Method for producing hetero-type monodisperse polyethylene glycol derivative
12030833 · 2024-07-09
Assignee
Inventors
Cpc classification
C07C213/06
CHEMISTRY; METALLURGY
C07C43/11
CHEMISTRY; METALLURGY
C07C67/31
CHEMISTRY; METALLURGY
C07C41/16
CHEMISTRY; METALLURGY
C07C43/11
CHEMISTRY; METALLURGY
C07C213/10
CHEMISTRY; METALLURGY
C07C217/08
CHEMISTRY; METALLURGY
C07C213/02
CHEMISTRY; METALLURGY
C07C67/31
CHEMISTRY; METALLURGY
C07C213/02
CHEMISTRY; METALLURGY
C07C217/08
CHEMISTRY; METALLURGY
C07C41/16
CHEMISTRY; METALLURGY
International classification
C07C213/06
CHEMISTRY; METALLURGY
C07C213/10
CHEMISTRY; METALLURGY
C08G65/30
CHEMISTRY; METALLURGY
C08G65/332
CHEMISTRY; METALLURGY
Abstract
A method for producing a hetero-type monodisperse polyethylene glycol, which includes (A) carrying out a nucleophilic substitution reaction between a compound of formula (2) and a compound of formula (3) so as to satisfy expression (F1) to obtain a compound of formula (4), (B) of carrying out a Michaels addition reaction of a compound of formula (5) to the compound of formula (4) at a temperature condition of 10? C. or lower to obtain a compound of formula (6), (C) detritylating or debenzylating the compound of the formula (6) to obtain a reaction product containing a compound of formula (7), (D) purifying the compound of formula (7) from the reaction product, (E) reacting the compound of formula (7) with phthalimide and performing dephthalimidation to obtain a compound of formula (8), and (F) subjecting the compound of formula (8) to an acid hydrolysis treatment to obtain a compound represented by formula (1).
Claims
1. A method for producing a hetero-type monodisperse polyethylene glycol represented by the formula (1), which comprises the following steps (A), (B), (C), (D), (E) and (F): ##STR00070## wherein, in the formula (1), a represents an integer of 6 to 12, Step (A): a step of carrying out a nucleophilic substitution reaction between a compound of the formula (2) and a compound of the formula (3) so as to satisfy the requirement of the expression (F1) to obtain a compound of the formula (4): ##STR00071## wherein, in the formula (2), b represents an integer of 3 to 9, ##STR00072## wherein, in the formula (3), L represents a mesyl group or a tosyl group, R.sup.1 represents a trityl group or a benzyl group, and c represents an integer of 3 to 9, ##STR00073## wherein, in the formula (4), R.sup.1 represents a trityl group or a benzyl group and a represents an integer of 6 to 12; Step (B): a step of carrying out Michael addition reaction of a compound of the formula (5) to the compound of the formula (4) obtained in the step (A) under a temperature condition of 10? C. or lower to obtain a compound of the formula (6), ##STR00074## wherein, in the formula (5), R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms, ##STR00075## wherein, in the formula (6), R.sup.1 represents a trityl group or a benzyl group, R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms, and a represents an integer of 6 to 12; Step (C): a step of detritylating or debenzylating the compound of the formula (6) obtained in the step (B) to obtain a reaction product containing a compound of the formula (7), ##STR00076## wherein, in the formula (7), R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms and a represents an integer of 6 to 12; Step (D): a step of purifying the compound of the formula (7) from the reaction product obtained in the step (C); Step (E): a step of reacting the compound of the formula (7) obtained in the step (D) with phthalimide and performing dephthalimidation to obtain a compound of the formula (8), ##STR00077## wherein, in the formula (8), R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms and a represents an integer of 6 to 12; and Step (F): a step of subjecting the compound of the formula (8) obtained in the step (E) to an acid hydrolysis treatment to obtain the compound represented by the formula (1).
2. The method according to claim 1, wherein the compound of the formula (7) is purified using dichloromethane or chloroform in the step (D).
3. The method according to claim 1, wherein the compound of the formula (7) is purified using water or an aqueous solution having a concentration of an inorganic salt of 10% by weight or less in the step (D).
4. The method according to claim 1, wherein the step (D) comprises a washing step at a temperature of 1 to 25? C.
5. A method for producing a hetero-type monodisperse polyethylene glycol represented by the formula (1), which comprises the following steps (A), (B), (C), (E), (F) and (G): ##STR00078## wherein, in the formula (1), a represents an integer of 13 to 40, Step (A): a step of carrying out a nucleophilic substitution reaction between a compound of the formula (2) and a compound of the formula (3) so as to satisfy the requirement of the expression (F1) to obtain a compound of the formula (4): ##STR00079## wherein, in the formula (2), b represents an integer of 3 to 37, ##STR00080## wherein, in the formula (3), L represents a mesyl group or a tosyl group, R.sup.1 represents a trityl group or a benzyl group, and c represents an integer of 3 to 37, ##STR00081## wherein, in the formula (4), R.sup.1 represents a trityl group or a benzyl group and a represents an integer of 13 to 40; Step (B): a step of carrying out Michael addition reaction of a compound of the formula (5) to the compound of the formula (4) obtained in the step (A) under a temperature condition of 10? C. or lower to obtain a compound of the formula (6), ##STR00082## wherein, in the formula (5), R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms, ##STR00083## wherein, in the formula (6), R.sup.1 represents a trityl group or a benzyl group, R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms, and a represents an integer of 13 to 40; Step (C): a step of detritylating or debenzylating the compound obtained in the step (B) to obtain a compound of the formula (7), ##STR00084## wherein, in the formula (7), R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms and a represents an integer of 13 to 40; Step (E): a step of reacting the compound of the formula (7) obtained in the step (C) with phthalimide and performing dephthalimidation to obtain a compound of the formula (8), ##STR00085## wherein, in the formula (8), R.sup.2 represents a hydrocarbon group having 1 to 6 carbon atoms and a represents an integer of 13 to 40; Step (F): a step of subjecting the compound of the formula (8) obtained in the step (E) to an acid hydrolysis treatment to obtain a reaction product containing the compound of the formula (1); and Step (G): a step of purifying the compound of the formula (1) from the reaction product obtained in the step (F).
6. The method according to claim 5, wherein an organic solvent to be used for the purifying is dichloromethane or chloroform in the step (G).
7. The method according to claim 5, wherein an aqueous solution to be used for the purifying is a basic aqueous solution of pH 8 or higher in the step (G).
8. The method according to claim 1, wherein R.sup.2 of the compound of the formula (5) is an isopropyl group or a tert-butyl group in the step (B).
9. The method according to claim 1, wherein flaky potassium hydroxide or powdery potassium hydroxide is used as a base in the step (B).
10. The method according to claim 2, wherein the compound of the formula (7) is purified using water or an aqueous solution having a concentration of an inorganic salt of 10% by weight or less in the step (D).
11. The method according to claim 2, wherein the step (D) comprises a washing step at a temperature of 1 to 25? C.
12. The method according to claim 3, wherein the step (D) comprises a washing step at a temperature of 1 to 25? C.
13. The method according to claim 6, wherein an aqueous solution to be used for the purifying is a basic aqueous solution of pH 8 or higher in the step (G).
14. The method according to claim 2, wherein R.sup.2 of the compound of the formula (5) is an isopropyl group or a tert-butyl group in the step (B).
15. The method according to claim 3, wherein R.sup.2 of the compound of the formula (5) is an isopropyl group or a tert-butyl group in the step (B).
16. The method according to claim 4, wherein R.sup.2 of the compound of the formula (5) is an isopropyl group or a tert-butyl group in the step (B).
17. The method according to claim 5, wherein R.sup.2 of the compound of the formula (5) is an isopropyl group or a tert-butyl group in the step (B).
18. The method according to claim 2, wherein flaky potassium hydroxide or powdery potassium hydroxide is used as a base in the step (B).
19. The method according to claim 3, wherein flaky potassium hydroxide or powdery potassium hydroxide is used as a base in the step (B).
20. The method according to claim 4, wherein flaky potassium hydroxide or powdery potassium hydroxide is used as a base in the step (B).
Description
EXAMPLES
(1) Hereinafter, the present invention will be described in more detail with reference to Examples, but the present invention is not limited to the following Examples.
(2) For the measurement of the monodisperse polyethylene glycol obtained in the present invention, JNM-ECP400 or JNM-ECP600 manufactured by JEOL Datum Co., Ltd. was used in .sup.1H-NMR analysis. A 5 mm? tube was used for the measurement, CDCl.sub.3 or CDOD.sub.3 was used as deuterated solvents, and tetramethylsilane (TMS) was used as an internal standard substance.
(3) The monodisperse polyethylene glycol having one-terminal trityl group which is obtained by the production method of the step (A) and is a compound represented by the above formula (4) (R.sup.1 is a trityl group), in which a is 8 or 12, contains a monodisperse polyethylene glycol impurities having both-terminal trityl group which is a compound represented by the formula (9). Although it is difficult to accurately quantify the content, the hydroxyl group of the monodisperse polyethylene glycol having one-terminal trityl group is reacted with methanesulfonyl chloride, and from .sup.1H-NMR measurement results of the resulting compound, the content of the monodisperse polyethylene glycol impurities having both-terminal trityl group contained in each step was estimated.
(4) In the reaction in each Example, since the exact number of moles of the monodisperse polyethylene glycol having one-terminal trityl group is unclear, a reagent equivalent was calculated on the assumption that all the amount was derived from the monodisperse polyethylene glycol having one-terminal trityl group.
Example 1
(5) Synthesis of compound 10 in which a in formula (1) is 8
(6) ##STR00022##
Example 1-1
(7) Synthesis of compound 4 in which a is 4, R.sup.1 is trityl group, and L is mesyl group in formula (3)
(8) ##STR00023##
(9) Tetraethylene glycol 1 (3416 g, 17.6 mol) and toluene (3.7 L) were added to a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer and the whole was dissolved under a nitrogen atmosphere, followed by azeotropic dehydration at 110 to 120? C. After the azeotropic dehydration, the mixture was cooled, triethylamine (736 ml, 5.3 mol), DMAP (54 g, 0.44 mol) and trityl chloride (TrtCl, 1226 g, 4.4 mol) were added, and the mixture was stirred at room temperature for 3 hours. After 3 hours, the disappearance of TrtCl was confirmed using thin layer chromatography (TLC), a 5% aqueous sodium dihydrogen phosphate solution (6.1 L) was added, and liquid separation was performed. The organic layer was washed once with a 5% aqueous sodium dihydrogen phosphate solution (6.1 L), once with a saturated aqueous sodium hydrogen carbonate solution (6.1 L), and once with a saturated aqueous sodium chloride solution (6.1 L). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 2 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compound 3.
(10) Compound 2
(11) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm):
(12) 2.4 (1H, t, C(OCH.sub.2CH.sub.2).sub.4OH),
(13) 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3)
(14) 3.45-3.85 (14H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.3OH, including one derived from compound 3),
(15) 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3)
(16) Yield: 1687 g
(17) ##STR00024##
(18) The reaction product containing the compound 2 (compound 2: 1672 g, less than 3.83 mol) and toluene (8.4 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (643 ml, 4.62 mol). Methanesulfonyl chloride (326 mL, 4.22 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 2 was confirmed by TLC analysis, a 5% aqueous sodium dihydrogen phosphate solution (8.4 L) was added, and liquid separation was performed. The organic layer was washed once with a 5% aqueous sodium dihydrogen phosphate solution (8.4 L), once with a saturated aqueous sodium hydrogen carbonate solution (8.4 L), and once with a saturated aqueous sodium chloride solution (8.4 L). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 4 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compound 3.
(19) Compound 4
(20) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm):
(21) 2.98 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3),
(22) 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3)
(23) 3.45-3.85 (12H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.2OCH.sub.2CH.sub.2, including one derived from compound 3),
(24) 4.33 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3),
(25) 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3)
(26) Yield: 1942 g
(27) From the .sup.1H-NMR measurement results of the compound 4 of Example 1-1, it was confirmed that the compound 3 was contained in an amount of about 6.2 mol %.
(28) A calculation expression of the compound 3 content on the basis of a ? 3.23 peak is expressed by the following expression.
(((2?[? 4.32])/4H)/([? 4.32]/2H))?100 (mol %)
(29) Further, the reaction product 2 obtained in Example 1-1 contains the compound 3 in an amount of about 8.8 wt %.
(Example 1-2, Step (A))
(30) Synthesis of Compound 5 in which a is 8 and R.sup.1 is Trityl Group in Formula (4)
(31) ##STR00025##
(32) Sodium hydride (215 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and after nitrogen substitution, MeCN (3.9 L) was added and the mixture was cooled to 0? C. MeCN (2.1 L) was mixed with tetraethylene glycol 1 (3660 g, 18.8 mol) azeotropically dehydrated with toluene (1.8 L), and this mixed solution was added dropwise over 2 hours. After completion of the dropwise addition, MeCN (2.1 L) was mixed with the reaction product containing the compound 4 (compound 4: 1942 g, less than 3.77 mol), and the mixed solution was added dropwise over 20 minutes. After completion of the dropwise addition, the reaction mixture was heated to 75? C. and stirred for 3 hours. After 3 hours, it was confirmed using TLC that the compound 4 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. A saturated aqueous ammonium chloride solution (3.9 L) and hexane (3 L) were added to the reaction mixture solution and liquid separation was performed. The lower layer from which the hexane layer (upper layer) had been removed was concentrated under reduced pressure, and toluene (9.7 L) was added to the residue. This toluene solution was washed once with a saturated aqueous ammonium chloride solution (5.8 L) and three times with a saturated aqueous sodium chloride solution (9.7 L). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 5 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction products contained the above compounds 3 and 6.
(33) Compound 4
(34) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm):
(35) 2.52 (1H, t, C(OCH.sub.2CH.sub.2).sub.8OH),
(36) 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6)
(37) 3.45-3.85 (30H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.7OH, including those derived from compounds 3 and 6),
(38) 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6)
(39) Yield: 2195 g
(Example 1-3, Step (B))
(40) Synthesis of Compound 7 in which a is 8, R.sup.1 is Trityl Group, and R.sup.2 is Tert-Butyl Group in Formula (6)
(41) ##STR00026##
(42) The reaction product containing the compound 5 (compound 5: 2190 g, less than 3.58 mol) and dichloromethane (11 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the whole was dissolved under a nitrogen atmosphere, followed by addition of powdery potassium hydroxide (21 g, 0.37 mol). After cooling to 5? C., tert-butyl acrylate (a compound in which R.sup.2 in the formula (5) is a tert-butyl group, 780 mL, 5.38 mol) was added dropwise, and the mixture was reacted at 5? C. for 1 hour. After the reaction, a saturated aqueous ammonium chloride solution (4.0 L) was added and liquid separation was performed. The organic layer was washed once with a saturated aqueous sodium chloride solution (5.5 L). The organic layer was concentrated under reduced pressure to obtain a reaction product containing the compound 4 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction products contained the above compounds 3 and 6.
(43) Compound 7
(44) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.49 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 3.45-3.85 (32H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) Yield: 2588 g (Yield: 98%)
(Example 1-4, Steps (C), (D))
(45) Synthesis of Compound 8 in which a is 8 and R.sup.2 is a Tert-Butyl Group in Formula (7)
(46) ##STR00027##
(47) A reaction product containing the compound 7 (compound 7: 2588 g, less than 3.49 mol) and methanol (12.9 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the whole was dissolved under a nitrogen atmosphere. Thereafter, p-toluenesulfonic acid monohydrate (338 g, 1.78 mol) and hexane (10 L) were added. After stirring at room temperature for 30 minutes, the hexane layer was removed, hexane (6.5 L) was added again, and the mixture was stirred for 30 minutes. After performing the same operation 5 times, the disappearance of the compounds 7, 3 and 6 was confirmed as a result of .sup.1H-NMR measurement, and a saturated aqueous sodium hydrogen carbonate solution (5.2 L) was added. The mixed solution was washed once with hexane (6.5 L) to remove trityl methyl ether. The methanol solution was concentrated under reduced pressure to obtain a crude product containing the compound 8. It was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained crude product contained the above compounds 1 and 9.
(48) Next, dichloromethane (12.9 L) was added to the crude product, and the mixture was washed three times with ion-exchanged water (12.9 L) and once with a saturated aqueous sodium chloride solution (12.9 L) under the condition of 10? C. From TLC analysis, the disappearance of the compounds 1 and 9 was confirmed. Magnesium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a purified product containing the compound 8 as a pale yellow transparent liquid.
(49) Compound 8
(50) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.6 (1H, t, H(OCH.sub.2CH.sub.2).sub.8OCH.sub.2CH.sub.2), 3.45-3.85 (34H, m, H(OCH.sub.2CH.sub.2).sub.8OCH.sub.2CH.sub.2) Yield: 1229 g (yield: 71%)
(Example 1-5, Step (E))
(51) Synthesis of Compound 9 in which a is 8 and R.sup.2 is Tert-Butyl Group in Formula (8)
(52) ##STR00028##
(53) The compound 8 (1229 g, 2.46 mol) and dichloromethane (4.9 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the whole was dissolved under a nitrogen atmosphere. Then, phthalimide (508 g, 3.45 mol), triphenylphosphine (906 g, 3.45 mol) were added. After stirring at room temperature for 30 minutes, diisopropyl azodicarboxylate (599 g, 2.96 mol) diluted with dichloromethane (1.2 L) was added dropwise, and the mixture was stirred for 1 hour. After 1 hour, the disappearance of the compound 8 was confirmed by TLC, the solvent was distilled off under reduced pressure, methanol (5.4 L) and ethylenediamine monohydrate (2 L, 24.7 mol) were added, and a reaction was carried out at 40? C. for 2 hours. A 6N aqueous hydrochloric acid solution was added thereto for neutralization, the solvent was distilled off under reduced pressure, sodium chloride and chloroform (6.1 L) were added for extraction, and a saturated aqueous sodium chloride solution (6.2 L) was added to this extract solution, followed by washing twice. A 1% aqueous sodium dihydrogen phosphate solution (6.2 L) was added to the organic layer, and the mixture was extracted three times. A mixed solvent of chloroform (2.2 L) and toluene (4 L) was added to the extracted aqueous solution, and the mixture was washed 7 times. After a 5N aqueous sodium hydroxide solution was added thereto for neutralization, sodium chloride was added, and extraction was performed four times with dichloromethane (6.2 L). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 9 as a pale yellow transparent liquid.
(54) Compound 9
(55) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.1 (2H, t, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.7), 3.45-3.85 (32H, m, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.7CH.sub.2CH.sub.2) Yield: 1149 g (yield: 94%)
(Example 1-6, Step (F))
(56) Synthesis of Compound 10 in which a in Formula (1) is 8
(57) ##STR00029##
(58) The compound 9 (850 g, 1.71 mol) and 1 M hydrochloric acid (4.2 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the compound was dissolved, followed by stirring at 50 to 55? C. for 1 hour. After cooling to 15? C., the mixture was neutralized with a 10 M aqueous sodium hydroxide solution, and sodium chloride and dichloromethane (4.3 L) were added, followed by washing. Further, this aqueous layer was washed twice with dichloromethane (4.3 L), adjusted to pH 2 with 6M hydrochloric acid, and concentrated under reduced pressure. Dichloromethane (2.6 L) and ethanol (2.2 L) were added, and the mixture was stirred and filtered, followed by concentration under reduced pressure. Then, after the operation of adding dichloromethane (2.6 L) and concentrating under reduced pressure was repeated three times, dichloromethane (2.6 L) was added to dissolve the resultant, and the solution was filtered. The filtrate was concentrated under reduced pressure to obtain the compound 10 as a pale yellow transparent liquid.
(59) Compound 10
(60) .sup.1H-NMR (CD.sub.3OD internal standard TMS); ? (ppm): 2.56 (2H, t, CH.sub.2CH.sub.2COOH), 3.17 (2H, t, H.sub.2NCH.sub.2CH.sub.2O), 3.6-3.9 (32H, m, CH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.7CH.sub.2CH.sub.2) Yield: 710 g (Yield: 87%) Purity: 99.8% (HPLC-RI)
(61) The HPLC measurement conditions used for the purity measurement are shown below.
(62) Apparatus: alliance manufactured by Waters Corporation.
(63) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(64) Detector: RI
(65) Developing solvent: a solution of methanol/5 mM ammonium acetate solution=15/85
(66) Flow rate: 1.0 mL/min
(67) Column temperature: 40? C.
(68) Sample concentration: 1 mg/mL
(69) Injection volume: 50 ?L
Example 2
(70) Synthesis of Compound 18 in which a in Formula (1) is 12
(71) ##STR00030##
Example 2-1
(72) Synthesis of Compound 11 in which a is 8, R.sup.1 is Trityl Group, and L is Mesyl Group in Formula (3)
(73) ##STR00031##
(74) The reaction product containing the compound 5 (compound 5: 841 g, less than 1.37 mol) and toluene (4.2 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (267 ml, 1.92 mol). Methanesulfonyl chloride (128 mL, 1.65 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 5 was confirmed by TLC analysis, a 5% aqueous sodium dihydrogen phosphate solution (4.2 L) was added, and liquid separation was performed. The organic layer was washed once with a 5% aqueous sodium dihydrogen phosphate solution (4.2 L), once with a saturated aqueous sodium hydrogen carbonate solution (4.2 L), and once with a saturated aqueous sodium chloride solution (4.2 L). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 11 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3 and 6.
(75) Compound 11
(76) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 3.07 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) 3.45-3.85 (28H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.6OCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 4.37 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) Yield: 945 g
(77) From the .sup.1H-NMR measurement results of the compound 11 of Example 2-1, it was confirmed that the compounds 3 and 6 were contained in an amount of about 11 mol % (compound 3: 6 mol %, compound 6: 5 mol %, rough estimation).
(78) A calculation expression of the contents of the compounds 3 and 6 on the basis of a ? 3.23 peak is expressed by the following expression.
(((2?[? 4.32])/4H)/([? 4.32]/2H))?100 (mol %)
(79) With regard to the content of the compound 3, a value calculated in Example 1-1 is applied.
(80) Further, the reaction product 5 used in Example 2-1 contains the compounds 3 and 6 in an amount of about 12.6 wt %.
(Example 2-2, Step (A)) Synthesis of Compound 12 in which a is 12 and R.SUP.1 .is Trityl Group in Formula (4)
(81) ##STR00032##
(82) Sodium hydride (78 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and after nitrogen substitution, MeCN (1.9 L) was added and the mixture was cooled to 0? C. MeCN (940 mL) was mixed with tetraethylene glycol 1 (1327 g, 6.83 mol) azeotropically dehydrated with toluene (660 mL), and this mixed solution was added dropwise over 2 hours. After completion of the dropwise addition, MeCN (940 mL) was mixed with the reaction product containing the compound 11 (compound 11: 944 g, less than 1.37 mol), and the mixed solution was added dropwise over 20 minutes. After completion of the dropwise addition, the reaction mixture was heated to 75? C. and stirred for 3 hours. After 3 hours, it was confirmed using .sup.1H-NMR that the compound 11 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. A saturated aqueous ammonium chloride solution (1.9 L) and hexane (1.4 L) were added to the reaction mixture solution and liquid separation was performed. The lower layer from which the hexane layer (upper layer) had been removed was concentrated under reduced pressure, and toluene (4.7 L) was added to the residue. This toluene solution was washed once with a saturated aqueous ammonium chloride solution (2.8 L) and three times with a saturated aqueous sodium chloride solution (4.7 L). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 12 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6 and 13.
(83) Compound 12
(84) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.56 (1H, t, C(OCH.sub.2CH.sub.2).sub.12OH), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) 3.45-3.85 (46H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.11OH, including those derived from compounds 3, 6 and 13), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) Yield: 1027 g
(Example 2-3, Step (B))
(85) Synthesis of Compound 14 in which a is 12, R.sup.1 is Trityl Group, and R.sup.2 is Tert-Butyl Group in Formula (6)
(86) ##STR00033##
(87) The reaction product containing the compound 12 (compound 12: 200 g, less than 253 mmol) and dichloromethane (1000 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the product was dissolved under a nitrogen atmosphere, followed by addition of powdery potassium hydroxide (1.4 g, 25 mmol). After cooling to 5? C., tert-butyl acrylate (a compound in which R.sup.2 in the formula (5) is a tert-butyl group, 55 mL, 380 mmol) was added dropwise, and the mixture was reacted at 5? C. for 2 hours. After the reaction, a saturated aqueous ammonium chloride solution (400 mL) was added and liquid separation was performed. The organic layer was washed once with a saturated aqueous sodium chloride solution (600 mL). The organic layer was concentrated under reduced pressure to obtain a reaction product containing the compound 14 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6 and 13.
(88) Compound 14
(89) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.49 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 3.45-3.85 (48H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.11OCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) Yield: 226 g (Yield: 97%)
(Example 2-4, Steps (C), (D))
(90) Synthesis of Compound 15 in which a is 12 and R.sup.2 is Tert-Butyl Group in Formula (7)
(91) ##STR00034##
(92) A reaction product containing the compound 14 (compound 14: 226 g, less than 246 mmol) and methanol (1.1 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube, and a stirrer, and the product was dissolved under a nitrogen atmosphere. Thereafter, p-toluenesulfonic acid monohydrate (23 g, 123 mmol) and hexane (900 mL) were added. After stirring at room temperature for 30 minutes, the hexane layer was removed, hexane (565 mL) was added again, and the mixture was stirred for 30 minutes. After performing the same operation four times, the disappearance of the compounds 14, 3, 6 and 13 was confirmed as a result of .sup.1H-NMR measurement, and a saturated aqueous sodium hydrogen carbonate solution (452 mL) was added. The mixed solution was washed once with hexane (565 mL) to remove trityl methyl ether. The methanol solution was concentrated under reduced pressure to obtain a crude product containing the compound 15. It was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained crude product contained the above compounds 1, 9 and 16.
(93) Next, dichloromethane (1.1 L) was added to the crude product, and the mixture was washed three times with ion-exchanged water (1.1 L) and once with a saturated aqueous sodium chloride solution (1.1 L) under the condition of 10? C. From TLC analysis, the disappearance of the compounds 1, 9 and 13 was confirmed. Magnesium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a purified product containing the compound 15 as a pale yellow transparent liquid.
(94) Compound 15
(95) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.6 (1H, t, H(OCH.sub.2CH.sub.2).sub.12OCH.sub.2CH.sub.2), 3.45-3.85 (50H, m, H(OCH.sub.2CH.sub.2).sub.12OCH.sub.2CH.sub.2) Yield: 118 g (yield: 71%)
(Example 2-5, Step (E))
(96) Synthesis of Compound 17 in which a is 12 and R.sup.2 is Tert-Butyl Group in Formula (8)
(97) ##STR00035##
(98) The compound 15 (117 g, 173 mmol) and dichloromethane (469 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the compound was dissolved under a nitrogen atmosphere. Then, phthalimide (36 g, 245 mmol) and triphenylphosphine (64 g, 244 mmol) were added. After stirring at room temperature for 30 minutes, diisopropyl azodicarboxylate (42 g, 208 mmol) diluted with dichloromethane (118 mL) was added dropwise, and the mixture was stirred for 1 hour. After 1 hour, the disappearance of the compound 15 was confirmed by TLC, the solvent was distilled off under reduced pressure, methanol (518 mL) and ethylenediamine monohydrate (141 mL, 1.73 mol) were added, and a reaction was carried out at 40? C. for 3 hours. A 6N aqueous hydrochloric acid solution was added thereto for neutralization, the solvent was distilled off under reduced pressure, sodium chloride and chloroform (585 mL) were added for extraction. A saturated aqueous sodium chloride solution (585 mL) was added to the resultant extract solution, and washing was performed twice. A 1% aqueous sodium dihydrogen phosphate solution (585 mL) was added to the organic layer, and the mixture was extracted four times. While a mixed solvent of chloroform (293 mL) and toluene (293 mL) was added to the extracted aqueous solution, washing was performed seven times. After a 5N aqueous sodium hydroxide solution was added thereto for neutralization, sodium chloride was added, and extraction was performed with dichloromethane (590 mL) four times. Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 17 as a pale yellow transparent liquid.
(99) Compound 17
(100) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.1 (2H, t, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.11), 3.45-3.85 (48H, m, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.11CH.sub.2CH.sub.2) Yield: 113 g (yield: 97%)
(Example 2-6, Step (F))
(101) Synthesis of Compound 18 in which a in Formula (1) is 12
(102) ##STR00036##
(103) The compound 17 (50 g, 74 mmol) and 1 M hydrochloric acid (250 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the compound was dissolved, followed by stirring at 50 to 55? C. for 1 hour. After cooling to 15? C., the mixture was neutralized with a 10M aqueous sodium hydroxide solution, and sodium chloride and dichloromethane (250 mL) were added for washing. The aqueous layer was again washed once with dichloromethane (250 mL), adjusted to pH 2 with 6M hydrochloric acid, and extracted with dichloromethane (250 mL) four times. The recovered organic layers were mixed and dehydrated over sodium sulfate, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 18 as a pale yellow solid.
(104) Compound 18
(105) .sup.1H-NMR (CD.sub.3OD internal standard TMS); ? (ppm): 2.56 (2H, t, CH.sub.2CH.sub.2COOH), 3.17 (2H, t, H.sub.2NCH.sub.2CH.sub.2O), 3.6-3.9 (48H, m, CH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.11CH.sub.2CH.sub.2) Yield: 44 g (Yield: 91%) Purity: 99.8% (HPLC-RI)
(106) The HPLC measurement conditions used for the purity measurement are shown below.
(107) Apparatus: alliance manufactured by Waters Corporation.
(108) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(109) Detector: RI
(110) Developing solvent: a solution of methanol/5 mM ammonium acetate solution=27.5/72.5
(111) Flow rate: 1.0 mL/min
(112) Column temperature: 40? C.
(113) Sample concentration: 2 mg/mL
(114) Injection volume: 50 ?L
Example 3
(115) Synthesis of Compound 40 in which a is 24 in Formula (1)
(116) ##STR00037##
Example 3-1
(117) Synthesis of Compound 19 in which a is 12, R.sup.1 is Trityl Group, and L is Mesyl Group in Formula (3)
(118) ##STR00038##
(119) The reaction product containing the compound 12 (compound 12: 90 g, less than 0.11 mol) and toluene (451 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (29 ml, 0.21 mol). Methanesulfonyl chloride (14 mL, 0.18 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 12 was confirmed by TLC analysis, a 5% aqueous sodium dihydrogen phosphate solution (450 mL) was added, and liquid separation was performed. The organic layer was washed once with a 5% aqueous sodium dihydrogen phosphate solution (450 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (450 mL), and once with a saturated aqueous sodium chloride solution (450 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 19 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6 and 13.
(120) Compound 19
(121) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 3.07 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) 3.45-3.85 (44H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.10OCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 4.37 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) Yield: 96 g
(122) From the .sup.1H-NMR measurement results of the compound 19 of Example 3-1, it was confirmed that the compounds 3, 6 and 13 were contained in an amount of about 15 mol % (compound 3: 6 mol %, compound 6: 5 mol %, compound 13: 4 mol %, rough estimation).
(123) A calculation expression of the content of the compounds 3, 6 and 13 on the basis of a ? 3.23 peak is expressed by the following expression.
(((2?[? 4.32])/4H)/([? 4.32]/2H))?100 (mol %)
(124) With regard to the content of the compounds 3 and 6, values calculated in Example 1-1 and 2-1 are applied.
(125) Further, the reaction product 12 used in Example 3-1 contains the compounds 3, 6 and 13 in an amount of about 15.9 wt %.
(Example 3-2, Step (A))
(126) Synthesis of Compound 20 in which a is 16 and R.sup.1 is Trityl Group in Formula (4)
(127) ##STR00039##
(128) Sodium hydride (6.3 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer. After nitrogen substitution, MeCN (192 mL) was added and the mixture was cooled to 0? C. MeCN (96 mL) was mixed with tetraethylene glycol 1 (108 g, 0.56 mol) azeotropically dehydrated with toluene (53 mL), and this mixed solution was added dropwise over 30 minutes. After completion of the dropwise addition, MeCN (96 mL) was mixed with the reaction product containing the compound 19 (compound 19: 96 g, less than 0.11 mol), and the mixed solution was added dropwise over 15 minutes. After completion of the dropwise addition, the reaction mixture was heated to 75? C. and stirred for 3 hours. After 3 hours, it was confirmed using .sup.1H-NMR that the compound 19 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. A saturated aqueous ammonium chloride solution (170 mL) and hexane (146 mL) were added to the reaction mixture solution and liquid separation was performed. The lower layer from which the hexane layer (upper layer) had been removed was concentrated under reduced pressure, and toluene (481 mL) was added to the residue. The toluene solution was washed once with a saturated aqueous ammonium chloride solution (260 mL) and three times with a saturated aqueous sodium chloride solution (480 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 20 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6, 13 and 21.
(129) Compound 20
(130) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.78 (1H, b, C(OCH.sub.2CH.sub.2).sub.16OH), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13 and 21) 3.45-3.85 (62H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.15OH, including those derived from compounds 3, 6, 13 and 21), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13 and 21) Yield: 103 g
Example 3-3
(131) Synthesis of Compound 22 in which a is 16, R.sup.1 is Trityl Group, and L is Mesyl Group in Formula (3)
(132) ##STR00040##
(133) The reaction product containing the compound 20 (compound 20: 100 g, less than 0.10 mol) and toluene (500 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (20 ml, 0.14 mol). Methanesulfonyl chloride (9.6 mL, 0.12 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 14 was confirmed by TLC analysis, a 5% aqueous sodium dihydrogen phosphate solution (500 mL) was added, and liquid separation was performed. The organic layer was washed once with a 5% aqueous sodium dihydrogen phosphate solution (500 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (500 mL), and once with a saturated aqueous sodium chloride solution (500 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 22 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6, 13 and 21.
(134) Compound 22
(135) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 3.07 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13 and 21) 3.45-3.85 (60H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.14OCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13 and 21), 4.37 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13 and 21) Yield: 106 g
(Example 3-4, Step (A))
(136) Synthesis of Compound 23 in which a is 20 and R.sup.1 is Trityl Group in Formula (4)
(137) ##STR00041##
(138) Sodium hydride (5.7 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and after nitrogen substitution, MeCN (208 mL) was added and the mixture was cooled to 0? C. MeCN (105 mL) was mixed with tetraethylene glycol 1 (97 g, 0.50 mol) azeotropically dehydrated with toluene (48 mL), and this mixed solution was added dropwise over 30 minutes. After completion of the dropwise addition, MeCN (105 mL) was mixed with the reaction product containing the compound 22 (compound 22: 109 g, less than 0.10 mol), and the mixed solution was added dropwise over 15 minutes. After completion of the dropwise addition, the reaction mixture was heated to 75? C. and stirred for 3 hours. After 3 hours, it was confirmed using .sup.1H-NMR that the compound 22 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. A saturated aqueous ammonium chloride solution (190 mL) and hexane (159 mL) were added to the reaction mixture solution and liquid separation was performed. The lower layer from which the hexane layer (upper layer) had been removed was concentrated under reduced pressure, and toluene (524 mL) was added to the residue. This toluene solution was washed once with a saturated aqueous ammonium chloride solution (285 mL) and three times with a saturated aqueous sodium chloride solution (520 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 23 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6, 13, 21 and 24.
(139) Compound 22
(140) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.64 (1H, b, C(OCH.sub.2CH.sub.2).sub.20OH), 3.23 (2H, t, (C-.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21 and 24) 3.45-3.85 (78H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.19OH, including those derived from compounds 3, 6, 13, 21 and 24), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21 and 24) Yield: 109 g
Example 3-5
(141) Synthesis of Compound 25 in which a is 20, R.sup.1 is Trityl Group, and L is Mesyl Group in Formula (3)
(142) ##STR00042##
(143) The reaction product containing the compound 23 (compound 23: 107 g, less than 0.094 mol) and toluene (585 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (18 ml, 0.13 mol). Methanesulfonyl chloride (8.7 mL, 0.11 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 14 was confirmed by TLC analysis, a 5% aqueous sodium dihydrogen phosphate solution (535 mL) was added, and liquid separation was performed. The organic layer was washed once with a 5% aqueous sodium dihydrogen phosphate solution (535 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (535 mL), and once with a saturated aqueous sodium chloride solution (535 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 25 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6, 13, 21 and 24.
(144) Compound 25
(145) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 3.07 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21 and 24) 3.45-3.85 (76H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.18OCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21 and 24), 4.37 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21 and 24) Yield: 113 g
(Example 3-6, Step (A))
(146) Synthesis of Compound 26 in which a is 24 and R.sup.1 is Trityl Group in Formula (4)
(147) ##STR00043##
(148) Sodium hydride (5.2 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and after nitrogen substitution, MeCN (221 mL) was added and the mixture was cooled to 0? C. MeCN (111 mL) was mixed with tetraethylene glycol 1 (88 g, 0.46 mol) azeotropically dehydrated with toluene (44 mL), and this mixed solution was added dropwise over 30 minutes. After completion of the dropwise addition, MeCN (111 mL) was mixed with the reaction product containing the compound 25 (compound 25: 113 g, less than 0.092 mol), and the mixed solution was added dropwise over 15 minutes. After completion of the dropwise addition, the reaction mixture was heated to 75? C. and stirred for 3 hours. After 3 hours, it was confirmed using .sup.1H-NMR that the compound 25 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. A saturated aqueous ammonium chloride solution (200 mL) and hexane (168 mL) were added to the reaction mixture solution and liquid separation was performed. The lower layer from which the hexane layer (upper layer) had been removed was concentrated under reduced pressure, and toluene (556 mL) was added to the residue. The toluene solution was washed once with a saturated aqueous ammonium chloride solution (300 mL) and three times with a saturated aqueous sodium chloride solution (555 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 26 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6, 13, 21, 24 and 27.
(149) Compound 26
(150) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.66 (1H, b, C(OCH.sub.2CH.sub.2).sub.24OH), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21, 24 and 27) 3.45-3.85 (94H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.23OH, including those derived from compounds 3, 6, 13, 21, 24 and 27), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21, 24 and 27) Yield: 115 g
(Example 3-7, Step (B))
(151) Synthesis of Compound 28 in which a is 24, R.sup.1 is Trityl Group, and R.sup.2 is Tert-Butyl Group in Formula (6)
(152) ##STR00044##
(153) The reaction product containing the compound 28 (compound 28: 5.02 g, less than 3.79 mmol) and dichloromethane (25 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the product was dissolved under a nitrogen atmosphere, followed by addition of powdery potassium hydroxide (128 mg, 2.28 mmol). After cooling to 5? C., tert-butyl acrylate (a compound in which R.sup.2 in the formula (5) is a tert-butyl group, 1.1 mL, 7.58 mmol) was added dropwise, and the mixture was reacted at 5? C. for 4 hours. After the reaction, a saturated aqueous ammonium chloride solution (10 mL) was added and liquid separation was performed. The organic layer was washed once with a saturated aqueous sodium chloride solution (15 mL). The organic layer was concentrated under reduced pressure to obtain a reaction product containing the compound 28 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6, 13, 21, 24 and 27.
(154) Compound 28
(155) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.49 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21, 24 and 27), 3.45-3.85 (96H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.23OCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21, 24 and 27), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6, 13, 21, 24 and 27) Yield: 5.31 g
(Example 3-8, Step (C))
(156) Synthesis of Compound 29 in which a is 24 and R.sup.2 is Tert-Butyl Group in Formula (7)
(157) ##STR00045##
(158) A reaction product containing the compound 28 (compound 28: 5.01 g, less than 3.46 mmol) and methanol (25 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube, and a stirrer, and the product was dissolved under a nitrogen atmosphere. Thereafter, p-toluenesulfonic acid monohydrate (329 mg, 1.72 mmol) and hexane (20 mL) were added. After stirring at room temperature for 30 minutes, the hexane layer was removed, hexane (13 mL) was added again, and the mixture was stirred for 30 minutes. After performing the same operation four times, the disappearance of the compounds 28, 3, 6, 13, 21, 24 and 27 was confirmed as a result of .sup.1H-NMR measurement, and a saturated aqueous sodium hydrogen carbonate solution (11 mL) was added. The mixed solution was washed once with hexane (13 mL) to remove trityl methyl ether. The methanol solution was concentrated under reduced pressure and a 20% aqueous sodium chloride solution (10 mL) and dichloromethane (10 mL) were added, followed by liquid separation. Magnesium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 29 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 1, 9, 16 30, 31 and 32.
(159) Compound 29
(160) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.7 (1H, b, H(OCH.sub.2CH.sub.2).sub.24OCH.sub.2CH.sub.2), 3.45-3.85 (98H, m, H(OCH.sub.2CH.sub.2).sub.24OCH.sub.2CH.sub.2, including those derived from compounds 1, 9, 16 30, 31 and 32) Yield: 3.13 g
(Example 3-9, Step (E))
(161) Synthesis of Compound 33 in which a is 24 and R.sup.2 is Tert-Butyl Group in Formula (8)
(162) ##STR00046##
(163) The reaction product containing the compound 29 (compound 29: 3.00 g, less than 2.49 mmol) and dichloromethane (12 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere. Then, phthalimide (641 mg, 4.36 mmol) and triphenylphosphine (1.14 g, 4.35 mmol) were added. After stirring at room temperature for 30 minutes, diisopropyl azodicarboxylate (756 mg, 3.74 mmol) diluted with dichloromethane (3 mL) was added dropwise, and the mixture was stirred for 3 hours. After 3 hours, the disappearance of the compound 29 was confirmed by TLC analysis, the solvent was distilled off under reduced pressure, methanol (13 mL) and ethylenediamine monohydrate (2 mL, 24.8 mol) were added, and a reaction was carried out at 40? C. for 4 hours. A 6N aqueous hydrochloric acid solution was added thereto for neutralization, the solvent was distilled off under reduced pressure, sodium chloride and chloroform (15 mL) were added for extraction, and while a saturated aqueous sodium chloride solution (15 mL) was added to the extract solution, washing was performed twice. A 1% aqueous sodium dihydrogen phosphate solution (15 mL) was added to this organic layer, and the mixture was extracted four times. While a mixed solvent of chloroform (8 mL) and toluene (8 mL) was added to the extracted aqueous solution, washing was performed seven times. After a 5N aqueous sodium hydroxide solution was added thereto for neutralization, sodium chloride was added, and extraction was performed four times with dichloromethane (15 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 33 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and.sup.1H-NMR measurement results that the obtained a reaction product contained the above compounds 34, 35, 36, 37, 38 and 39.
(164) Compound 33
(165) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.1 (2H, t, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.23), 3.45-3.85 (96H, m, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.23CH.sub.2CH.sub.2, including those derived from compounds 34, 35, 36, 37, 38 and 39) Yield: 2.45 g
(Example 3-10, Steps (F), (G))
(166) Synthesis of Compound 40 in which a in Formula (1) is 24
(167) ##STR00047##
(168) The reaction product containing the compound 33 (compound 33: 1.22 g, less than 1.00 mmol) and 1 M hydrochloric acid (6 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved, followed by stirring at 50 to 55? C. for 2 hours. After 2 hours, the disappearance of the compound 33 was confirmed by TLC. The mixture was cooled to room temperature.
(169) Then, a 10M aqueous sodium hydroxide solution was added to the reaction solution to adjust the pH to 10, and while dichloromethane (6 mL) were added, washing was performed twice. The solution was adjusted to pH 2 with 6M hydrochloric acid, and after addition of sodium chloride, extraction was performed with dichloromethane (6 mL) three times. The recovered organic layers were mixed and dehydrated over sodium sulfate, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 40 (1.16 g) as a pale yellow oily solid. After tetrahydrofuran (5 mL) was added to the obtained compound 40 for dissolution, the solution was cooled to 10? C. and hexane (24 mL) cooled to 10? C. was added to effect crystallization. The crystals were collected by filtration and washed the crystals with hexane (10 mL) cooled to 10? C. The crystals were collected by filtration and dried under vacuum to obtain the compound 40 as white powdery crystals.
(170) Compound 40
(171) .sup.1H-NMR (CD.sub.3OD internal standard TMS); ? (ppm): 2.56 (2H, t, CH.sub.2CH.sub.2COOH), 3.17 (2H, t, H.sub.2NCH.sub.2CH.sub.2O), 3.6-3.9 (96H, m, CH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.23CH.sub.2CH.sub.2) Yield: 893 mg
Example 4
(172) In order to calculate the content of the compound 5 in which a is 8 in the compound where R.sup.1 in the formula (4) is a trityl group when the content of the compound having trityl groups at both terminals is removed from the reaction product containing the compound 5 obtained in Example 1-2, derivatization of the compound 42 was performed.
(173) Synthesis of Compound 41 in which a is 8, R.sup.1 is Trityl Group, and L is Tosyl Group in Formula (3)
(174) ##STR00048##
(175) After the reaction product containing the compound 5 (compound 5: 5 g, less than 8.2 mmol) and dichloromethane (25 mL) was added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the product was dissolved under a nitrogen atmosphere, triethylamine (1.2 mL, 8.6 mmol), 4-dimethylaminopyridine (100 mg, 0.82 mmol) and TsCl (1.4 g, 7.3 mmol) were added, and the mixture was stirred at room temperature for 4 hours. After 4 hours, the disappearance of TsCl was confirmed by .sup.1H-NMR analysis, a 1M aqueous hydrochloric acid solution (25 mL) was added, and liquid separation was performed. The organic layer was washed once with a 1M aqueous hydrochloric acid solution (25 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (25 mL), and once with a saturated aqueous sodium chloride solution (25 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing compound 41 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and the .sup.1H-NMR measurement results that the obtained reaction products contained the above compounds 3 and 6.
(176) Compound 41
(177) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 3.45-3.85 (28H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.6OCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 4.16 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 7.35 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) Yield: 6.1 g
(178) Synthesis of Compound 42 Having Ethylene Glycol Chain Length of 8 Units Having Hydroxyl Group at One Terminal and Tosyl Group at One Terminal
(179) ##STR00049##
(180) Next, the reaction product containing compound 42 (compound 42: 6.1 g, less than 8.0 mmol) and methanol (25 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer. After the compound was dissolved under a nitrogen atmosphere, p-toluenesulfonic acid monohydrate (0.78 g, 4.1 mmol) and hexane (20 mL) were added. After stirring at room temperature for 30 minutes, the hexane layer was removed, hexane (13 mL) was added again, and the mixture was stirred for 30 minutes. After performing the same operation six times, the disappearance of the compounds 42, 3 and 6 was confirmed as a result of .sup.1H-NMR measurement, and a saturated aqueous sodium hydrogen carbonate solution (10 mL) was added. The mixed solution was washed twice with hexane (13 mL) to remove trityl methyl ether. The methanol solution was concentrated under reduced pressure, and dichloromethane (25 mL) was added for extraction. Magnesium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 42 as a pale yellow transparent liquid.
(181) Compound 42
(182) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 2.73 (1H, t, H(OCH.sub.2CH.sub.2).sub.8), 3.45-3.85 (30H, m, (OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2), 4.16 (2H, t, (OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.35 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) Yield: 3.0 g Purity: 99.7% (HPLC-RI)
(183) The HPLC measurement conditions used for the purity measurement are shown below.
(184) Apparatus: alliance manufactured by Waters Corporation.
(185) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(186) Detector: RI
(187) Developing solvent: a solution of methanol/5 mM ammonium acetate=50/50
(188) Flow rate: 0.6 mL/min
(189) Column temperature: 40? C.
(190) Sample concentration: 0.2 mg/mL
(191) Injection volume: 40 ?L
(192) The purity value is the ratio of the peak area of the compound 43 to the total peak area detected over a retention time of 10 to 40 min.
Example 5
(193) In order to calculate the content of compound 12 in which a is 12 in the compound where R.sup.1 in the formula (4) is a trityl group when the content of the compound having trityl groups at both terminals is removed from the reaction product containing the compound 12 obtained in Example 2-2, derivatization of the compound 44 was performed.
(194) Synthesis of Compound 43 in which a is 12, R.sup.1 is Trityl Group, and L is Tosyl Group in Formula (3)
(195) ##STR00050##
(196) After the reaction product containing compound 12 (compound 12: 5 g, less than 6.3 mmol) and dichloromethane (25 mL) was added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the product was dissolved under a nitrogen atmosphere, triethylamine (0.93 mL, 6.6 mmol), 4-dimethylaminopyridine (77 mg, 0.63 mmol) and TsCl (0.97 g, 5.4 mmol) were added, and the mixture was stirred at room temperature for 4 hours. After 4 hours, the disappearance of TsCl was confirmed by .sup.1H-NMR analysis, a 1M aqueous hydrochloric acid solution (25 mL) was added, and liquid separation was performed. The organic layer was washed once with a 1M aqueous hydrochloric acid solution (25 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (25 mL), and once with a saturated aqueous sodium chloride solution (25 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing compound 43 as a pale yellow transparent liquid. Moreover, it was confirmed from TLC analysis and the .sup.1H-NMR measurement results that the obtained reaction product contained the above compounds 3, 6 and 13.
(197) Compound 43
(198) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 3.45-3.85 (44H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.10OCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 4.16 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 7.35 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) Yield: 6.1 g
(199) ##STR00051##
(200) Next, the reaction product containing the compound 43 (compound 43: 6.1 g, less than 6.5 mmol) and methanol (25 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer. After the compound was dissolved under a nitrogen atmosphere, p-toluenesulfonic acid monohydrate (0.60 g, 3.2 mmol) and hexane (20 mL) were added. After stirring at room temperature for 30 minutes, the hexane layer was removed, hexane (13 mL) was added again, and the mixture was stirred for 30 minutes. After performing the same operation six times, the disappearance of the compounds 43, 3, 6 and 13 was confirmed as a result of .sup.1H-NMR measurement, and a saturated aqueous sodium hydrogen carbonate solution (10 mL) was added. The mixed solution was washed twice with hexane (13 mL) to remove trityl methyl ether. The methanol solution was concentrated under reduced pressure and dichloromethane (25 mL) was added, followed by extraction. Magnesium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 44 as a pale yellow transparent liquid.
(201) Compound 44
(202) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 2.73 (1H, t, H(OCH.sub.2CH.sub.2).sub.8), 3.45-3.85 (46H, m, (OCH.sub.2CH.sub.2).sub.11OCH.sub.2CH.sub.2), 4.16 (2H, t, (OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.35 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) Yield: 3.3 g Purity: 99.7% (HPLC-RI)
(203) The HPLC measurement conditions used for the purity measurement are shown below.
(204) Apparatus: alliance manufactured by Waters Corporation.
(205) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(206) Detector: RI
(207) Developing solvent: a solution of methanol/5 mM ammonium acetate=55/45
(208) Flow rate: 0.6 mL/min
(209) Column temperature: 40? C.
(210) Sample concentration: 0.2 mg/mL
(211) Injection volume: 40 ?L
(212) The purity value is the ratio of the peak area of the compound 44 to the total peak area detected over a retention time of 11 to 40 min.
Comparative Example 1, Production Method for Obtaining Compound 10 Via Tosylation Step
(213) A compound 10 in which a is 8 in the formula (1) was synthesized by the production method described in Patent Literature 4.
Comparative Example 1-1
(214) Synthesis of Compound 41 in which a is 4, R.sup.1 is Trityl Group, and L is Tosyl Group in Formula (3)
(215) ##STR00052##
(216) Tetraethylene glycol 1 (200 mL, 1.15 mol) and toluene (50 mL) were added to a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the compound was dissolved under a nitrogen atmosphere and then azeotropically dehydrated at 110 to 120? C. After the azeotropic dehydration, the mixture was cooled, pyridine (18 ml, 0.22 mol) and trityl chloride (TrtCl, 40 g, 0.14 mol) were added, and the mixture was stirred at room temperature for 3 hours. After 3 hours, the disappearance of TrtCl was confirmed using TLC, and ion-exchanged water (200 mL) was added. Toluene (100 mL) was added to the obtained mixed solution, liquid separation was performed, and the organic layer was washed once with a mixed solution of ion-exchanged water (80 mL) and a saturated aqueous sodium chloride solution (20 mL), once with a 1M aqueous hydrochloric acid solution (50 mL), and four times with a saturated aqueous sodium chloride solution (50 mL). Sodium sulfate was added to the obtained organic layer, which was dried and filtered. Toluene (50 mL) was added to the filtrate, and azeotropic dehydration was performed three times to obtain a reaction product containing the compound 2 as a pale yellow transparent liquid. Moreover, it was confirmed by ESI-MS measurement that the obtained reaction product contained the above compound 3.
(217) Compound 2
(218) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.4 (1H, t, C(OCH.sub.2CH.sub.2).sub.4OH), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3), 3.45-3.85 (14H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.3OH, including one derived from compound 3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3) MS (ESI.sup.+): Compound 2 454.5 [M+NH.sub.4].sup.+, Compound 3 696.9 [M+NH.sub.4].sup.+ Yield: 63.8 g
(219) ##STR00053##
(220) Next, the reaction product containing compound 2 (compound 2: 62.8 g, less than 0.14 mol) and toluene (314 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer. After dissolution under a nitrogen atmosphere, triethylamine (24 mL, 0.17 mol) was added. Methanesulfonyl chloride (12.2 mL, 0.16 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 2 was confirmed by TLC analysis, and a 1M aqueous hydrochloric acid solution (314 mL) was added, followed by liquid separation. The organic layer was washed once with a 1M aqueous hydrochloric acid solution (314 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (314 mL), and once with a saturated aqueous sodium chloride solution (314 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 4 as a pale yellow transparent liquid. Moreover, it was confirmed by ESI-MS measurement that the obtained reaction product contained the above compound 3.
(221) Compound 4
(222) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.98 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3), 3.45-3.85 (12H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.2OCH.sub.2CH.sub.2, including one derived from compound 3) 4.33 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including one derived from compound 3) MS (ESI.sup.+): Compound 4 532.4 [M+NH.sub.4].sup.+, Compound 3 696.8 [M+NH.sub.4].sup.+ Yield: 70.4 g
(223) From .sup.1H-NMR measurement results of the compound 4 of Comparative Example 1-1, it was confirmed that the compound 3 was contained in an amount of about 4.8 mol %.
(224) A calculation expression of the compound 3 content on the basis of a ? 3.23 peak is expressed by the following expression.
(((2?[? 4.32])/4H)/([? 4.32]/2H))?100 (mol %)
(225) Further, the reaction product 2 obtained in Comparative Example 1-1 contains the compound 3 in an amount of about 6.9 wt %.
Comparative Example 1-2
(226) Synthesis of compound 5 in which a is 8 and R.sup.1 is trityl group in formula (4)
(227) ##STR00054##
(228) Sodium hydride (7.8 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and after nitrogen substitution, MeCN (141 mL) was added and the mixture was cooled to 0? C. MeCN (70 mL) was mixed with tetraethylene glycol 1 (213 g, 1.10 mol) azeotropically dehydrated with toluene (107 mL), and this mixed solution was added dropwise over 30 minutes. After completion of the dropwise addition, MeCN (70 mL) was mixed with the reaction product containing the compound 4 (compound 4: 70.4 g, less than 0.14 mol), and the mixed solution was added dropwise over 15 minutes. After completion of the dropwise addition, the reaction mixture was heated to 80? C. and stirred for 3 hours. After 3 hours, it was confirmed using .sup.1H-NMR that the compound 4 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. The reaction mixture solution was concentrated under reduced pressure, and toluene (352 mL) was added to the residue. The toluene solution was washed twice with a saturated aqueous ammonium chloride solution (352 mL) and three times with a saturated aqueous sodium chloride solution (352 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 5 as a pale yellow transparent liquid. Moreover, it was confirmed from ESI-MS measurement that the obtained reaction products also contained the above compounds 3 and 6.
(229) Compound 5
(230) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.52 (1H, t, C(OCH.sub.2CH.sub.2).sub.8OH), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) 3.45-3.85 (30H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.7OH, including those derived from compounds 3 and 6), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) MS (ESI.sup.+): Compound 5 630.8 [M+NH.sub.4].sup.+, Compound 3 696.8 [M+NH.sub.4].sup.+, Compound 6 1048.4 [M+NH.sub.4].sup.+ Yield: 82.4 g
Comparative Example 1-3
(231) Synthesis of Compound 41 in which a is 8, R.sup.1 is Trityl Group, and L is Tosyl Group in Formula (3)
(232) ##STR00055##
(233) Tetrahydrofuran (200 mL) was added to the reaction product containing the compound 5 (Compound 5: 77.8 g, less than 0.14 mol) in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and the product was dissolved under a nitrogen atmosphere. Then, the solution was cooled to 0? C. An aqueous sodium hydroxide solution (20 g, 0.5 mol/60 mL) was added, and the mixture was stirred at 0? C. for 20 minutes. A tosyl chloride/tetrahydrofuran solution (30 g, 0.16 mnol/60 mL) was added dropwise to the reaction mixture over 30 minutes, and the mixture was stirred at 0? C. for 1.5 hours. After 1.5 hours, after confirming the disappearance of the compound 5 by TLC, the mixture was stirred at room temperature for 12.5 hours in order to make excess tosyl chloride disappear. After 12.5 hours, the disappearance of tosyl chloride was confirmed by TLC and ion-exchanged water (30 mL) and diethyl ether (50 mL) were added. The mixture was washed once with a saturated aqueous sodium hydrogen carbonate solution (50 mL) and three times with a saturated aqueous sodium chloride solution (50 mL). Sodium sulfate was added to the organic layer for dehydration, followed by dehydration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing compound 41 as a pale yellow transparent liquid. Moreover, it was confirmed by ESI-MS measurement that the obtained reaction product contained the above compounds 3 and 6.
(234) Compound 41
(235) MS (ESI.sup.+): Compound 41 785.2 [M+NH.sub.4].sup.+, Compound 3 697.0 [M+NH.sub.4].sup.+, Compound 6 1048.7 [M+NH.sub.4].sup.+ Yield: 86.1 g (Yield: 88%)
Comparative Example 1-4
(236) Synthesis of Compound 42 Having an Ethylene Glycol Chain Length of 8 Units and Having a Hydroxyl Group at One Terminal and a Tosyl Group at One Terminal
(237) ##STR00056##
(238) The reaction product containing the compound 41 (compound 41: 85.6 g, less than 0.144 mol), methanol (20 mL), palladium carbon (Pd/C, 2 g) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, hydrogen substitution was performed, and the mixture was stirred at room temperature for 18 hours. After 18 hours, the disappearance of the compound 42 was confirmed by TLC, and Pd/C was removed by celite filtration. Ion-exchanged water (130 mL) was added to the filtrate, and formed triphenylmethane was filtered. Since triphenylmethane remained in the filtrate, it was washed with hexane (100 mL) five times to remove triphenylmethane. The methanol/ion-exchanged water layer was concentrated under reduced pressure to obtain a crude product containing the compound 41. Then, dichloromethane (120 mL) was added to the crude product and the mixture was washed three times with ion-exchanged water (100 mL) and twice with a saturated aqueous sodium chloride solution (100 mL) under the condition of 20? C. Sodium sulfate was added to the organic layer, which was dried and filtered. The filtrate was concentrated under reduced pressure to obtain a purified product of the compound 42 as a pale yellow transparent liquid.
(239) Compound 42
(240) Purified Product
(241) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 2.73 (1H, t, H(OCH.sub.2CH.sub.2).sub.8), 3.45-3.85 (30H, m, (OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2), 4.16 (2H, t, (OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.35 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) MS (ESI.sup.+): Compound 42 542.4 [M+NH.sub.4].sup.+ Crude product MS (ESI.sup.+): Compound 42 542.4 [M+NH.sub.4].sup.+, Compound 1 212.7 [M+NH.sub.4].sup.+, Compound 9 564.5 [M+NH.sub.4].sup.+ Yield: 52.0 g (yield: 89%) Purity: 96.7% (HPLC-RI)
(242) The HPLC measurement conditions used for the purity measurement are shown below.
(243) Apparatus: alliance manufactured by Waters Corporation.
(244) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(245) Detector: RI
(246) Developing solvent: a solution of methanol/5 mM ammonium acetate=50/50, Flow rate: 0.6 mL/min
(247) Column temperature: 40? C.
(248) Sample concentration: 0.2 mg/mL
(249) Injection volume: 40 ?L
(250) The purity value is the ratio of the peak area of the compound 42 to the total peak area detected over the retention time of 10 to 40 min.
Comparative Example 1-5
(251) Synthesis of Compound 45 Having an Ethylene Glycol Chain Length of 8 Units and Having a Tosyl Group at One Terminal and a Tert-Butyl Ester Group at One Terminal
(252) ##STR00057##
(253) The compound 45 (1.0 g, 1.91 mmol), tert-butyl acrylate (1.8 mL, 19 mmol), and toluene (25 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and they were dissolved under a nitrogen atmosphere. Thereafter, the mixture was cooled to 0? C., powdery potassium hydroxide (53 mg, 0.9 mmol) was added, and the mixture was reacted at 0? C. for 1 hour. After the reaction, ion-exchanged water (20 mL) was added and liquid separation was performed. The organic layer was washed once with a saturated aqueous sodium chloride solution (20 mL). The organic layer was concentrated under reduced pressure to obtain the compound 45 as a pale yellow transparent liquid.
(254) Compound 45
(255) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.45 (9H, s, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 2.50 (2H, t, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 3.45-3.85 (32H, m, CH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2), 4.16 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.34 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) MS (ESI.sup.+): Compound 45 670.6 [M+NH.sub.4].sup.+ Yield: 1.06 g (yield: 85%)
Comparative Example 1-6
(256) Synthesis of Compound 46 Having an Ethylene Glycol Chain Length of 8 Units and Having a Phthalimide Group at One Terminal and a Tert-Butyl Ester Group at One Terminal
(257) ##STR00058##
(258) After the compound 45 (1.06 g, 1.60 mmol) and acetonitrile (25 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the compound was dissolved under a nitrogen atmosphere, phthalimide potassium salt (520 mg, 2.80 mmol) was added, and the mixture was stirred at 80? C. for 8 hours. The disappearance of the compound 45 was confirmed by NMR, and the reaction solution was concentrated. After adding dichloromethane (7 mL) and filtrating the solid content, the filtrate was washed once with a 0.1M aqueous sodium hydroxide solution (7 mL) and once with a saturated aqueous sodium chloride solution (10 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 46 as a pale yellow transparent liquid.
(259) Compound 46
(260) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.45 (9H, s, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 2.50 (2H, t, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 3.45-3.85 (32H, m, CH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2), 3.90 (2H, t, OCH.sub.2CH.sub.2-phthalimide), 7.71 (2H, dd, -phthalimide), 7.80 (2H, dd, -phthalimide) MS (ESI.sup.+): Compound 46 945.7 [M+NH.sub.4].sup.+ Yield: 965 mg (yield: 95%)
Comparative Example 1-7
(261) Synthesis of compound 9 in which a is 8 and R.sup.2 is tert-butyl group in formula (8)
(262) ##STR00059##
(263) The compound 46 (510 mg, 0.80 mmol), ethanol (10 mL) and hydrazine monohydrate (334 mg, 6.70 mmol) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and a reaction was performed at 85? C. for 45 minutes. After cooling to room temperature, in order to dissolve precipitated white solid, a 12% aqueous potassium carbonate solution (5 mL) was added and then the mixed solution was concentrated under reduced pressure. Next, ion-exchanged water (3 mL) was added, concentrated hydrochloric acid (0.6 mL) was added to adjust the pH to 3, and the solid mass was filtered. The filtrate was washed three times with dichloromethane and the aqueous layer was saturated by adding sodium chloride. This aqueous solution was extracted 5 times with dichloromethane, and the organic layer was dehydrated over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the compound 9 as a pale yellow transparent liquid.
(264) Compound 9
(265) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.1 (2H, t, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.7), 3.45-3.85 (32H, m, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.7CH.sub.2CH.sub.2) Yield: 400 mg (yield: 99%)
Comparative Example 1-8
(266) Synthesis of Compound 10 in which a in Formula (1) is 8
(267) ##STR00060##
(268) After the compound 9 (400 mg, 0.80 mmol) and 1M hydrochloric acid (0.5 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, the compound was dissolved and then the mixture was stirred at 55? C. for 5 hours. After cooling to 15? C., a 10M aqueous sodium hydroxide solution was added to adjust the pH to 5. Dichloromethane (10 mL) was added to the solid mass obtained by performing azeotropic dehydration of water with toluene (5 mL) twice, and the mixture was filtered. The filtrate was concentrated under reduced pressure to obtain the compound 10 as a pale yellow transparent liquid.
(269) Compound 10
(270) .sup.1H-NMR (CD.sub.3OD, internal standard TMS); ? (ppm): 2.56 (2H, t, CH.sub.2CH.sub.2COOH), 3.17 (2H, t, H.sub.2NCH.sub.2CH.sub.2O), 3.6-3.9 (32H, m, CH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.7CH.sub.2CH.sub.2) Yield: 320 mg (yield: 83%) Purity: 95.6% (HPLC-RI)
(271) The HPLC measurement conditions used for the purity measurement are shown below.
(272) Apparatus: alliance manufactured by Waters Corporation.
(273) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(274) Detector: RI
(275) Developing solvent: a solution of methanol/5 mM ammonium acetate=15/85,
(276) Flow rate: 1.0 mL/min
(277) Column temperature: 40? C.
(278) Sample concentration: 1 mg/mL
(279) Injection volume: 50 ?L
Comparative Example 2, Production Method for Obtaining Compound 18 Via Tosylation Step
(280) Compound 18 in which a in formula (1) is 12 was synthesized by the production method described in Patent Literature 4.
Comparative Example 2-1
(281) Synthesis of Compound 11 in which a is 8, R.sup.1 is Trityl Group, and L is Mesyl Group in Formula (3)
(282) ##STR00061##
(283) The reaction product containing the compound 5 (compound 5: 72.7 g, less than 0.12 mol) and toluene (350 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (20 ml, 0.14 mol). Methanesulfonyl chloride (10 mL, 0.13 mol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 5 was confirmed by TLC analysis, a 1M aqueous hydrochloric acid solution (100 mL) was added, and liquid separation was performed. The organic layer was washed once with a 1M aqueous hydrochloric acid solution (100 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (100 mL), and once with a saturated aqueous sodium chloride solution (100 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 11 as a pale yellow transparent liquid. Moreover, it was confirmed from ESI-MS measurement that the obtained reaction product contained the above compounds 3 and 6.
(284) Compound 11
(285) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 3.07 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) 3.45-3.85 (28H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.6OCH.sub.2CH.sub.2, including those derived from compounds 3 and 6), 4.37 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3 and 6) MS (ESI.sup.+): Compound 11 708.3 [M+NH.sub.4].sup.+, Compound 3 696.4 [M+NH.sub.4].sup.+, Compound 6 1048.5 [M+NH.sub.4].sup.+ Yield: 80.1 g
(286) From the .sup.1H-NMR measurement results of the compound 11 of Comparative Example 2-1, it was confirmed that the compounds 3 and 6 were contained in an amount of about 9.5 mol % (compound 3: 4.8 mol %, compound 6: 4.7 mol %, rough estimation).
(287) A calculation expression of the contents of the compounds 3 and 6 on the basis of a ? 3.23 peak is expressed by the following expression.
(((2?[? 4.32])/4H)/([? 4.32]/2H))?100 (mol %)
(288) As the content of the compound 3, the value calculated in Comparative Example 1-1 is applied.
(289) Further, the reaction product 5 used in Comparative Example 2-1 contains the compounds 3 and 6 in an amount of about 11.7 wt %.
Comparative Example 2-2
(290) Synthesis of Compound 12 in which a is 12 and R.sup.1 is Trityl Group in Formula (4)
(291) ##STR00062##
(292) Sodium hydride (6.6 g) was placed in a reactor fitted with a thermometer, a nitrogen inlet tube and a stirrer, and after nitrogen substitution, MeCN (200 mL) was added and the mixture was cooled to 0? C. MeCN (50 mL) was mixed with tetraethylene glycol 1 (180 g, 0.93 mol) azeotropically dehydrated with toluene (50 mL), and this mixed solution was added dropwise over 30 minutes. After completion of the dropwise addition, MeCN (50 mL) was mixed with the reaction product containing the compound 11 (compound 11: 80.1 g, less than 0.12 mol), and the mixed solution was added dropwise over 15 minutes. After completion of the dropwise addition, the reaction mixture was heated to 80? C. and stirred for 3 hours. After 3 hours, it was confirmed using .sup.1H-NMR that the compound 11 had disappeared, and the mixture was allowed to cool until the temperature became 40? C. or lower. The reaction mixture solution was concentrated under reduced pressure, and toluene (200 mL) was added to the residue. This toluene solution was washed twice with a saturated aqueous ammonium chloride solution (100 mL) and three times with a saturated aqueous sodium chloride solution (100 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 12 as a pale yellow transparent liquid. Moreover, it was confirmed from ESI-MS and .sup.1H-NMR measurement that the obtained reaction product contained the above compounds 3, 6 and 13.
(293) Compound 12
(294) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.56 (1H, t, C(OCH.sub.2CH.sub.2).sub.12OH), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) 3.45-3.85 (46H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.11OH, including those derived from compounds 3, 6 and 13), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) MS (ESI.sup.+): Compound 12 806.4 [M+NH.sub.4].sup.+, Compound 3 696.8 [M+NH.sub.4].sup.+, Compound 6 1048.1 [M+NH.sub.4].sup.+, Compound 13 1400.9 [M+NH.sub.4].sup.+ Yield: 85.4 g
Comparative Example 2-3
(295) Synthesis of Compound 43 in which a is 12, R.sup.1 is Trityl Group, and L is Tosyl Group in Formula (3)
(296) ##STR00063##
(297) After the reaction product containing the compound 12 (compound 12: 57.3 g, less than 72.7 mmol) and dichloromethane (280 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, the product was dissolved under a nitrogen atmosphere, triethylamine (10 mL, 73 mol), 4-dimethylaminopyridine (888 mg, 7.27 mmol) and TsCl (12.5 g, 65.5 mmol) were added, and the mixture was stirred at room temperature for 4.5 hours. After 4.5 hours, the disappearance of TsCl was confirmed by .sup.1H-NMR analysis, a 1M aqueous hydrochloric acid solution (150 mL) was added, and liquid separation was performed. The organic layer was washed once with a 1M aqueous hydrochloric acid solution (150 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (150 mL), and once with a saturated aqueous sodium chloride solution (150 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 43 as a pale yellow transparent liquid. Moreover, it was confirmed from ESI-MS measurement that the obtained reaction product contained the above compounds 3, 6 and 13.
(298) Compound 43
(299) MS (ESI.sup.+): Compound 43 960.3 [M+NH.sub.4].sup.+, Compound 3 696.3 [M+NH.sub.4].sup.+, Compound 6 1048.2 [M+NH.sub.4].sup.+, Compound 13 1400.8 [M+NH.sub.4].sup.+ Yield: 69.1 g (yield: 101%)
Comparative Example 2-4
(300) Synthesis of Compound 44 Having Ethylene Glycol Chain Length of 12 Units and Having Hydroxyl Group at One Terminal and Tosyl Group at One Terminal
(301) ##STR00064##
(302) The reaction product containing compound 43 (compound 43: 69.1 g, less than 73.3 mmol) and methanol (550 L) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer. After the compound was dissolved under a nitrogen atmosphere, p-toluenesulfonic acid monohydrate (6.97 g, 36.7 mmol) and hexane (200 mL) were added. After stirring at room temperature for 30 minutes, the hexane layer was removed, hexane (200 mL) was added again, and the mixture was stirred for 30 minutes. After performing the same operation six times, the disappearance of the compounds 43, 3, 6 and 13 was confirmed as a result of .sup.1H-NMR measurement, and a saturated aqueous sodium hydrogen carbonate solution (200 mL) was added. The mixed solution was washed twice with hexane (200 mL) to remove trityl methyl ether. The methanol solution was concentrated under reduced pressure, and dichloromethane (200 mL) was added and the mixture was washed three times with ion-exchanged water (200 mL) and once with a saturated aqueous sodium chloride solution (200 mL) under the condition of 20? C. or lower. Magnesium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 44 as a pale yellow transparent liquid.
(303) Compound 44
(304) Purified Product
(305) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 2.73 (1H, t, H(OCH.sub.2CH.sub.2).sub.8), 3.45-3.85 (46H, m, (OCH.sub.2CH.sub.2).sub.11OCH.sub.2CH.sub.2), 4.16 (2H, t, (OCH.sub.2CH.sub.2).sub.7OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.35 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) MS (ESI.sup.+): Compound 44 718.3 [M+NH.sub.4].sup.+ Crude product MS(ESI.sup.+): Compound 44 718.3 [M+NH.sub.4].sup.+, Compound 1 212.3 [M+NH.sub.4].sup.+, Compound 9 564.5 [M+NH.sub.4].sup.+, Compound 16 916.4 [M+NH.sub.4].sup.+ Yield: 30.3 g (yield: 59%) Purity: 94.9% (HPLC-RI)
(306) The HPLC measurement conditions used for the purity measurement are shown below.
(307) Apparatus: alliance manufactured by Waters Corporation.
(308) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc.
(309) Detector: RI
(310) Developing solvent: a solution of methanol/5 mM ammonium acetate=55/45
(311) Flow rate: 0.6 mL/min
(312) Column temperature: 40? C.
(313) Sample concentration: 0.2 mg/mL
(314) Injection volume: 40 ?L
(315) The purity value is the ratio of the peak area of the compound 44 to the total peak area detected over the retention time of 11 to 40 min.
Comparative Example 2-5
(316) Synthesis of Compound 47 Having Ethylene Glycol Chain Length of 12 Units and Having Tosyl Group at One Terminal and Tert-Butyl Ester Group at One Terminal
(317) ##STR00065##
(318) The compound 44 (4.96 g, 7.08 mmol), tert-butyl acrylate (3.09 mL, 21.2 mmol), and toluene (100 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and they were dissolved under a nitrogen atmosphere. Thereafter, the mixture was cooled to 0? C., powdery potassium hydroxide (199 mg, 3.54 mmol) was then added, and the mixture was reacted at 0? C. for 1 hour. After the reaction, a saturated aqueous ammonium chloride solution (50 mL) was added and liquid separation was performed. The organic layer was washed once with a saturated aqueous sodium chloride solution (50 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 47 as a pale yellow transparent liquid.
(319) Compound 47
(320) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.45 (9H, s, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 2.45 (3H, s, OSO.sub.2-Ph-CH.sub.3), 2.50 (2H, t, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 3.45-3.85 (48H, m, CH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.11OCH.sub.2CH.sub.2), 4.16 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2-Ph-CH.sub.3), 7.34 (2H, d, OSO.sub.2-Ph-CH.sub.3), 7.80 (2H, d, OSO.sub.2-Ph-CH.sub.3) MS (ESI.sup.+): Compound 47 847.0 [M+NH.sub.4].sup.+ Yield: 5.43 g (Yield: 93%)
Comparative Example 2-6
(321) Synthesis of Compound 48 Having Ethylene Glycol Chain Length of 12 Units and Having Phthalimide Group at One Terminal and Tert-Butyl Ester Group at One Terminal
(322) ##STR00066##
(323) After the compound 47 (5.43 g, 6.55 mmol) and acetonitrile (45 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the compound was dissolved under a nitrogen atmosphere, phthalimide potassium salt (1.58 g, 8.52 mmol) was added, and the mixture was stirred at 80? C. for 18 hours. The disappearance of the compound 48 was confirmed by NMR, and the reaction solution was concentrated. After adding dichloromethane (50 mL) to the residue and filtering the solid mass, the filtrate was washed once with a 0.1M aqueous sodium hydroxide solution (50 mL) and once with a saturated aqueous sodium chloride solution (50 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain the compound 48 as a pale yellow transparent liquid.
(324) Compound 48
(325) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.45 (9H, s, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 2.50 (2H, t, (CH.sub.3).sub.3COCOCH.sub.2CH.sub.2), 3.45-3.85 (48H, m, CH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.11OCH.sub.2CH.sub.2), 3.90 (2H, t, OCH.sub.2CH.sub.2-phthalimide), 7.71 (2H, dd, -phthalimide), 7.80 (2H, dd, -phthalimide) MS (ESI.sup.+): Compound 48 828.1 [M+NH.sub.4].sup.+ Yield: 4.26 g (yield: 81%)
Comparative Example 2-7
(326) Synthesis of Compound 17 in which a is 12 and R.sup.2 is Tert-Butyl Group in Formula (8)
(327) ##STR00067##
(328) The compound 48 (4.26 g, 5.30 mmol), ethanol (60 mL) and hydrazine monohydrate (3.86 mL, 79.5 mmol) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, and a reaction was performed at 85? C. for 1 hour. After cooling to room temperature, in order to dissolve precipitated white solid, a 12% aqueous potassium carbonate solution (5 mL) was added and then the mixed solution was concentrated under reduced pressure. Ion-exchanged water (20 mL) was added to the residue, concentrated hydrochloric acid was added to adjust the pH to 3, and the solid mass was filtered. Sodium chloride was added to the filtrate, which was extracted twice with dichloromethane (20 mL). The organic layer was dehydrated over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the compound 17 as a pale yellow transparent liquid.
(329) Compound 17
(330) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 1.44 (9H, s, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 2.50 (2H, t, CH.sub.2CH.sub.2COOC(CH.sub.3).sub.3), 3.1 (2H, t, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.11), 3.45-3.85 (48H, m, H.sub.2NCH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.11CH.sub.2CH.sub.2) Yield: 3.13 g (yield: 88%)
Comparative Example 2-8
(331) Synthesis of Compound 18 in which a in Formula (1) is 12
(332) ##STR00068##
(333) After the compound 17 (3.13 g, 4.65 mmol) and 1M hydrochloric acid (3 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer, the compound was dissolved and then the mixture was stirred at 55? C. for 2 hours. After cooling to 15? C., the mixture was diluted with ion-exchanged water (5 mL) and was washed with dichloromethane (10 mL) three times. A 2M aqueous sodium hydroxide solution was added to the aqueous layer to adjust the pH to 9. The aqueous solution was washed three times with dichloromethane (10 mL) and the aqueous layer was saturated by adding sodium chloride. This aqueous solution was extracted three times with chloroform (10 mL), and the organic layer was dehydrated over sodium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain the compound 18 as a pale yellow transparent liquid.
(334) Compound 18
(335) .sup.1H-NMR (CD.sub.3OD, internal standard TMS); ? (ppm): 2.56 (2H, t, CH.sub.2CH.sub.2COOH), 3.17 (2H, t, H.sub.2NCH.sub.2CH.sub.2O), 3.6-3.9 (48H, m, CH.sub.2CH.sub.2O(CH.sub.2CH.sub.2O).sub.11CH.sub.2CH.sub.2) Yield: 2.32 g (yield: 76%) Purity: 98.0% (HPLC-RI)
(336) The HPLC measurement conditions used for the purity measurement are shown below.
(337) Apparatus: alliance manufactured by Waters Corporation,
(338) Column: Inertsil ODS-3 (column size: 4.6 mm?25 cm, particle size 5 ?m) manufactured by GL Science Inc,
(339) Detector: RI,
(340) Developing solvent: a solution of methanol/5 mM ammonium acetate=27.5/72.5,
(341) Flow rate: 1.0 mL/min,
(342) Column temperature: 40? C.,
(343) Sample concentration: 2 mg/mL, Injection volume: 50 ?L
Comparative Example 3
(344) Calculation Expression of Contents of Compounds 3, 6 and 13 Having Trityl Groups at Both Terminals in Reaction Product Containing Compound 12 Obtained in Comparative Example 2-2
(345) Synthesis of Compound 19 in which a is 12, R.sup.1 is Trityl Group, and L is Mesyl Group in Formula (3)
(346) ##STR00069##
(347) The reaction product containing the compound 12 (compound 12: 25 g, less than 317 mmol) and toluene (125 mL) were added to a reaction vessel fitted with a thermometer, a nitrogen inlet tube and a stirrer and the product was dissolved under a nitrogen atmosphere, followed by addition of triethylamine (5.3 ml, 38 mmol). Methanesulfonyl chloride (2.7 mL, 35 mmol) was added dropwise at 10? C., and the mixture was stirred at room temperature for 2 hours. After 2 hours, the disappearance of the compound 12 was confirmed by TLC analysis, a 1M aqueous hydrochloric acid solution (50 mL) was added, and liquid separation was performed. The organic layer was washed once with a 1M aqueous hydrochloric acid solution (50 mL), twice with a saturated aqueous sodium hydrogen carbonate solution (50 mL), and once with a saturated aqueous sodium chloride solution (50 mL). Sodium sulfate was added to the organic layer for dehydration, followed by filtration. The filtrate was concentrated under reduced pressure to obtain a reaction product containing the compound 19 as a pale yellow transparent liquid. Moreover, it was confirmed from ESI-MS measurement that the obtained reaction product contained the above compounds 3, 6 and 13.
(348) Compound 19
(349) .sup.1H-NMR (CDCl.sub.3, internal standard TMS); ? (ppm): 3.07 (3H, s, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 3.23 (2H, t, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) 3.45-3.85 (44H, m, OCH.sub.2CH.sub.2(OCH.sub.2CH.sub.2).sub.10OCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13), 4.37 (2H, t, OCH.sub.2CH.sub.2OSO.sub.2CH.sub.3), 7.21-7.47 (15H, m, (C.sub.6H.sub.5).sub.3COCH.sub.2CH.sub.2, including those derived from compounds 3, 6 and 13) MS (ESI.sup.+): Compound 19 884.9 [M+NH.sub.4].sup.+, Compound 3 696.8 [M+NH.sub.4].sup.+, Compound 6 1049.4 [M+NH.sub.4].sup.+, Compound 13 1401.3 [M+NH.sub.4].sup.+ Yield: 26 g
(350) From the .sup.1H-NMR measurement results of the compound 19 of Comparative Example 3, it was confirmed that the compounds 3, 6 and 13 were contained in an amount of about 9.8 mol % (compound 3: 4.8 mol %, compound 6: 4.7 mol %, compound 13: 0.3 mol %, rough estimation).
(351) A calculation expression of the contents of the compounds 3, 6 and 13 content on the basis of a ? 3.23 peak is expressed by the following expression.
(((2?[? 4.32])/4H)/([? 4.32]/2H))?100 (mol %)
(352) As the compounds 3 and 6, the values calculated in Comparative Examples 1-1 and 2-1 are applied.
(353) Further, the reaction product 12 used in Comparative Example 3 contains the compounds 3, 6 and 13 in an amount of about 9.7 wt %.
(354) Comparison of total yields until obtaining compound 10 of hetero monodisperse polyethylene glycol having amino group and carboxyl group at respective both terminals obtained using compound 5 of hetero monodisperse polyethylene glycol having hydroxyl group and trityl group at respective both terminals as raw material
(355) Table 1 shows total yields when the formula (1) was produced from the formula (4) in the case where a was 8 or 12.
(356) TABLE-US-00001 TABLE 1 Value of a Example Comparative Example 8 64% 58% 12 72% 34%
(357) The total yields were calculated following the following expression.
Case of a=8: (Value obtained by multiplying yields in respective steps until obtaining compound 10 from compound 5)?(Pure content of compound 10 in which a is 8)/(Pure content of compound 5 in which a is 8)
Case of a=12: (Value obtained by multiplying yields in respective steps until obtaining compound 18 from compound 12)?(Pure content of compound 18 in which a is 12)/(Pure content of compound 12 in which a is 12)
(358) In the expression for determining the total yield, the pure content of the compound 5 in which a is 8 is a value obtained by multiplying a value obtained by subtracting the contents of the compounds 3 and 6 having trityl groups at both terminals from the amount of the reaction product containing the compound 5 obtained in Example 1-2 by the purity of the compound 42 obtained in Example 4. Further, it is a value obtained by multiplying a value obtained by subtracting the contents of the compounds 3 and 6 having trityl groups at both terminals from the amount of the reaction product containing the compound 5 obtained in Comparative Example 1-2 by the purity of the compound 42 obtained in Comparative Example 1-4.
(359) In the expression for determining the total yield, the pure content of the compound 12 in which a is 12 is a value obtained by multiplying a value obtained by subtracting the contents of the compounds 3, 6 and 13 having trityl groups at both terminals from the amount of the reaction product containing the compound 12 obtained in Example 2-2 by the purity of the compound 44 obtained in Example 5. Further, it is a value obtained by multiplying a value obtained by subtracting the contents of the compounds 3, 6 and 13 having trityl groups at both terminals from the amount of the reaction product containing the compound 5 obtained in Comparative Example 2-2 by the purity of the compound 44 obtained in Comparative Example 2-4.
(360) In the expressions for determining the total yields, the pure contents of the compound 10 in which a is 8 and the compound 18 in which a is 12 correspond to the purities determined by HPLC, respectively.
(361) In the conventional production method, the hydroxyl group is once converted into a tosyl group in order to purify impurities containing trityl groups at both terminals, which are different in chain length and have specific molecular weights, formed as by-products in the chain length extension step. On the other hand, by removing the impurities without this step, the number of steps is smaller than before, and the presence of the residual unreacted raw material and the formation of reaction by-products which may cause a yield decrease can be suppressed, so that the yield could be improved.
INDUSTRIAL APPLICABILITY
(362) According to the present invention, it is possible to provide a novel method for producing a highly pure hetero-type monodisperse polyethylene glycol having an amino group and a carboxyl group at respective both terminals.
(363) Although the present invention has been described in detail and with reference to specific embodiments, it will be apparent to those skilled in the art that various changes and modifications can be made without departing from the spirit and scope of the present invention.
(364) This application is based on a Japanese patent application filed on Mar. 29, 2019 (Japanese Patent Application No. 2019-065528), the contents of which are incorporated herein by reference.