-butyldimethylsiloxy) phenyl] -1-methoxy-1-propene (compound [3
9]) was obtained as a colorless oily substance with a yield of 144 mg and a yield of 84.0%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.17 (s, 6H), 0.97 (s, 9
H), 1.29 (s, 9H), 3.23 (s, 3H),
3.77 (s, 2H), 4.29 (s, 2H),
6.80 (ddd, J = 8.0, 2.5 and
1.0Hz, 1H), 6.85 (s with
fine coupling, 1H), 6.97 (d
with fine coupling, J =
7.6 Hz, 1 H), 7.18 (dd, J = 8.
0 and 7.6 Hz, 1H), 7.17 to 7.35.
(M, 5 H) ppm IR (liquid film); 2956, 293
6, 1634, 1598, 1580, 1262 cm ^-1 Mass (m / z,%); 440 (M ⁺ , 22), 38.
3 (19), 335 (28), 334 (100), 33
3 (29), 293 (52), 277 (42), 235
(13), 91 (65).

(Example 9)

[Chemical 47]

Compound [39] 49 synthesized in Reference Example 27
mg (0.111 mmol) and 4 mg of TPP were added to 20 ml of dichloromethane, and the mixture was stirred at 0 ° C. under an oxygen atmosphere. This solution was irradiated with a Na lamp (180 W) for 4 hours. The reaction mixture was concentrated, applied to a silica gel column, and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 200: 1 to give 3-benzyloxymethyl-3-t.
-Butyl-4- [3- (t-butyldimethylsiloxy)
Phenyl] -4-methoxy-1,2-dioxetane (Compound [40]) was obtained as a colorless oily substance in 42 mg, yield 79.9%.

¹ HNMR (300 MHz, CDCl ₃ );
δ0.17 (broad s, 6H), 0.98
(S, 9H), 1.29 (s, 9H), 3.05
(S, 3H), 3.57 (d, J = 10.2Hz, 1
H), 3.71 (d, J = 11.5 Hz, 1
H), 3.84 (d, J = 10.2 Hz, 1
H), 3.84 (d, J = 11.5 Hz, 1H), 6.
83-6.90 (m, 1H), 6.95-7.3
0 (m, 8H) ppm IR (liquid film); 2960, 293
6,1602,1586,1256,1096 cm ^-1 Mass (m / z,%); 472 (M ⁺ , trac
e), 440 (2), 266 (26), 210 (2
2), 209 (100), 177 (20), 149 (1
1), 91 (41).

(Reference Example 28)

Embedded image

Compound [17] synthesized in Reference Example 10.
00 g (14.4 mmol) was added to 30 ml of anhydrous DMSO, and 3.26 g (29.1 mmol) of potassium t-butoxide was added to the solution stirred at room temperature under an argon atmosphere and stirred for 15 minutes. 2.7 ml (28.8 mmol) of isopropyl bromide was added to this solution, and the mixture was stirred for 4 hours. 11.02 g of potassium t-butoxide (9
8.2 mmol) and isopropyl bromide 9.3.
ml (99.0 mmol) was added in 5 portions over 24 hours. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. Apply the concentrate to a silica gel column,
When it was poured out with a mixed solvent of hexane and ethyl acetate of 9: 1, 2-t-butyl-3-isopropoxy-3-
Ethyl (3-methoxyphenyl) -2-propenoate (Compound [41]) was obtained as a colorless oil in 3.73 g, yield 81.0%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.86 (t, J = 7.1 Hz, 3H), 1.
15 (d, J = 6.2 Hz, 6 H), 1.30 (
s, 9H), 3.79 (s, 3H), 3.79 (q, J
= 7.1 Hz, 2 H), 3.87 (hept, J
= 6.2 Hz, 1 H), 6.83 (dd d, J =
8.2, 2.6 and 0.8Hz, 1H), 6.8
7 (s with fine couple)
g, 1H), 6.91 (dd, J = 7.5 and
0.9 Hz, 1 H), 7.21 (dd, J = 8.2)
and 7.5 Hz, 1H) ppm IR (liquid film); 2976,171.
8, 1632, 1598, 1580 cm ^-1 Mass (m / z,%); 320 (37), 275 (1
2), 263 (15), 233 (24), 232 (9
4), 217 (55), 176 (20), 135 (10
0).

(Reference Example 29)

[Chemical 49]

Compound [41] synthesized in Reference Example 28.3.
72 g (11.6 mmol) was added to 35 ml of anhydrous toluene, and the mixture was stirred at -78 ° C under an argon atmosphere. 15.0 ml (26.4 mmol) of diisobutylaluminum hydride (25% hexane solution) was added to this solution, and the mixture was stirred for 4 hours. The reaction mixture was poured into a mixed solution of water and ethyl acetate stirred at 0 ° C., stirred for 20 minutes, and then filtered through Celite.
The organic layer was separated, washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and was poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 4: 1 to give 2-t-butyl-3-isopropoxy-3- (3
-Methoxyphenyl) -2-propen-1-ol (Compound [42]) was obtained as a colorless oily substance with a yield of 2.47 g and a yield of 76.4%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.89 (t, J = 5.6 Hz, 1H), 1.
12 (d, J = 6.2Hz, 6H), 1.33 (
s, 9H), 3.75 (hept, J = 6.2 Hz, 1
H), 3.82 (s, 3H), 3.89 (d, J
= 5.6 Hz, 2H), 6.82 to 6. 92
(M, 3H), 7.23 to 7.31 (m, 1H) ppm IR (liquid film); 3460, 297.
6, 1628, 1598, 1580 cm ^-1 Mass (m / z,%); 278 (M ⁺ , 31), 26.
1 (20), 219 (46), 218 (24), 203
(54), 135 (100), 107 (20).

(Reference Example 30)

Embedded image

Compound [42] synthesized in Reference Example 29
195 g (4.30 mmol) was added to 12 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 325 mg of 60% sodium hydride (8.13 mmo)
1) and 1.20 ml neopentyl bromide (9.
(53 mmol) was added and the mixture was heated with stirring at 100 ° C. for 1 hour.
The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate of 20: 1.
1.194 g of t-butyl-1-isopropoxy-1- (3-methoxyphenyl) -3-neopentyloxy-1-propene (Compound [43]) was obtained as a colorless oily substance in a yield of 79.8%. Was given.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.88 (s, 9H), 1.12 (d, J =
6.2Hz, 6H), 1.29 (s, 9H), 2.7
6 (s, 2H), 3.56 (s, 2H), 3.77
(Hept, J = 6.2 Hz, 1H), 3.80
(S, 3H), 6.84 (ddd, J = 8.0,
2.7 and 1.0 Hz, 1H), 6.87 (s)
with fine coupling, 1
H), 6.94 (d with fine co
upling, J = 7.5 Hz, 1H), 7.22 (d
d wit hine coupling, J =
8.0 and 7.5 Hz, 1 H) ppm IR (liquid film); 2956,286
8, 1632, 1598, 1580 cm ^-1 Mass (m / z,%); 348 (M ⁺ , 50), 29
1 (40), 261 (18), 219 (43), 218
(23), 203 (75), 179 (27), 135
(100), 107 (18).

(Reference Example 31)

[Chemical 51]

60% sodium hydride 389 mg (9.
(73 mmol) in anhydrous DMF (15 ml) at 0 ° C. under argon atmosphere, and ethanethiol (0.8 m) was added to the suspension.
1 (10.8 mmol) was added and stirred for 20 minutes. Compound [43] 1.522 synthesized in Reference Example 30 was added to this solution.
g (4.37 mmol) was dissolved in 10 ml of anhydrous DMF and added, and the mixture was heated with stirring at 120 ° C. for 6 hours. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and eluted with a mixed solvent of hexane and ethyl acetate 10: 1 and then 4: 1 to give 2-t-butyl-1- (3-hydroxyphenyl).
-1-Isopropoxy-3-neopentyloxy-1-
1.208 g of propene (compound [44]), yield 8
Obtained as a pale yellow oil at 2.7%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.90 (s, 9H), 1.12 (d, J =
6.1Hz, 6H), 1.27 (s, 9H), 2.7
6 (s, 2H), 3.53 (s, 2H), 3.79
(Hept, J = 6.1 Hz, 1H), 4.65
(S, 1H), 6.78 (ddd, J = 8.1,
2.6 and 0.9Hz, 1H), 6.84 (s)
with fine coupling, 1
H), 6.92 (d with fine co
upling, J = 7.6 Hz, 1H), 7.18 (d
d, J = 8.1 and 7.6 Hz, 1H) pp
m IR (liquid film); 3400, 296.
0,2872,1628 cm ^-1 Mass (m / z,%); 334 (M ⁺ , 41), 27
7 (35), 247 (23), 205 (55), 204
(37), 189 (72), 165 (26), 121
(100), 93 (16).

(Reference Example 32)

Embedded image

Compound [44] 12 synthesized in Reference Example 31
2 mg (0.365 mmol) was dissolved in 2 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 0.15 ml (1.08 mmol) of triethylamine and 110 mg (0.
730 mmol) was added and the mixture was stirred overnight. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and hexane and ethyl acetate 25:
When it was poured out with the mixed solvent of 1, 2-t-butyl-1
-[3- (t-butyldimethylsiloxy) phenyl]-
121 mg of 1-isopropoxy-3-neopentyloxy-1-propene (compound [45]), yield 73.9.
% As a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.18 (s, 6H), 0.87 (s, 9
H), 0.98 (s, 9H), 1.11 (d, J = 6.
2 Hz. 6H), 1.28 (s, 9H), 2.7
5 (s, 2H), 3.58 (s, 2H), 3.7
4 (hept, J = 6.2 Hz, 1H), 6.7
5 to 6.82 (m, 2H), 6.93 (d with
fine cou pling, J = 7.6 Hz,
1H), 7.15 (dd, J = 7.6 a nd
7.4 Hz, 1 H) ppm IR (liquid film); 2956, 286
4,1630,1596,1578,1260,108
6 cm ^-1 Mass (m / z,%); 448 (M ⁺ , 100), 3
91 (70), 361 (26), 319 (56), 31
8 (25), 303 (52), 279 (30), 261
(74), 235 (45).

(Embodiment 10)

Embedded image

Compound [45] 68 synthesized in Reference Example 32
mg (0.152 mmol) and 4 mg of TPP were dissolved in 20 ml of dichloromethane, and the mixture was stirred at 0 ° C. under an oxygen atmosphere. This solution was irradiated with a Na lamp (180 W) for 2 hours. The reaction mixture was concentrated, applied to a silica gel column, and then poured out with a mixed solvent of hexane and ethyl acetate of 100: 1. 3-t-butyl-4- [3- (t
-Butyldimethylsiloxy) phenyl] -4-isopropoxy-3-neopentyloxymethyl-1,2-dioxetane (compound [46]) 52 mg, yield 71.4
% As a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.21 (broad s, 6H), 0.72
(S, 9H), 0.90 to 1.10 (m, 12H)
, 1.15 to 1.30 (m, 3H), 1.32 (s,
9H), 2.10 to 2.24 (m, 1H), 2.
56 (d, J = 8.3 Hz.1H), 3.3 2
(D, J = 10, 1 Hz, 1H), 3.40 to 3.64.
(M, 2H), 6.70 to 6.96 (m, 2
H), 7.14 to 7.40 (m, 2H) pp m IR (liquid film); 2960, 293.
6,2868,1602,1586,1256,110
0 cm ^-1 Mass (m / z,%); 448 (M ⁺ -32, 7),
294 (45), 238 (25), 237 (67), 2
35 (25), 196 (36), 195 (100), 1
67 (19), 135 (21), 71 (54), 57
(70).

(Reference Example 33)

[Chemical 54]

Compound [32] 65 synthesized in Reference Example 21
2 mg (2.00 mmol) and 2- (2-methoxyethoxy) ethyl bromide 0.55 ml (4.05 m
mol) in THF 4 ml, and under argon atmosphere,
Stir at room temperature. 408m of sodium hydroxide in this solution
g (10.2 mmol), 67 mg (0.208 mmol) of tetrabutylammonium bromide and 0.
1 ml was added and the mixture was heated under reflux for 8 hours and 40 minutes. To this solution was further added 0.60 ml (4.42 mmol) of 2- (2-methoxyethoxy) ethyl bromide, 530 mg (13.3 mmol) of sodium hydroxide and 69 mg (0.214 mmo) of tetrabutylammonium bromide.
1) was added and the mixture was heated under reflux overnight. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 4: 1, and 1- (3-benzyloxyphenyl) -2-t-butyl-1-methoxy-3- [2- ( Two
-Methoxyethoxy) ethoxy] -1-propene (Compound [47]) was obtained as a colorless oily substance in a yield of 69.1% (591 mg).

¹ HNMR (300 MHz, CDCl ₃ );
δ1.28 (s, 9H), 3.22 (s, 3
H), 3.34 (s, 3H), 3.37 to 3.42 (
m, 2H), 3.46 to 3.51 (m, 2H),
3.54 to 3.61 (m, 4H), 3.77
(S, 2H), 5.07 (s, 2H), 6.92-
7.02 (m, 3H), 7.22 to 7.48 (m, 6
H) ppm IR (liquid film); 2876,163
6, 1596, 1580 cm ^-1 Mass (m / z,%); 428 (M ⁺ , 49), 37.
1 (10), 309 (16), 308 (13), 293
(26), 251 (67), 217 (29), 91 (1
00).

(Reference Example 34)

[Chemical 55]

Compound [47] 45 synthesized in Reference Example 33
1 mg (1.05 mmol) and 10% Pd-C, 5
4 mg of a 5: 2 mixed solvent of ethyl acetate and methanol 7
The mixture was added to ml and stirred at room temperature for 2.5 hours under hydrogen atmosphere. The reaction mixture was filtered through Celite, and the filtrate was concentrated. The concentrate was applied to a silica gel column and poured out with a 2: 1 mixed solvent of hexane and ethyl acetate to give 2-t-butyl-1- (3-hydroxyphenyl) -1-methoxy-.
3- [2- (2-methoxyethoxy) ethoxy] -1-
Propene (compound [48]) 300 mg, yield 84.
Obtained as a colorless oil at 2%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 1.26 (s, 9H), 3.29 (s, 3
H), 3.40 to 3.46 (m, 2H), 3.44 (
s, 3H), 3.62 (s, 2H), 3.60-
3.74 (m, 6H), 6.80-6.87
(M, 2H), 7.18 to 7.25 (m, 2H) pp
m IR (liquid film); 3380, 295.
6,2928,2876,1634,1596,158
2 cm ^-1 Mass (m / z,%); 338 (M ⁺ , 77), 28
1 (33), 219 (30), 218 (23), 203
(73), 161 (100), 103 (36), 59
(31).

(Reference Example 35)

[Chemical 56]

Compound [48] 13 synthesized in Reference Example 34
5 mg (0.399 mmol) was dissolved in 2 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. 0.11 ml (0.789 mmol) of triethylamine was added to this solution.
And t-butyldimethylchlorosilane 78 mg (0.
518 mmol) was added and the mixture was stirred for 1.5 hours. 0.10 ml (0.717 mmo) of triethylamine was added to this solution.
l) and t-butyldimethylchlorosilane 58 mg
(0.385 mmol) was further added and stirred for 1 hour. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and was poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 4: 1.
-Butyl-1- [3- (t-butyldimethylsiloxy)
Phenyl] -1-methoxy-3- [2- (2-methoxyethoxy) ethoxy] -1-propene (compound [4
9]) was obtained as a colorless oily substance in a yield of 173 mg and a yield of 95.8%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.19 (s, 6H), 0.99 (s, 9
H), 1.28 (s, 9H), 3.22 (s, 3H),
3.37 (s, 3H), 3.34 to 3.40
(M, 2H), 3.49 to 3.62 (m, 6
H), 3.78 (s, 2H), 6.77 to 6.83 (
m, 2H), 6.95 (d with fine)
coupling, J = 7.6 Hz, 1H),
7.19 (dd, J = 8.7 and 7.6H
z, 1H) ppm IR (liquid film); 2956, 293.
6,2864,1636,1596,1578 cm ^-1 Mass (m / z,%); 452 (M ⁺ , 55), 39
5 (11), 333 (27), 317 (45), 376
(29), 275 (100).

(Embodiment 11)

[Chemical 57]

Compound [49] 54 synthesized in Reference Example 35
mg (0.119 mmol) and 10 mg of TPP were dissolved in 20 ml of dichloromethane, and the mixture was stirred at 0 ° C. under an oxygen atmosphere. This solution was irradiated with a Na lamp (180 W) for 8 hours. The reaction solution was concentrated, applied to a silica gel column, and then eluted with dichloromethane and then with a mixed solvent of dichloromethane and ethyl acetate at a ratio of 25: 1.
-Butyl-4- [3- (t-butyldimethylsiloxy)
Phenyl] -4-methoxy-3- [2- (2-methoxyethoxy) ethoxymethyl] -1,2-dioxetane (compound [50]) was obtained as a pale yellow oily substance in 38 mg in a yield of 65.7%. It was

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.20 (s, 6H), 0.99 (s, 9
H), 1.28 (s, 9H), 2.66 to 2.77 (
m, 1H), 2.99 to 3.10 (m, 1H),
3.04 (s, 3H), 3.20 to 3.32
(M, 2H), 3.35 (s, 3H), 3.40-
3.52 (m, 4H), 3.61 (d, J = 10.3)
Hz, 1H), 3. 77 (d with fin
e coupling, J = 10.3 Hz, 1
H), 6.81-6.87 (m, 1H), 6.92-
7.30 (m, 3 H) ppm IR (liquid film); 2936, 288
8, 1604, 1586, 1256, 1104 cm ^-1 Mass (m / z,%); 452 (M ⁺ -32, 1),
266 (27), 210 (22), 209 (100),
177 (19).

(Reference Example 36)

Embedded image

Compound [19] 96 synthesized in Reference Example 12
0 mg (3.84 mmol) was dissolved in 10 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 330 mg of 60% sodium hydride (8.25 mmo
l) and 0.6 ml of ethyl iodide (7.50 mmol)
Were sequentially added, and the mixture was stirred at room temperature for 5.5 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate is applied to a silica gel column with 20: 1 hexane and ethyl acetate.
When it was poured out with the mixed solvent of 2-t-butyl-3-
1.00 g of ethoxy-1-methoxy-1- (3-methoxyphenyl) -1-propene (compound [51]),
Obtained as a colorless oil in a yield of 93.7%.

¹ HNMR (300 MHz, CDCl ₃ );
δ1.13 (t, J = 7.0 Hz, 3H), 1.
29 (s, 9H), 3.25 (s, 3H), 3.2
9 (q, J = 7.0 Hz, 2H), 3.69 (s, 2
H), 3.82 (s, 3H), 6.84 to 6.9.
0 (m, 1H), 6.94 to 6.99 (m, 2
H), 7.22 to 7.29 (m, 1H) ppm IR (liquid film); 2956, 286.
8, 1636, 1598, 1580 cm ^-1 Mass (m / z,%); 278 (M ⁺ , 31), 26
3 (8), 233 (22), 221 (100), 217
(42).

(Reference Example 37)

Embedded image

Compound [51] synthesized in Reference Example 36
00 g (3.60 mmol) and 60% sodium hydride 305 mg (7.63 mmol) in anhydrous DMF10.
The mixture was added to ml and stirred at 0 ° C. under an argon atmosphere. 0.53 ml (7.16 mmo of ethanethiol was added to this solution.
1) was added, and the mixture was stirred for 10 minutes and then heated and stirred at 120 ° C for 3 hours. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine and water, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a 7: 1 mixed solvent of hexane and ethyl acetate to give 2-t-butyl-3-ethoxy-
1- (3-hydroxyphenyl) -1-methoxy-1-
Propene (compound [52]) 492 mg, yield 51.
Obtained as a colorless oil at 8%.

¹ HNMR (300 MHz, CDCl ₃ );
δ1.13 (t, J = 7.0 Hz, 3H), 1.
28 (s, 9H), 3.24 (s, 3H), 3.2
9 (q, J = 7.0 Hz, 2H), 3.69 (s, 2
H), 4.78 to 4.83 (m, 1H), 6.8.
1 (ddd, J = 8.0, 2.6 and 0.
9Hz, 1H), 6.87 (s with fine
coupling, 1H), 6.94 (d wi)
th fine coupling, J = 7.6
Hz, 1H), 7.22 (dd, J = 8.0 and
7.6 Hz, 1 H) ppm IR (liquid film); 3320, 295
6,2872,1634,1596,1582 cm ^-1 Mass (m / z,%); 264 (M ⁺ , 30), 24
9 (6), 219 (13), 207 (100), 203
(64), 161 (51).

(Reference Example 38)

Embedded image

Compound [52] 12 synthesized in Reference Example 37
4 mg (0.470 mmol) of anhydrous DMF 1.5 ml
And was stirred at room temperature under an argon atmosphere. 69 mg (1.01 mmol) of imidazole and 137 mg (0.90 of t-butyldimethylchlorosilane) were added to this solution.
(9 mmol) was added and the mixture was stirred for 8 hours and 40 minutes. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine and water, dried over magnesium sulfate, and concentrated.
The concentrate was applied to a silica gel column and was poured out with a 1: 1 mixed solvent of hexane and dichloromethane.
-Butyl-1- [3- (t-butyldimethylsiloxy)
Phenyl] -3-ethoxy-1-methoxy-1-propene (Compound [53]) was obtained as a colorless oily substance with a yield of 152 mg, 85.6%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.20 (s, 6H), 0.99 (s, 9
H), 1.12 (t, J = 7.0 Hz, 3H), 1.2
8 (s, 9H), 3.23 (s, 3H), 3.2
6 (q, J = 7.0Hz, 2H), 3.70
(S, 2H), 6.80 (ddd, J = 8.1,
2.5 and 1.0 Hz, 1H, 6.86 (sw)
it fine coupling, 1H),
6.95 (d with fine c oupl
ing, J = 7.6 Hz, 1H), 7.19 (dd, J
= 8.1 and 7.6 Hz, 1H) ppm IR (liquid film); 2956, 293
6,2864,1634,1598,1578,126
0,1086 cm ^-1 Mass (m / z,%); 378 (M ⁺ , 31), 33
3 (13), 322 (25), 321 (100), 31
9 (15), 317 (31).

(Example 12)

[Chemical formula 61]

Compound [53] 10 synthesized in Reference Example 38
5 mg (0.278 mmol) and 4 mg of TPP were dissolved in 30 ml of dichloromethane, and the mixture was stirred under an oxygen atmosphere at 0 ° C. This solution was irradiated with a Na lamp (180 W) for 7 hours. The reaction mixture was concentrated, applied to a silica gel column and poured out with a mixed solvent of hexane and dichloromethane of 2: 1. 3-t-butyl-4- [3- (t-butyldimethylsiloxy) phenyl] -3-ethoxymethyl- 4-methoxy-1,2-dioxetane (compound [5
4]) was obtained as a pale yellow oily substance in a yield of 88 mg and a yield of 77.3%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.20 (s, 3H), 0.20 (s, 3
H), 0.81 (t, J = 7.0 Hz, 3H), 0.9
9 (s, 9H), 1.28 (s, 9H), 2.4
8 to 2.62 (m, 1 H), 2.94 (dq, J
= 9.2 and 7.0 Hz, 1H), 3. 0
4 (s, 3H), 3.52 (d, J = 10.1Hz, 1
H), 3.71 (d, J = 10.1 Hz, 1
H), 6.80 to 7.30 (m, 4H) pp m IR (liquid film); 2960, 293.
6,1604,1586,1256,1106 cm ^-1 Mass (m / z,%); 378 (M ⁺ -32,8),
321 (13), 266 (25), 208 (24), 2
09 (100), 177 (33), 149 (18).

(Reference Example 39)

Embedded image

Compound [52] 48 synthesized in Reference Example 37
3 mg (1.83 mmol) was added to 6 ml of anhydrous toluene, and the mixture was stirred at 0 ° C under an argon atmosphere. 0.31 ml (2.22 mmol) of triethylamine was added to this solution, followed by 2-chloro-1,3,2-dioxaphosphorane-2.
-Oxide (0.175 ml, 1.89 mmol) was added, and the mixture was stirred at 0 ° C for 10 minutes and then at room temperature for 50 minutes.
The reaction mixture was concentrated, diethyl ether was added, and the insoluble material was filtered off. When the filtrate was concentrated, 3- (2-t-butyl-3-ethoxy-1-methoxy-1-propene-1-
Phenyl) phenyl ethylene phosphate (compound [5
The crude product of 5]) was obtained as a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ1.15 (t, J = 7.0 Hz, 3H), 1.
28 (s, 9H), 3.24 (s, 3H), 3.3
0 (q, J = 7.0 Hz, 2H), 3.66 (s, 2
H), 4.26 to 4.60 (m, 4H), 7.1
2 to 7.38 (m, 4H) ppm

(Reference Example 40)

[Chemical formula 63]

680 mg of the crude product of the compound [55] synthesized in Reference Example 39 was added to 8 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 94 mg (1.82 mmol) of sodium cyanide (95%) was added and stirred overnight. The reaction mixture was concentrated and 28% aqueous ammonia 5
ml and 2 ml of THF were added and stirred for 1 day. The reaction mixture was concentrated, the concentrate was dissolved in water and washed with hexane.
When the aqueous layer was freeze-dried, 733 mg of a crude product of sodium ammonium 3- (2-t-butyl-3-ethoxy-1-methoxy-1-propen-1-yl) phenyl phosphate (Compound [56]) was obtained. , Was obtained as a colorless amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ1.13 (t, J = 7.0 Hz, 3H), 1.
31 (s, 9H), 3.28 (s, 3H), 3.2
9 (q, J = 7.0 Hz, 2H), 3.79 (s, 2
H), 7.03 (d with fine cou
pling, J = 7.1 Hz, 1H), 7.20
(Broad s, 1H), 7.29 (dd, J = 8.
3 and d 7.1 Hz, 1H), 7.35 (d,
J = 8.3 Hz, 1 H) ppm IR (KBr); 29
60, 2868, 1634, 1600, 1580, 12
96,1110 cm ^-1 Mass (FAB-pos, m / z,%); 389
([M + H-NH ₄ + Na] ^+, 28), 343 (2
4), 329 (23), 125 (100), 11
5 (19).

(Example 13)

[Chemical 64]

Compound [56] 10 synthesized in Reference Example 40
6 mg (0.277 mmol) and 4 mg of TPP were dissolved in 30 ml of dichloromethane, and the mixture was stirred at 0 ° C under an oxygen atmosphere. Add 8 to this solution using a Na lamp (180W).
Light irradiation was performed for an hour. The reaction mixture was concentrated, methanol was added to the concentrate, the insoluble material was filtered, and the mixture was concentrated again. The concentrate was dissolved in a mixed solvent of methanol (2 ml) and a 0.1% sodium hydrogen carbonate aqueous solution (2 ml), and filtered through a 0.45 µ polytetrafluoroethylene filter. 0.3 ml of the filtrate was subjected to HPLC using a column for preparative reversed phase C18 fractionation, and the fraction eluted with a gradient of 0.1% aqueous sodium hydrogen carbonate solution and acetonitrile was freeze-dried. Methanol was added to the obtained lyophilized product to concentrate the soluble portion, and 3-t-butyl-3-ethoxymethyl-4-methoxy-4- (3'-
Phosphoryloxy) phenyl-1,2-dioxetane
A crude purified product of the disodium salt (Compound [57]) was obtained as a colorless amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.87 (t, J = 7.0 Hz, 3H), 1.
30 (s, 9H), 2.54 to 2.68 (m, 1
H), 2.97 (dq, J = 9.0 and 7.0H
z, 1H), 3. 05 (s, 3H), 3.51
(D, J = 10.2 Hz, 1H), 3.76
(D, J = 10.2 Hz, 1H), 6.96 to 7.10
(M, 1H), 7.24 to 7.44 (m, 2
H), 7.56 to 7.68 (m, 1H) ppm Mass (FAB-pos, m / z,%); 443.
([M + Na] ⁺ , 20), 421 ([M + H] ⁺ , 2
4), 299 (22), 277 (23), 207
(17), 115 (100).

(Reference Example 41)

Embedded image

Compound [19] 31 synthesized in Reference Example 12
6 mg (1.26 mmol) was dissolved in 4 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution 180 mg (2.64 mmol) of imidazole and t-
Butyldimethylchlorosilane 286mg (1.90mm
was added and stirred for 1 hour. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine and water, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a 2: 1 mixed solvent of hexane and dichloromethane, and 2-t-butyl-3- (t-butyldimethylsiloxy) -1-methoxy-1- (3-methoxyphenyl) was obtained. -1-Propene (Compound [58]) was obtained as a colorless oily substance in a yield of 95.2% (438 mg).

¹ HNMR (300 MHz, CDCl ₃ );
δ-0.12 (s, 6H), 0.86 (s, 9)
H), 1.28 (s, 9H), 3.23 (s, 3H)
, 3.81 (s, 3H), 3.89 (s, 2H),
6.82-6.91 (m, 1H), 6.88
(S, 1H), 6.96 (d with fin
ecoupling, J = 7.5 Hz, 1H), 7.1
8 to 7.29 (m, 1H) ppm IR (liquid film); 2956, 293.
2,2860, 1638, 1596, 1580, 125
4,1046 cm ^-1 Mass (m / z,%); 364 (M ⁺ , 5), 308
(24), 307 (100), 251 (19), 233
(65), 201 (61), 177 (17)

(Reference Example 42)

[Chemical formula 66]

Compound [58] 48 synthesized in Reference Example 41
4 mg (1.33 mmol) and 60% sodium hydride 106 mg (2.65 mmol) in anhydrous DMF 5 m
In addition to 1, the mixture was stirred at 0 ° C. under an argon atmosphere. 0.19 ml (2.57 mmol) of ethanethiol was added to this solution, and the mixture was stirred for 15 minutes, and then heated and stirred at 110 ° C. for 3 hours. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine and water, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and was poured out with a mixed solvent of hexane and ethyl acetate of 10: 1. 2-t-butyl-3- (t-butyldimethylsiloxy) -1- (3-hydroxyphenyl) -1
-Methoxy-1-propene (compound [59]) was 221
It was obtained as a colorless amorphous solid in mg, yield 47.5%.

¹ HNMR (300 MHz, CDCl ₃ );
δ-0.10 (s, 6H), 0.87 (s, 9)
H), 1.27 (s, 9H), 3.24 (s, 3H)
, 3.88 (s, 2H), 4.57 to 4.67.
(M, 1H), 6.79 (ddd, J = 8.1,
2.6 and 0.9Hz, 1H), 6.85
(With swine fine coupling, 1H),
6.94 (d with fine couple
ing, J = 7.6 Hz, 1H), 7. 20 (d
d, J = 8.1 and 7.6 Hz, 1H) ppm IR (KBr); 3320, 2956, 2860, 16
42, 1598, 1254, 1048 cm ^-1 Mass (m / z,%); 350 (M ⁺ , 4), 294.
(22), 293 (100), 261 (14), 237
(19), 219 (64), 203 (18), 187
(76), 163 (22), 161 (22), 161
(18), 119 (24)

(Reference Example 43)

Embedded image

Compound [59] 27 synthesized in Reference Example 42
8 mg (0.794 mmol) was dissolved in 3 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution 115 mg (1.69 mmol) of imidazole and t
-Butyldimethylchlorosilane 228 mg (1.51 m
mol) was added and the mixture was stirred overnight. The reaction mixture was poured into saturated saline and extracted with ethyl acetate. The extract layer was washed with saturated brine and water, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and dichloromethane at a ratio of 3: 1 to give 2-t-butyl-3- (t-butyldimethylsiloxy) -1- [3-
270 mg of (t-butyldimethylsiloxy) phenyl] -1-methoxy-1-propene (compound [60]),
Obtained as a colorless oil in a yield of 73.3%.

¹ HNMR (300 MHz, CDCl ₃ );
δ-0.14 (s, 6H), 0.19 (s, 6
H), 0.85 (s, 9H), 0.98 (s, 9H)
, 1.28 (s, 9H), 3.22 (s, 3H),
3.90 (s, 2H), 6.76 to 6.83
(M, 2H), 6.96 (d with fine
coupling, J = 7.6 Hz, 1H), 7.1
7 (dd, J = 8.8 and 7.6 Hz, 1
H) ppm IR (liquid film); 2960, 293
6,2860,1640,1596,1578,125
6,1046 cm ^-1 Mass (m / z,%); 464 (M ⁺ , 5), 408
(34), 407 (100), 351 (19), 334
(20), 333 (66), 302 (21), 301
(80).

(Example 14)

[Chemical 68]

Compound [60] 80 synthesized in Reference Example 43
mg (0.172 mmol) and TPP 2 mg were dissolved in dichloromethane 20 ml, and the mixture was stirred at room temperature under an oxygen atmosphere. This solution was irradiated with a Na lamp (180 W) for 3 hours. The reaction mixture was concentrated, applied to a silica gel column, and poured out with a mixed solvent of hexane and dichloromethane at a ratio of 4: 1. 3-t-butyl-3-[(t-butyldimethylsiloxy) methyl] -4- [3- (t -Butyldimethylsiloxy) phenyl] -4-methoxy-1,2-
Dioxetane (compound [61]) 70 mg, yield 8
Obtained as a colorless oil at 1.9%.

¹ HNMR (300 MHz, CDCl ₃ );
δ-0.44 (s, 3H), -0.21 (s, 3
H), 0.20 (s, 6H), 0.73 (s, 9)
H), 0.98 (s, 9H), 1.31 (s, 9H),
3.00 (s, 3 H), 3.64 (d, J = 1
0.8Hz, 1H), 4.08 (d, J = 10.
8 Hz, 1H), 6.78 to 7.28 (m, 4H) pp
m IR (liquid film); 2960, 293.
2,2860,1602,1586,1256,108
6 cm ^-1 Mass (m / z,%); 464 (M ⁺ -32), 26
6 (28), 210 (22), 209 (100), 17
7 (19) 173 (73), 115 (23).

(Reference Example 44)

[Chemical 69]

Compound [44] 50 synthesized in Reference Example 31
4 mg (1.51 mmol) was added to 6 ml of anhydrous toluene, and the mixture was stirred at 0 ° C under an argon atmosphere. 0.25 ml (1.79 mmol) of triethylamine was added to this solution, followed by 2-chloro-1,3,2-dioxaphosphorane-2.
-Oxide (0.136 ml, 0.147 mmol) was added, and the mixture was stirred at 0 ° C for 10 minutes and then at room temperature for 3 hours. The reaction mixture was concentrated, diethyl ether was added, and the insoluble material was filtered off. When the filtrate was concentrated, 3- (2-t-butyl-1-isopropoxy-3-neopentyloxy-1-
A crude purified product of propen-1-yl) phenylethylene phosphate (compound [62]) was obtained as 664 mg of a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.90 (s, 9H), 1.12 (d, J =
6.2Hz, 6H), 1.28 (s, 9H), 2.7
7 (s, 2H), 3.51 (s, 2H), 3.74
(Hept, J = 6.2 Hz, 1H), 4.20
~ 4.57 (m, 4H), 7.11 to 7.35
(M, 4H) ppm

(Reference Example 45)

Embedded image

Compound [62] 66 synthesized in Reference Example 44
4 mg (1.51 mmol) was added to 7 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. 80 mg (1.55 mmo) of sodium cyanide (95%) was added to this solution.
1) was added and stirred overnight. The reaction mixture was concentrated, the concentrate was dissolved in water and lyophilized to give sodium 3-
(2-t-Butyl-1-isopropoxy-3-neopentyloxy-1-propen-1-yl) phenyl-2 ′
A crude product of -cyanoethyl phosphate (Compound [63]) was obtained as 730 mg as a colorless amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.93 (s, 9H), 1.17 (d, J =
6.1 Hz, 6H), 1.33 (s, 9H), 2.8
0 (t, J = 6.2 Hz, 2H), 2.81 (s,
2H), 3.64 (s, 2H), 3.87 (he
pt, J = 6.1 Hz, 1H), 4.15 (d
t, J = 7.8 and 6.2 Hz, 2H), 7.0
4 to 7.40 (m, 4H) ppm

(Reference Example 46)

Embedded image

710 mg of the crude product of compound [63] synthesized in Reference Example 45 was added to 3 ml of THF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 5 ml of 28% ammonia water was added and stirred for 2 days. The reaction mixture was concentrated, the concentrate was dissolved in water and washed with hexane. When the aqueous layer was freeze-dried, ammonium sodium 3- (2-t
A crude purified product of -butyl-1-isopropoxy-3-neopentyloxy-1-propen-1-yl) phenyl phosphate (Compound [64]) was obtained as a colorless amorphous solid (562 mg).

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.92 (s, 9H), 1.15 (d, J =
6.2Hz, 6H), 1.33 (s, 9H), 2.8
0 (s, 2H), 3.65 (s, 2H), 3.89
(Hept, J = 6.2 Hz, 1H), 7.05
(D, J = 7.5 Hz, 1 H), 7.13 (sw)
it fine coupling, 1H), 7.2
5 (dd, J = 8.2 and 7.5 Hz, 1
H), 7.37 (d with fine co)
upling, J = 8.2 Hz, 1H) ppm IR (KBr); 2956, 2868, 1627, 15
99, 1578, 1294, 1110 cm ^-1 Mass (FAB-pos, m / z,%); 459
([M + H-NH ₄ + Na] ^+, 28), 431 (2
2), 329 (100), 307 (43), 12
5 (67), 115 (35).

(Example 15)

Embedded image

Compound [64] 19 synthesized in Reference Example 46
8 mg (0.438 mmol) and TPP 4 mg were melt | dissolved in 30 ml of dichloromethane, and it stirred at 0 degreeC in oxygen atmosphere. Add 4 to this solution using a Na lamp (180W).
Light irradiation was performed for an hour. The reaction mixture was concentrated, methanol was added to the concentrate, the insoluble material was filtered, and the mixture was concentrated again. The concentrate was dissolved in a mixed solvent of methanol (1 ml) and 0.1% sodium hydrogen carbonate aqueous solution (1 ml), and filtered through a 0.45 µ polytetrafluoroethylene filter. HP using a polymer-based reverse phase C18 preparative column
The fraction was subjected to LC, and the fraction eluted with a gradient of 0.1% aqueous sodium hydrogen carbonate solution and acetonitrile was freeze-dried. The obtained lyophilized product was dissolved in water and subjected to HPLC using a polymer-based reverse phase C18 preparative column, and the fraction desalted with a gradient of water and acetonitrile was lyophilized to give 3-t-butyl. -4-isopropoxy-
80 mg of 3-neopentyloxymethyl-4- (3'-phosphoryloxy) phenyl-1,2-dioxetane disodium salt (Compound [65]), yield 37.4%
Obtained as an amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ 0.79 (s, 9H), 1.00 to 1.13
(M, 3H), 1.21 (d, J = 6.1Hz, 3
H), 1.37 (s, 9H), 2.24 to 2.37.
(M, 1H), 2.60 (d, J = 8.2 Hz,
1H), 3.23 to 3.40 (m, 1H), 3.
50-3.70 (m, 2H), 6.82-6.87
(M, 1H), 7.20 to 7.40 (m, 1H
H), 7.52 to 7.70 (m, 2H) ppm IR (KBr); 2976, 2872, 1588, 12
70,1104 cm ^-1 Mass (FAB-pos, m / z,%); 513
([M + Na] ⁺ , 17), 491 ([M + H] ⁺ , 3
7), 429 (50), 407 (29), 327
(52), 305 (100), 263 (38), 125
(65), 115 (49)

(Reference Example 47)

Embedded image

Compound [32] 82 synthesized in Reference Example 21
4 mg (2.53 mmol) was dissolved in 10 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution, 202 mg of 60% sodium hydride (5.05 mmo
l), 2-methoxyethyl bromide 0.48 ml
(5.11 mmol) and tetrabutylammonium bromide 73 mg (0.226 mmol) were added, and 1
The mixture was heated and stirred at 00 ° C for 5 hours. To this solution, 220 mg (5.50 mmol) of 60% sodium hydride and 0.50 ml (5.32 mmo) of 2-methoxyethyl bromide.
l) and tetrabutylammonium bromide 81m
g (0.310 mmol) was added and the mixture was added at
The mixture was heated and stirred for an hour. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine, dried over magnesium sulfate, and concentrated. The concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate of 10: 1 to give 1- (3-benzyloxyphenyl) -2-
630 mg of t-butyl-1-methoxy-3- (2-methoxyethoxy) -1-propene (compound [66]),
The yield was 64.9%.

Mp: 48.0 to 49.0 ° C. (colorless columnar crystals, recrystallized from methanol) ¹ HNMR (300 MHz, CDCl ₃ ); δ1.29
(S, 9H), 3.22 (s, 3H), 3.30
(S, 3H), 3.34 to 3.40 (m, 2
H), 3.42 to 3.48 (m, 2H), 3.78.
(S, 2H), 5.08 (s, 2H), 6.92
~ 7.00 (m, 2H), 7.00 to 7.04
(M, 1H), 7.22 to 7.48 (m, 6H) ppm IR (KBr); 2952, 2928, 2900, 16
28, 1596, 1582 cm ^-1 Mass (m / z,%); 384 (M ⁺ , 27), 32
7 (10), 309 (11), 293 (21), 251
(55), 161 (13), 91 (100).

(Reference Example 48)

[Chemical 74]

Compound [66] 34 synthesized in Reference Example 47
1 mg (0.888 mmol) and 10% Pd-C,
30 mg was added to 4.5 ml of a 2: 1 mixed solvent of ethyl acetate and methanol, and the mixture was stirred under a hydrogen atmosphere at room temperature for 5 hours. The reaction mixture was filtered through Celite, and the filtrate was concentrated. The concentrate was applied to a silica gel column and was poured out with a mixed solvent of hexane and ethyl acetate at a ratio of 5: 1 to give 2-t-butyl-1- (3-hydroxyphenyl) -1-methoxy-3- (2-methoxyethoxy). ) -1-Propene (Compound [67]) was obtained as a colorless oil in a yield of 236 mg and 90.4%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 1.27 (s, 9H), 3.34 (s, 3
H), 3.41 to 3.47 (m, 2H), 3.48 (
s, 3H), 3.59 to 3.65 (m, 2H),
3.64 (broads, 2H), 6.77
(S with fine coupling, 1
H), 6.83 (ddd, J = 8.0, 2.6 an
d 0.9 Hz, 1 H), 6.89 (d wit
h fine coupling, J = 7. 7H
z, 1H), 7.22 (dd, J = 8.0 and
7.7 Hz, 1 H), 7.41 (broad
s, 1H) ppm IR (liquid film); 3384,295
2, 2836, 1634, 1596, 1582 cm ^-1 Mass (m / z,%); 294 (M ⁺ , 32), 23.
7 (24), 219 (20), 218 (24), 203
(87) 162 (26), 161 (100).

(Reference Example 49)

[Chemical 75]

Compound [67] 10 synthesized in Reference Example 48
5 mg (0.357 mmol) was dissolved in 2 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. To this solution was added 52 mg (0.764 mmol) of imidazole and t
-Butyldimethylchlorosilane 98 mg (0.65 mm
ol) was added and the mixture was stirred for 5 hours. The reaction mixture was poured into water and extracted with ethyl acetate. The extract layer was washed with saturated brine and water, dried over magnesium sulfate, and concentrated. When the concentrate was applied to a silica gel column and poured out with a mixed solvent of hexane and ethyl acetate of 20: 1, 2-t-butyl-1-
[3- (t-butyldimethylsiloxy) phenyl] -1
124 mg of -methoxy-3- (2-methoxyethoxy) -1-propene (compound [68]), yield 85.1%
And was obtained as a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.19 (s, 6H), 0.99 (s, 9
H), 1.28 (s, 9H), 3.22 (s, 3H),
3.33 (s, 3H), 3.31 to 3.37
(M, 2H), 3.41 to 3.47 (m, 2
H), 3.80 (s, 2H), 6.77 to 6.84 (
m, 1H). 6.81 (d, J = 1.4Hz, 1
H), 6.94 (d w it fine cou
pling, J = 7.6 Hz, 1H), 7.19
(Dd, J = 8.7 and 7.6 Hz, 1H) pp
m IR (liquid film); 2956, 293.
6, 1636, 1596, 1578, 1260 cm ^-1 Mass (m / z,%); 408 (M ⁺ , 33), 35.
1 (12), 333 (22), 317 (47), 276
(29), 275 (100), 249 (29), 219
(35), 179 (27), 121 (40).

(Example 16)

[Chemical 76]

Compound [68] 55 synthesized in Reference Example 49
mg (0.134 mmol) and 4 mg of TPP were dissolved in 20 ml of dichloromethane, and the mixture was stirred under an oxygen atmosphere at room temperature. This solution was irradiated with a Na lamp (180 W) for 6.5 hours. The reaction mixture was concentrated, applied to a silica gel column, and then eluted with dichloromethane and a mixed solvent of dichloromethane and ethyl acetate at a ratio of 50: 1.
Butyl-4- [3- (t-butyldimethylsiloxy) phenyl] -4-methoxy-3- (2-methoxyethoxy) methyl-1,2-dioxetane (Compound [69])
Was obtained as a yellow oily substance in a yield of 42 mg and a yield of 70.8%.

¹ HNMR (300 MHz, CDCl ₃ );
δ 0.20 (s, 6H), 0.99 (s, 9
H), 1.28 (s, 9H), 2.69 (dt, J = 1)
0.5 and 4.7 Hz, 1H), 3.02
(Dt, J = 10.5 and 5.3 Hz, 1
H), 3.04 (s, 3H), 3.16 (dd,
J = 5.3 and 4.7 Hz, 2H), 3.23
(S, 3H), 3. 61 (d, J = 10.3H
z, 1H), 3.77 (d, J = 10.3H z,
1H), 6.85 (d with fine coup
ling, J = 8.0 Hz, 1H), 6.90-
7.20 (m, 2H), 7.26 (dd, J =
8.0 and 7.6 Hz, 1H) ppm IR (liquid film); 2960,293
6,1602,1586,1256,1108 cm ^-1 Mass (m / z,%); 408 (M ⁺ -32,3),
266 (27), 210 (22), 209 (100),
177 (19), 89 (21).

(Reference Example 50)

Embedded image

Compound [67] 41 synthesized in Reference Example 48
5 mg (1.41 mmol) was added to 6 ml of anhydrous toluene, and the mixture was stirred at 0 ° C under an argon atmosphere. 0.24 ml (1.72 mmol) of triethylamine was added to this solution, followed by 2-chloro-1,3,2-dioxaphosphorane-2.
-Oxide (0.13 ml, 0.141 mmol) was added, and the mixture was stirred at 0 ° C for 20 minutes and then at room temperature for 1 hour and 20 minutes. The reaction mixture was concentrated, diethyl ether was added, and the insoluble material was filtered off. When the filtrate was concentrated, 3- [2-t
A crude product of -butyl-1-methoxy-3- (2-methoxyethoxy) -1-propen-1-yl] phenylethylene phosphate (Compound [70]) was obtained as a colorless oil.

¹ HNMR (300 MHz, CDCl ₃ );
δ 1.28 (s, 9H), 3.24 (s, 3
H), 3.35 (s, 3H), 3.32 to 3.52 (
m, 4H), 3.37 (s, 2H), 4.30 ~.
4.60 (m, 4H), 7.12 to 7.39
(M, 4H) ppm

(Reference Example 51)

Embedded image

560 mg of the crude product of the compound [70] synthesized in Reference Example 50 was added to 8 ml of anhydrous DMF, and the mixture was stirred at room temperature under an argon atmosphere. 73 mg (1.42 mmol) of sodium cyanide (95%) was added to this solution, and the mixture was stirred overnight. The reaction mixture was concentrated and 28% aqueous ammonia 4
ml was added and stirred for 1 day. The reaction mixture was concentrated, the concentrate was added to water and washed with hexane. When the aqueous layer was freeze-dried, sodium ammonium 3- [2-t-butyl-1-methoxy-3- (2-methoxyethoxy) -1-
A crude purified product of propen-1-yl] phenyl phosphate (Compound [71]) was obtained as 525 mg as a colorless amorphous solid.

¹ HNMR (300 MHz, CD ₃ OD);
δ1.32 (s, 9H), 3.29 (s, 3
H), 3.36 (s, 3H), 3.35-3.42 (
m, 2H), 3.45 to 3.51 (m, 2H),
3.84 (s, 2H), 7.05 (d with
fine coupling, J = 6.8H
z, 1H), 7.22 (broads, 1H), 7.
26-7.37 (m, 2H) ppm IR (KBr); 2956, 1636, 1600, 15
78,1292,1218 cm ^-1 Mass (FAB-pos, m / z,%); 419
([M + H-NH ₄ + Na] ^+, 84), 343 (3
7), 329 (26), 321 (48), 125
(100).

(Example 17)

Embedded image

Compound [71] 15 synthesized in Reference Example 51
1 mg (0.366 mmol) and 4 mg of TPP were dissolved in 30 ml of dichloromethane, and the mixture was stirred at 0 ° C under an oxygen atmosphere. Add 7 to this solution using a Na lamp (180W).
Light irradiation was performed for an hour. The reaction mixture was concentrated, methanol was added to the concentrate, the insoluble material was filtered, and the mixture was concentrated again. The concentrate was dissolved in a mixed solvent of methanol (3 ml) and 0.1% aqueous sodium hydrogen carbonate solution (3 ml), and filtered through a 0.45 µ polytetrafluoroethylene filter.
HPLC using a polymer-based reverse phase C18 preparative column
Then, the fraction eluted with a gradient of 0.1% aqueous sodium hydrogen carbonate solution and acetonitrile was freeze-dried. The obtained lyophilized product was dissolved in water and subjected to HPLC using a polymer-based reverse phase C18 preparative column, and the fraction desalted with a gradient of water and acetonitrile was lyophilized to give 3-t-butyl. -4-methoxy-3-
[(2-Methoxyethoxy) methyl] -4- (3'-phosphoryloxy) phenyl-1,2-dioxetane disodium salt (Compound [72]) 43 mg, yield 2
Obtained as an amorphous solid at 6.0%.

¹ HNMR (300 MHz, CD ₃ OD);
δ1.31 (s, 9H), 2.70 to 2.79.
(M, 1H), 2.96 to 3.08 (m, 1H),
3.06 (s, 3H), 3.20 to 3.30 (m, 2
H), 3.28 (s, 3H), 3.61 (d, J
= 10.4 Hz, 1H), 3.81 (d, J = 1)
0.4 Hz, 1 H), 6.98 to 7.14 (m, 1
H), 7.26 to 7.42 (m, 2H), 7.5
6 to 7.68 (m, 1H) ppm IR (KBr); 1605, 1585, 1281, 11
12 cm ^-1 Mass (FAB-pos, m / z,%); 473
([M + Na] ⁺ , 18), 451 ([M + H] ⁺ , 1
0), 401 (60), 299 (100), 27
7 (56), 125 (56), 115 (28).

* Test Example 1 * 3-t-butyl-4-methoxy-3 obtained in Example 6
-Neopentyloxymethyl-4- (3'-phosphoryloxy) phenyl-1,2-dioxetane disodium salt (compound [26]) was added to a quaternary ammonium salt BDMQ0 at a concentration of 0.2 mg / ml. 0.4 mg /
ml, dissolved in 0.1 M diethanolamine-hydrochloric acid buffer solution (pH 10.0) containing 1 mM magnesium chloride and 0.05% sodium azide, and after stirring, 300 μl of this solution was placed in an assay cartridge and incubated. After 90 minutes incubation, EI
Alkaline phosphatase solution for A (Boehringer Mannheim KK) (3 mg / 0.3 ml) was added to 0.15M.
Sodium chloride, 1 mM magnesium chloride, 0.1 mM
50 mM containing zinc chloride and 0.1% sodium azide
20 μl of an enzyme solution prepared by diluting 154 times with Tris / Cl buffer (pH 7.2) was added, and after stirring, the amount of luminescence was measured at 37 ° C. over time. For comparison, the amount of luminescence of commercially available AMPPD was measured under the same conditions. The result is shown in FIG.

* Test Example 2 * 3-t-butyl-4-methoxy-3-neopentyloxymethyl-4- (3'-phosphoryloxy) phenyl-1,2-dioxetane disodium obtained in Example 6 1 mg of salt (compound [26]) was added to methanol d
_It was dissolved in ₄ (0.35 ml) and heated in a constant temperature bath at 60 ° C. ¹ H NMR was measured every 2-3 hours. As a result, 3-t-butyl-4-methoxy-3-neopentyloxymethyl-4- (3′-phosphoryloxy) phenyl-1,2-dioxetane disodium salt (compound [26]) at 60 ° C. The half-life was estimated to be 18.6 hours.

Commercially available AMPPD (3- (2'-spiroadamantane) -4-methoxy-4- (3 "-phosphoryloxy) phenyl-1,2-dioxetane disodium salt) was also measured in the same manner. The half-life at was estimated to be 5.5 hours.

[0230]

INDUSTRIAL APPLICABILITY The 1,2-dioxetane derivative of the present invention is excellent in thermal stability, has a short time after the start of light emission until the amount of light emission reaches the maximum value, and has a high maximum light emission amount. It has characteristics. That is, it is possible to save the trouble such as necessary adjustment and temperature control when storing. Since the maximum amount of light emission is reached in a short time after the start of light emission and a high amount of light emission is obtained, measurement can be performed in a short time, and application to a high sensitivity analysis system is easy. Furthermore, the molecule is designed so that it can be easily bound to amino acids or peptides, and it can be easily applied to labeled substances and the like.

[Brief description of drawings]

FIG. 1 shows 3-t-butyl-4-methoxy-3-neopentyloxymethyl-4- (3′-phosphoryloxy) phenyl-1,2-dioxetane disodium salt (compound [26]) and alkaline phosphite. It is a figure which shows the relationship between the light emission intensity and time at the time of making it light-emit using tase. The results of AMPPD are also shown for comparison.

Claims (3)

[Claims] 1. A compound of the general formula The 1,2-dioxetane derivative represented by the formula (wherein R ¹ is a hydrogen atom, an alkyl group or —Si (R ⁹ R ¹⁰ R ¹¹ ), and R ² , R ³ , R ⁵ and R ⁶ are hydrogen atoms. Or an alkyl group, R ⁴ is an alkyl group, a hydroxyl group, an alkoxyl group or —OSi (R ⁹ R ¹⁰ R ¹¹ ), R ⁷ is an alkyl group, Ar is unsubstituted or substituted with —R ⁸ . An aryl group, R ⁸ is an alkoxyl group, —OSi (R ⁹ R ¹⁰ R ¹¹ ).
Alternatively, it is a phosphate group. R ⁹ , R ¹⁰ and R ¹¹ are alkyl groups. ). 2. A general formula: The 1,2-dioxetane derivative according to claim 1, represented by the formula (wherein R ^{1 to} R ⁸ are the same as above). 3. A general formula: The 1,2-dioxetane derivative according to claim 1, represented by the formula (wherein R ^{1 to} R ⁸ are the same as above).