Synthesis and Biological Evaluation of Symmetrical 2,4,6-Trisubstituted 1,3,5-Triazine Derivatives

Abstract

We describe the synthesis and biological evaluation of newly designed 2,4,6-trisubstituted symmetrical 1,3,5-triazine (TAZ) derivatives. Among the tested trisubstituted symmetrical TAZ derivatives, various C₃- or C_S-symmetrical alkoxy-amino-substituted TAZ derivatives showed significant antiviral activity against herpes simplex virus type 1 (HSV-1) and/or cytotoxic activity against Vero cells. The structure–activity relationships for anti-HSV-1 activity of these symmetrical 2,4,6-trisubstituted TAZ derivatives are also described. Experimental results indicated that a C_S-symmetrical TAZ structure with introduction of two alkoxy groups and one amine moiety seems to be the minimally required structure for anti-HSV-1 activity.

Molecular recognition of two-fold (C₂) or three-fold (C₃) symmetrical geometry macromolecules is one of the common features in many important biological responses,^1,2) and we have therefore already designed a few symmetrical target molecules for the purpose of finding biologically active new leads or candidates.^3–5) With reference to the molecular symmetry, small molecules having C₃-, C_S-, or C₂-symmetrical geometry frequently appear in various biologically active compounds.^6–8) Such small symmetrical molecules are usually constructed on a corresponding symmetrical template.

From this point of view, we have recently reported some molecular modifications of tris(2-aminoethyl)amine (TAEA) derivatives to C₃- or C_S-symmetrical tripodal receptor type molecules and the results of biological evaluation of these symmetrical compounds.³⁾ We have also reported an interesting lectin-like property for sugar recognition of some of these tripodal receptor type TAEA molecules.⁴⁾

In order to achieve a suprafacial three-dimensional interaction of a bioactive symmetrical molecule for its binding site, the nature of substituents in the molecule is thought to be very important for preferential interactions. Such interactions are dictated largely by van der Waals interactions or formation of hydrogen bonds. The introduction of amine (basic nitrogen atom) and/or a bivalent oxygen group such as a hydroxy or alkoxy group into a C₃-symmetrical 1,3,5-triazine (TAZ) template seems to be interesting, because it is well known that amine functionalities behave as both hydrogen acceptors and hydrogen donors in appropriate circumstances. These facts may indicate that molecules with some amine and/or bivalent oxygen functionalities on a heterocyclic C₃-symmetrical TAZ template can produce a property for suprafacial hydrogen-bonding interaction.

For an extension of our studies, we planned to investigate the synthesis of 2,4,6-trisubstituted symmetrical TAZ derivatives^9–16) as well as to evaluate their biological properties. In this paper, we report the results of new synthetic routes for C₃- or C_S-symmetrical 2,4,6-trisubstituted TAZ derivatives in the target molecules together with the results of anti-herpes simplex virus type 1 (HSV-1) activity and cytotoxic activity against Vero cell of the obtained symmetrical 2,4,6-trisubstituted TAZ derivatives.

Results and Discussion

Synthesis

To begin with, we carried out a few synthetic trials for the preparation of target symmetrical triacylamino-substituted TAZ (2) derivatives by using 2,4,6-triamino-1,3,5-triazine (TATAZ) as a starting material. However, our attempts to separate the target products (2x or 2y) after acylation reactions of TATAZ with acylating agents failed¹⁷⁾ (Fig. 1).

Fig. 1. Target Triacylamino-Substituted TAZ

We therefore attempted using 2,4,6-trichloro-1,3,5-triazine (TCTAZ; 1) as a starting material to prepare target molecules, and we examined substitution reactions by amine or alcohol nucleophiles under various reaction conditions shown in a synthetic review by Blotny.¹⁸⁾ As a result, we obtained many types of target trisubstituted symmetrical TAZ derivatives. The obtained results are summarized in Table 1.

Table 1. TAZ Derivatives from Reactions of TCTAZ (1) and Amines (XH)

Entry	Amine XH	Ratio of 1 : amine : (additive)	Conditionsa)	Solvent	Products (yield %)
1		1 : 3.3	Reflux 15 min	AcOH	2a (83)
2		1 : 3.3	Reflux 1.5 h	AcOH	2b·HCl (82)
3		1 : 6	MW 50 W, 160°C, 5 min	Dioxane	2d (70)
4		1 : 6	MW 50 W, 100°C, 5 min	EtOH	2d (24), 3dq (36), 4dqq (4)
5		1 : 2	MW 50 W, 160°C, 5 min	i-PrOH	3dr (2), 5d (12), 6d (11)
6		1 : 4 : (10 DIPEAb))	MW 50 W, 160°C, 10 min	i-PrOH	2d (8), 3dr (4), 5d (27)
7		1 : 2.5 : (5 DIPEAb))	MW 50 W, 160°C, 10 min	i-PrOH	2d (9), 3dr (1), 5d (88)
8		1 : 2.5 : (5 DIPEAb))	MW 50 W, 160°C, 10 min	n-HepOH	3ds (3), 5d (26)
9		1 : 2 : (2 TEAc))	0°C, 1 h to rt 17 h	CH2Cl2	5d (57)
10		1 : 6	180°C, 1 h		2e (29)
11		1 : 2 : (2 TEAc))	0°C, 1 h to rt 18 h	dry CH2Cl2	5f (33)
12		1 : 3	rt 1 h	dry THF	5g·3HCl (20)
13		1 : 3.3d)	Reflux 1 d	AcOH	Ag (46)

a) MW means microwave irradiation. b) N,N-Diisopropylethylamine. c) Triethylamine. d) The target compound could not be isolated.

As shown in Table 1, triamino-substituted C₃-symmetrical derivatives (2a,¹⁹⁾ 2b, and 2d) were obtained predominantly in good yields from reactions of TCTAZ (1) with the corresponding amine nucleophile (Entries 1–3).

Trials for the preparation of alkoxy-diamino-trisubstituted TAZ (3) by nucleophilic substitutions of TCTAZ with an aliphatic amine (dH) and an alcohol (qH, rH, or sH) as nucleophiles under various reaction conditions (Entries 4–9) resulted in the formation of a mixture of a few types of target trisubstituted TAZ derivatives. Among these entries, the highest yield (88%) of diamino-chloro-trisubstituted TAZ derivative (5d) was obtained from microwave-assisted reaction with i-PrOH as a solvent in the presence of N,N-diisopropylethylamine (DIPEA) (Entry 7). Reactions of TCTAZ with an amine nucleophile (dH or fH) under mild conditions (room temperature) or reactions using an alcohol as a solvent acting as a nucleophile gave diamino-monochloro-substituted TAZ derivatives (5d or 5f²⁰⁾) and monoamino-dichloro-substituted TAZ product (6d) (Entries 5–9, and 11).

Using the method reported by Azarifar et al.,²¹⁾ reactions of TCTAZ (1) with a racemic 3-hydroxypiperidine (eH) as an amine nucleophile also resulted in the formation of triamino-substituted TAZ (2e) in 29% yield (Entry 10).

The reaction of TCTAZ with N-methylpiperazine (gH) proceeded smoothly even at room temperature to afford the corresponding diamino-monochloro-TAZ derivative (5g·HCl) in 20% yield (Entry 12). Dimerization between two intermediates TAZs (5g and 2g) occurred in the reaction of TCTAZ with a nucleophile (gH) at a high reaction temperature, resulting in the formation of dimeric triazine (Ag), which had a bridged structure with two TAZ rings (Entry 13, see Experimental for details). The reaction mechanism for formation of the dimerized compound Ag via triamino-TAZ (2g)²²⁾ is shown in Chart 1.

Chart 1. Formation Mechanism of Dimeric TAZ Derivative (Ag)

In terms of selective formation of targeted symmetrical alkoxy-amino-TAZ derivatives, we found that the above-described synthetic strategy for the preparation of alkoxy-amino-substituted (trisubstituted) TAZ from TCTAZ (1) has disadvantages as a method for preparation of targeted symmetrical alkoxy-amino-TAZ derivatives (3 or 4).

Mono- or diamino-substituted triazines generally showed poor reactivity against weak nucleophiles such as alcohols. On the other hand, alkoxy-substituted chloro-TAZs had higher reactivities as substrates in further nucleophic substitutions with amine nucleophiles than those of amine-substituted chloro-TAZ derivatives. Therefore, for the purpose of preparation of designed alkoxy-amino-TAZ derivatives as our target molecules, the synthetic procedure via alkoxy-substituted chloro-TAZ is expected to be superior to the procedure via amine-substituted chloro-TAZs.

Chart 2 shows the strategic point of synthetic pathways via alkoxy-chloro-TAZ derivatives (7, 8) to alkoxy-amino-TAZ derivatives (3, 4). Our method via alkoxy-substituted chloro-TAZ intermediates for target alkoxy-amino-TAZs consisting of stepwise nucleophilic substitutions gave moderate to excellent results as shown in Table 2.

Chart 2

Table 2. Synthesis of Alkoxy-amino-TAZ Derivatives (3 and 4) from Reactions of TCTAZ (1) with Various Alcohols (ROH) and Amines (XH)

Entry	Reagents	Ratio of reagents and/or (additive)a)	Conditionsb)	Solvent	Products (yield %)c)
14	i) 1, sH	1 : sH : dH : (collidine) =	i) Collidine, 0°C, 1 h	Dry acetone	3ds (73), 4dss (4), 9sss (1)
	ii) dH	1 : 1.3 : 4 : (1)	ii) MW 50 W, 160°C, 10 min	Dry dioxane
15	i) 1, sH	1 : sH : dH : (collidine : DIPEA)=	i) Collidine, 0°C, 1 h	Dry acetone	3ds (1), 4dss (47)
	ii) dH	1 : 2.6 : 2 : (2 : 2)	ii) DIPEA, rt 6 h	Dry acetonitrile
16	1, sH	1: sH : (collidine)=1 : 1.3 : (1)	0°C, 1 h	Dry acetone	7s (70), 9sss (1)
17	7s, dH	7s : dH=1 : 6.3	MW 50 W, 160°C, 10 min	Dry dioxane	3ds (85)
18	7s, eH	7s : eH : (DIPEA)=1 : 3 : (3)	rt 2 h	Dry acetonitrile	3es (92)
19	7s, hH	7s : hH : (DIPEA)=1 : 2 : (3)	rt 20 h	Dry acetonitrile	3hs (69)
20	7s, iH	7s : iH : (DIPEA)=1 : 3 : (3)	rt 21 h	Dry acetonitrile	3is (37)
21	i) 1, tH	1 : tH : dH : (collidine)=	i) Collidine, rt 3 h	Dry acetone	3dt (74)
	ii) dH	1 : 1.3 : 4 : (1)	ii) MW 300 W, 160°C, 10 min	Dry dioxane
22	i) 1, tH	1 : tH : dH : (NaOH)=	i) Aq. NaOH, rt 2 h	MeOH	4dpp (20), 4dpt (8),
	ii) dH	1.1 : 1 : 5 : (4)	ii) MW 50 W, 150°C, 15 min	Dry dioxane	9 ppt (21),d) 9ptt (22)d)
23	i) 1, uH	1 : uH : dH : (collidine)=	i) Collidine, rt 3 h	Dry acetone	3du (29), Bdu (9), Cdu (3)
	ii) dH	1 : 1.3 : 4 : (1)	ii) MW 300 W, 160°C, 10 min	Dry dioxane

a) DIPEA stands for N,N-diisopropylethylamine. b) MW stands for microwave irradiation. c) Yield obtained from TCTAZ (1). d) Yields of 9ppt and 9ptt based on tH were 23% and 49%, respectively.

Among the results, it is noteworthy that when using an isolated monoalkoxy-dichloro-TAZ derivative (7s),²³⁾ reactions with amine nucleophiles (Entries 17–20) gave the corresponding target alkoxy-diamino-trisubstituted TAZ derivatives (3ds, 3es, 3hs, and 3is) in moderate to excellent yields. These facts apparently indicate that the strategy is advantageous for the purpose of preparation of designed trisubstituted symmetrical alkoxy-amino-TAZ derivatives as our target molecules (3 or 4). The results of stepwise one-pot reactions of TCTAZ with other alcohols and then amine nucleophiles are also shown in Table 2 (Entries 21–23).

From these reactions with TCTAZ as a starting material, we could prepare many target symmetrical TAZ derivatives (3 and 4) including 3dq, 3ds, 3dt, 3du, 3es, 3hs, 3is, 4dpp, 4dpt, 4dqq, and 4dss. In some runs, we also obtained trialkoxy-substituted TAZ derivatives (9ppt, 9ptt, and 9sss²⁴⁾).

All structures of the synthesized compounds were easily confirmed by spectroscopic and analytical data. The geometries of C₃- or C_S-symmetrical structures of target TAZ derivatives in this article were also confirmed by ¹³C-NMR spectroscopic data.

Antiviral Activity and Discussion

Synthesized TAZ derivatives were evaluated for anti-HSV-1 activities by plaque reduction assays²⁵⁾ and were also evaluated for their cytotoxicities against Vero cells. The results are summarized in Table 3, and aciclovir²⁶⁾ is also shown as a positive control. Calculated log P values for the compounds are also shown in Table 3. There were few significant correlations between log P values and EC₅₀ values (anti-HSV-1 activity) or between log P values and IC₅₀ values (cytotoxicity against Vero cells) among the compounds listed in Table 3. However, regarding structural features of the evaluated 2,4,6-trisubstituted TAZ derivatives, we obtained interesting results for bioactivities of symmetrical derivatives using a TAZ template.

Table 3. Antiviral Activity (EC₅₀), Cytotoxicity (IC₅₀) against HSV-1, and Calculated Log P^a)

Compound		EC50 (µM)	IC50 (µM)	Log P	Compound		EC50 (µM)	IC50 (µM)	Log P
	2a	>100	>200	6.64		3hs	>50	55.3	8.43
	2b·HCl	>100c)	>100d)	5.51e)		3is	>3.1	21.4	2.88
	2d	>100	>100	2.43		4dpp	92.1	>200	1.74
	2e	>100	>200	1.96		4dss	38.0	178.7	6.72
	3dq	395.6	>200	2.42		5d	247.5	266.0	2.39
	3ds	25–30	25–30	4.57		5f	>200	>200	2.67
	3dt	>6.3	42.2	3.52		9ptt	>200	>200	4.28
	3du	123.5	>200	3.36		Ag	112.1	>200	3.79
	3es	>200	>200	4.27		Bdu	>200	>200	5.03f)
					Aciclovirb)		1.1	>444	−0.76

a) Log P was calculated by using ChemDraw Ultra 10.0. b) Data were taken from ref. 26. c) Compound (2b·HCl) at a concentration of 100 µM reduced plaque formation by ca. 10%. d) Compound (2b·HCl) showed cytotoxicity at a concentration of 200 µM. e) A value of the free amine (2b). f) Log P was calculated by using CAChe 6.1.12.

Among the prepared trisubstituted TAZ derivatives, C₃-symmetrical TAZ derivatives (2a, 2b·HCl, 2d, 2e²⁷⁾) showed no significant anti-HSV-1 activity or cytotoxic activity at a concentration of less than 100 µM.

However, two C_S-symmetrical derivatives (3ds and 3es²⁷⁾), in which a hydroxylated cyclic amine moiety in the C₃-symmetrical TAZ molecules (2d and 2e) had been substituted with an alkoxy group (HepO), showed different properties of those bioactivities. In compound 3ds, both the antiviral activity and cytotoxicity were increased (EC₅₀ and IC₅₀=25–50 µM), but compound 3es did not show any significant change in either of the biological activities at a concentration of less than 200 µM. In the C_S-symmetrical derivatives (3dt) substituted with a different alkoxy group, similar effects regarding anti-HSV-1 activity and cytotoxicity are also observed (EC₅₀=>6.3 µM and IC₅₀=42.2 µM). In the CS-symmetrical compounds (3dq and 3du) having a different alkoxy group, weak anti-HSV-1 activity also appeared in both compounds (see Table 3). In the other C_S-symmetrical derivative (3is) in which a hydroxylated amine moiety (d) in the C_S-symmetrical molecule (3ds) had been replaced with a different amine functionality (i) bearing an ether-linked alcohol, both anti-HSV-1 activity and increased cytotoxicity were observed (EC₅₀=>3.1 µM and IC₅₀=21.4 µM). C_S-Symmetrical compound 3hs also showed both anti-HSV-1 activity and cytotoxicity (EC₅₀=>50 µM and IC₅₀=55.3 µM) comparable to those of compound 3ds. These results suggest the importance of the presence of hydroxy groups and also the importance of the introduction of an alkoxy group for each biological activity.

Regarding antiviral activities of these symmetrical derivatives, comparison of the activities of two C_S-symmetrical compounds (3ds and 4dss) is particularly interesting. C_S-Symmetrical compound 4dss, modified by additional introduction of the same alkoxy group (s: HepO) into the TAZ ring of 3ds instead of an amino moiety (d), showed almost the same level of antiviral activity (EC₅₀=38.0 µM) and decreased cytotoxicity (IC₅₀=178.7 µM). The results indicate that this modification of 3ds to 4dss gave a better selectivity index (SI=IC₅₀/EC₅₀) for the antiviral compound, though obtained SI value was still small, providing an easy way for generating an antiviral lead of this 2,4,6-trisubstituted TAZ series. With respect to functional groups included in the molecule, the observed anti-HSV-1 activity of this molecule 4dss indicates that the distance of one set of two functional groups [an amine moiety containing a hydroxy group and an alkoxy group] seems to be important for a preferential required structure for anti-HSV-1 activity. In fact, compound 4dpp in which two long-chain alkoxy groups of 4dss were replaced by two methoxy groups still had considerable activity (EC₅₀=92.1 µM) with low cytotoxicity (IC₅₀=>200 µM). In addition, both the observed anti-HSV-1 and cytotoxic activities of compound 5d were weak (EC₅₀=247.5 µM and IC₅₀=266.0 µM) and neither of the TAZ derivatives (5f and 9ptt) showed any significant anti-HSV-1 or cytotoxic activity at a concentration of less than 200 µM. These results suggest the importance of the presence of hydroxy groups and an alkoxy group in these C_S-symmetrical TAZ molecular structures for the development of anti-HSV-1 agents.

Concerning the biological activities of dimeric TAZ derivatives, compound Ag showed only weak anti-HSV-1 activity (EC₅₀=112.1 µM) and no cytotoxic activity at a concentration of less than 200 µM, but dimeric TAZ derivative Bdu did not show any significant biological activities at a concentration of less than 200 µM. Although there is still some uncertainty, the results seem to indicate that the tested dimeric symmetrical TAZ derivatives have a different mode of antiviral or cytotoxic activity.

Regarding biological activities, the obtained results in this TAZ series provide interesting information for further molecular modifications targeting antiviral compounds. Thus, the TAZ template seems to be a preferable template for antiviral compounds rather than the TAEA template,³⁾ because antiviral activities of most of the C_S-symmetrical compounds in this study were observed at lower concentrations than those of the corresponding cytotoxicities. In addition, many trisubstituted target C_S-symmetrical TAZ derivatives described in this paper showed wide ranges of antiviral (anti-HSV-1) activities.

In the search for new antiviral active candidates or new leads, the results indicate that this heterocyclic TAZ template is a potential new scaffold for designing antiviral active molecules. Considering the nature of an introduced substituent characterizing the lipophilicity of a molecule, further modifications for finding more promising antiviral leads with better selectivity indexes are now under investigation. Based on the results of our previous studies on a few types of symmetrical compounds,^3–5) we speculate that a symmetrical molecule constructed on a symmetric template or with a linker may be an efficient structural feature. For the elucidation of sugar recognition properties of some of the highly bioactive symmetrical compounds such as 3ds, 3dt, 3is, and 4dss described in this article, we are also planning to carry out calorimetric experiments.

Experimental

Melting points were determined using a micro melting point apparatus (Yanaco MP-S3) without correction. IR spectra were measured by a Shimadzu FTIR-8100 IR spectrophotometer. Low- and high-resolution mass spectra (LR-MS and HR-MS) were obtained by a JEOL JMS HX-110 double-focusing model equipped with an FAB ion source interfaced with a JEOL JMA-DA 7000 data system. ^lH- and ¹³C-NMR spectra were obtained by JEOL JNM A-500. Chemical shifts were expressed in δ ppm downfield from an internal tetramethylsilane (TMS) signal for ^lH-NMR and the carbon signal of the corresponding solvent [CDCl₃ (77.00 ppm), CD₃OD (49.00 ppm), DMSO-d₆ (39.50 ppm), and pyridine-d₅ (149.80 ppm)] for ¹³C-NMR. The signal assignments were confirmed by two dimensional (2D)-NMR analyses; ¹H–¹H 2D correlation spectroscopy (COSY), ¹H–¹³C heteronuclear multiple-quantum coherence (HMQC), and ¹H–¹³C heteronuclear multiple-bond connectivity (HMBC). Microanalyses were performed with a Yanaco MT-6 CHN corder. Routine monitoring of reactions was carried out using precoated Kieselgel 60F₂₅₄ plates (E. Merck). Microwave irradiation experiments were carried out in a CEM Discover Focused Microwave System. Centrifugal or flash column chromatography was performed on silica gel (Able-Biott, Fuji Silysia FL-60D, or Kanto 60N) with a UV detector. Commercially available starting materials were used without further purification.

General Procedure for Substitution Reactions of TCTAZ with Amines

TCTAZ (1, 1–10 mmol) was added to a stirred solution of an amine in an appropriate solvent (6–75 mL) with or without an additive, and the resulting mixture was refluxed immediately or stirred or was subjected to microwave irradiation (MW) under the conditions shown in Table 1. The obtained products were purified by chromatography or recrystallization from appropriate solvents, and the yields are also summarized in Table 1. Typical protocols for the preparation of triamino-TAZ derivatives 2a (Entry 1) and 2d (Entry 3) are shown below in detail.

4,4′,4″-(1,3,5-Triazine-2,4,6-triyltriimino)trisbenzoic Acid 1,1′,1″-Triethylester (2a) [96474-94-1]¹⁹⁾

Entry 1: TCTAZ (1; 1.84 g, 10 mmol) was added to a stirred solution of ethyl 4-aminobenzoate (aH, 5.45 g, 33 mmol) in glacial acetic acid (AcOH, 75 mL) and the resulting solution was refluxed immediately for 15 min. The precipitated white solids were collected by filtration, washed with boiling water (ca. 20 mL×3), and dried at 90°C in vacuo to afford product 2a (4.74 g, 83% yield) with high purity (>98% confirmed by NMR). Purification by recrystallization from i-PrOH gave an analytically pure product 2a (3.06 g, 54% yield) as colorless crystals: mp 222–225°C (from i-PrOH) [lit.¹⁹⁾ mp 158°C (EtOH)]. IR (KBr) cm⁻¹: 3450 (NH of amine), 1720 (C=O), 1280, 1105 (C–O). ¹H-NMR (DMSO-d₆) δ: 1.34 (9H, t, J=7.0 Hz, CH₃), 4.31 (6H, q, J=7.0 Hz, CH₂), 7.92 (6H, dm, J=8.7 Hz, H3, 5), 8.00 (6H, t, J=8.7 Hz, H2, 6), 9.90 (3H, s, NH). ¹³C-NMR (DMSO-d₆) δ: 14.15 (CH₃), 60.23 (CH₂), 119.44 (C2, 6), 123.14 (C4), 129.75 (C3, 5), 144.13 (C1), 163.66 (C=N), 165.40 (C=O). Positive-ion FAB-MS m/z: 571(M+H⁺). HR-FAB-MS m/z: 571.2303 (Calcd for C₃₀H₃₁N₆O₆: 571.2305). Anal. Calcd for C₃₀H₃₀N₆O₆·2.5H₂O: C, 58.53; H, 5.73; N, 13.65. Found: C, 58.51; H, 5.44; N, 13.67.

N²,N⁴,N⁶-Tris(1,3-benzodioxol-5-yl)-1,3,5-triazine Hydrochloride (2b·HCl)

Entry 2: In the same manner as that for the preparation of 2a (Entry 1), by the reaction of 1 and 3,4-methylenedioxyaniline (bH), product 2b·HCl (82%) with high purity (>98% confirmed by NMR) was obtained. Recrystallization from MeOH gave an analytically pure product 2b·HCl (42%) as a white powder: mp 272–274°C (from MeOH). IR (KBr) cm⁻¹: 3410 (NH of amine), 1240 (C–N), 1040 (C–O). ¹H-NMR (DMSO-d₆) δ: 6.01 (6H, s, H2), 6.87 (3H, d, J=8.2 Hz, H7), 6.99 (3H, d, J=8.2 Hz, H6), 7.35 (3H, br s, H4), 10.14 (3H, br s, NH). ¹³C-NMR (DMSO-d₆) δ: 101.13 (C2), 104.12 (C4), 107.85 (C7), 114.92 (C6), 131.72 (C5), 143.82 (C7a), 147.20 (C3a). ¹H-NMR (pyridine-d₅) δ: 5.95 (6H, s, H2), 6.88 (3H, d, J=8.2 Hz, H7), 7.41 (3H, dd, J=8.2, 1.5 Hz, H6), 7.84 (3H, s, H4), 8.64 (3H, br s, NH or HCl), 10.22 (3H, br s, HCl or NH). ¹³C-NMR (pyridine-d₅, 149.80 ppm) δ: 101.44 (C2), 104.39 (C4), 108.23 (C7), 114.34 (C6), 135.47 (C5), 143.47 (C7a), 148.11 (C3a), 165.46 (C2′). Positive-ion FAB-MS m/z: 487 (M+H⁺). HR-FAB-MS m/z: 487.1365 (Calcd for C₂₄H₁₉N₆O₆: 487.1366). Anal. Calcd for C₂₄H₁₈N₆O₆·HCl·0.7H₂O: C, 53.83; H, 3.84; N, 15.69. Found: C, 53.88; H, 3.83; N, 15.49.

1,1′,1″-(1,3,5-Triazine-2,4,6-trityl)tris[(piperidine-4-yl)methanol] (2d)

Entry 3: A mixture of 1 (0.74 g, 4.0 mmol) and 4-piperidinemethanol (dH, 2.76 g, 24 mmol) in dioxane (8 mL) was subjected to MW at 160°C (50 W) for 5 min with stirring. After evaporation of the solvent, the residue was dissolved in a mixture of EtOAc (40 mL) and aqueous saturated NaHCO₃ (40 mL) and then separated. The aqueous layer was extracted with EtOAc (40 mL×2). The combined organic layer was dried (MgSO₄). Evaporation of the solvent gave product 2d (1.17 g, 70%). Recrystallization from propionitrile gave 2d (0.86 g, 51%) as colorless crystals: mp 198–201°C (from EtCN). IR (KBr) cm⁻¹: 3400 (OH of alcohol), 1250 (C–N of t-amine), 1040 (C–O of alcohol). ¹H-NMR (DMSO-d₆) δ: 0.99 (6H, dm, J=12.5 Hz, H3′β, 5′β), 1.58 (3H, m, H4′), 1.66 (6H, dm, J=12.5 Hz, H3′α, 5′α), 2.70 (6H, dt, J=12.5, 2.1 Hz, H2′β, 6′β), 3.25 (6H, d, J=5.6 Hz, H1), 4.40 (3H, t, J=5.3 Hz, OH), 4.60 (6H, br d, J=12.5 Hz, H2′α, 6′α). ¹³C-NMR (DMSO-d₆) δ: 28.38 (C3′, 5′), 38.74 (C4′), 42.60 (C2′, 6′), 65.74 (C1), 164.68 (C2″). Positive-ion FAB-MS m/z: 421 (M+H⁺). HR-FAB-MS m/z: 421.2929 (Calcd for C₂₁H₃₇N₆O₃: 421.2927). Anal. Calcd for C₂₁H₃₆N₆O₃: C, 59.98; H, 8.63; N, 19.98. Found: C, 59.92; H, 8.59; N, 19.98.

1,1′-(6-Ethoxy-1,3,5-triazine-2,4-diyl)bis[(piperidin-4-yl)methanol] (3dq) and 1-(4,6-Diethoxy-1,3,5-triazine-2-yl)-(piperidine-4-yl)-methanol (4dqq)

Entry 4: In the same manner as that for the preparation of 2d (Entry 3), the reaction of 1 and dH was carried out in EtOH. After evaporation, the residue was purified by flash chromatography (CH₂Cl₂ : EtOH=90 : 10) to give 2d (24%), 3dq (36%), and 4dqq (4%) as a colorless oil. An analytical sample 3dq was obtained by recrystallization from AcOEt.

3dq: Colorless crystals, mp 133–134°C (from AcOEt). IR (KBr) cm⁻¹: 3345 (OH of alcohol), 1190, 1145 (C–N of t-amine), 1025 (C–O of alcohol). ¹H-NMR (CD₃OD) δ: 1.13 (4H, m, H3′β, 5′β), 1.33 (3H, t, J=7.0 Hz, H2‴), 1.73 (2H, m, H4′), 1.75 (4H, br t, J=12 Hz, H3′α, 5′α), 2.82 (4H, dt, J=12.8, 2.3 Hz, H2′β, 6′β), 3.41 (4H, d, J=6.1 Hz, H1), 4.33 (2H, q, J=7.0 Hz, H1‴), 4.71 (4H, br d, J=13 Hz, H2′α, 6′α). ¹³C-NMR (CD₃OD) δ: 14.86 (C2‴), 29.79 (C3′, 5′), 40.31 (C4′), 44.54 (C2′, 6′), 63.28 (C1‴), 67.79 (C1), 166.96 (C2″, 4″), 172.10 (C6″). Positive-ion FAB-MS m/z: 352 (M+H⁺). HR-FAB-MS m/z: 352.2347 (Calcd for C₁₇H₃₀N₅O₃: 352.2349). Anal. Calcd for C₁₇H₂₉N₅O₃: C, 58.10; H, 8.32; N, 19.93. Found: C, 57.89; H, 8.24; N, 19.76.

4dqq: ¹H-NMR (CD₃OD) δ: 1.19 (2H, m, H3′β, 5′β), 1.38 (6H, q, J=7.0 Hz, H2‴), 1.75 (1H, m, H4′α), 1.80 (2H, m, H3′α, 5′α), 2.92 (2H, dt, J=10.8, 2.4 Hz, H2′β, 6′β), 3.42 (2H, d, J=7.0 Hz, H1), 4.36 (4H, q, J=7.0 Hz, H1‴), 4.76 (2H, m, H2′α, 6′α). ¹³C-NMR (CD₃OD) δ: 14.67 (C2‴), 29.69 (C3′, 5′), 40.06 (C4′), 44.82 (C2′, 6′), 64.23 (C1‴), 67.55 (C1), 167.40 (C2″), 172.90 (C4″, 6″). Positive-ion FAB-MS m/z: 283 (M+H⁺). HR-FAB-MS m/z: 283.1778 (Calcd for C₁₃H₂₃N₄O₃; 283.1770).

1-(4,6-Dichloro-1,3,5-triazine-2-yl)-4-piperidinemethanol (6d)

Entry 5: In the same manner as that for the preparation of 2d (Entry 3), the reaction of 1 and dH was carried out in i-PrOH. After removal of the white precipitated solid (dH·HCl), the residue obtained by evaporation was purified by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=97 : 2.7 : 0.3→93 : 6.5 : 0.5) to give products 6d (11%) as a yellow oil, 5d (12%), and 3dr (2%).

6d: Positive-ion FAB-MS m/z: 263 (M+H⁺). HR-FAB-MS m/z: 263.0460 (Calcd for C₉H₁₃Cl₂N₄O: 263.0466). ¹H-NMR (CDCl₃) δ: 1.26 (2H, m, H3′β, 5′β), 1.44 (0.5H, br s, OH), 1.58 (0.5H, br s, OH), 1.84 (1H, m, H4′α), 1.89 (2H, dm, J=13.4 Hz, H3′α, 5′α), 2.97 (2H, dt, J=13.0, 2.7 Hz, H2′β, 6′β), 3.55 (2H, d, J=6.1 Hz, H1), 4.78 (2H, dt, J=6.7, 2.3 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 28.40 (C3′, 5′), 38.50 (C4′), 44.26 (C2′, 6′), 67.01 (C1), 163.75 (C2″), 170.26 (C4″, 6″).

1,1′-(6-Isopropoxy-1,3,5-triazine-2,4-diyl)bis[(piperidin-4-yl)methanol] (3dr)

Entry 6: In the same manner as that for the preparation of 2d (Entry 3), the reaction of 1 and dH was carried out with DIPEA in i-PrOH. After evaporation of the solvent, the resulting residue was purified by centrifugal chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=93 : 6.5 : 0.5) to give 5d (27%), 2d (8%), and 3dr (4%).

3dr: Positive-ion FAB-MS m/z: 366 (M+H⁺). HR-FAB-MS m/z: 366.2509 (Calcd for C₁₈H₃₂ N₅O₃: 366.2505). ¹³C-NMR (CDCl₃) δ: 21.93 (Me), 28.47 (C3′, 5′), 39.10 (C4′), 43.22 (C2′, 6′), 67.66 (C1), 69.02 (C1‴), 165.79 (C2″, 4″), 171.03 (C6″).

1,1′-(6-Chloro-1,3,5-triazine-2,4-diyl)bis[(piperidin-4-yl)methanol] (5d)

Entry 7: In the same manner as that described in Entry 6 except for the ratio of reagents (dH and DIPEA), the reaction was carried out under the conditions shown in Table 1. Purification by open column chromatography (n-hexane : EtOAc=20 : 80→0 : 100) gave 5d (88%), 2d (9%), and 3dr (1%). An analytical sample 5d was obtained by recrystallization from CH₂Cl₂ as white powder.

Entry 8: In the same manner as that in Entry 7 except for the use of n-heptanol instead of i-PrOH, the reaction was carried out. Purification of the products by open column chromatography (n-hexane : EtOAc=20 : 80→0 : 100) gave 5d (26%) and 3ds (3%).

Entry 9: To a solution of 1 (10.0 mmol) in dry CH₂Cl₂ (20 mL) was added a solution of dH (20 mmol) and TEA (20 mmol) in dry CH₂Cl₂ (20 mL). After stirring for 1 h at 0°C, the reaction mixture was allowed to warm up to room temperature and then stirred for another 17 h. After addition of a solution of KOH (100 mmol) in water (60 mL), the resulting mixture was extracted with CH₂Cl₂ (30 mL×2) and then the organic layer was washed with aqueous NaHCO₃ (20 mL) and brine (20 mL×2). The organic layer was dried and the solvent was evaporated to give crude 5d as an oil, which was crystallized by addition of CH₂Cl₂ to form a white solid. Recrystallization of crude 5d from CH₂Cl₂ gave an analytical pure sample 5d (57%) as a white powder.

5d: mp 154–155°C (from CH₂Cl₂). IR (KBr) cm⁻¹: 3400 (OH), 1570 (N=C). ¹H-NMR (CDCl₃) δ: 1.20 (4H, dq, J=7.3, 4.3 Hz, H3′β, 5′β), 1.45 (2H, br s, OH), 1.76 (2H, m, H4′α), 1.79 (4H, t, J=11.9 Hz, H3′α, 5′α), 2.83 (4H, dt, J=13.1, 2.1 Hz, H2′β, 6′β), 3.52 (4H, d, J=5.8 Hz, H1), 4.73 (4H, d, J=13.1 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 28.53 (C3′, 5′), 38.94 (C4′), 43.46 (C2′, 6′), 67.48 (C1), 164.32 (C2″, 4″), 169.61 (C6″). Positive-ion FAB-MS m/z: 342 (M+H⁺). HR-FAB-MS m/z: 342.1689 (Calcd for C₁₅H₂₅ClN₅O₂: 342.1697). Anal. Calcd for C₁₅H₂₄ ClN₅O₂: C, 52.70; H, 7.08; N, 20.49. Found: C, 52.58; H, 7.12; N, 20.20.

1,1′,1″-(1,3,5-Triazine-2,4,6-triyl)tris-3-piperidinol (2e²⁷⁾)

Entry 10: A mixture of 1 (10 mmol) and 3-hydroxypiperidine (eH, 60 mmol) was heated. After addition of water (100 mL) and then ether (100 mL), the insoluble solid material was filtered and then the obtained product was dissolved in MeOH (300 mL). After addition of ether (100 mL), the resulting precipitates were collected. Recrystallization of the crude product from EtOH gave pure product 2e (29%) as colorless crystals: mp 264–265°C (from EtOH). IR (KBr) cm⁻¹: 3340 (OH of alcohol), 1230 (C–N of t-amine), 1060 (C–O of alcohol). ¹H-NMR (DMSO-d₆) δ: 1.32 (6H, m, H4, 5), 1.67 (3H, m, H5), 1.88 (3H, m, H4), 2.68 (3H, m, H2), 2.83 (3H, m, H6), 3.38 (3H, m, H3), 4.28 (3H, m, H6), 4.43 (3H, m, H2), 4.79 (3H, m, OH). ¹³C-NMR (DMSO-d₆) δ: 22.90 (C5), 33.50 (C4), 42.46 (C6), 49.91 (C2), 65.27 (C3), 164.72 (C2′). Positive-ion FAB-MS m/z: 379 (M+H⁺). HR-FAB-MS m/z: 379.2456 (Calcd for C₁₈H₃₁N₆O₃: 379.2458). Anal. Calcd for C₁₈H₃₀N₆O₃: C, 57.12; H, 7.99; N, 22.21. Found: C, 57.03; H, 7.96; N, 22.01.

3,3′,3″,3‴-[(6-Chloro-1,3,5-triazine-2,4-diyl)-dinitrilo]tetrakispropanenitrile (5f) [856812-02-7]²⁰⁾

Entry 11: In a manner similar to that described in Entry 9, the reaction of 1 with fH was carried out. Evaporation of the solvent gave crude product 5f as a pale yellow solid, which was washed with CH₂Cl₂ (15 mL) to give 5f (33%) as a white solid. An analytical sample 5f was obtained by recrystallization from CH₂Cl₂ as colorless needles: mp 156–157°C (from CH₂Cl₂) [lit.²⁰⁾ mp 149–151°C]. IR (KBr) cm⁻¹: 2250 (CN), 805 (C–Cl). ¹H-NMR (DMSO-d₆) δ: 2.84 (4H, t, J=6.7 Hz, H2_A), 2.88 (4H, t, J=6.7 Hz, H2_B), 3.84 (4H, t, J=6.7 Hz, H3_A), 3.89 (4H, t, J=6.7 Hz, H3_B). ¹³C-NMR (DMSO-d₆) δ: 15.52 (C2_B), 15.79 (C2_A), 42.86 (C3_A), 43.06 (C3_B), 118.84 (CN_A), 119.08 (CN_B), 164.34 (C2′), 168.58 (C6′). Positive-ion FAB-MS m/z: 358 (M+H⁺). HR-FAB-MS m/z: 358.1299 (Calcd for C₁₅H₁₇N₉Cl: 358.1295). Anal. Calcd for C₁₅H₁₆N₉Cl·0.5H₂O: C, 49.12; H, 4.67; N, 34.37. Found: C, 48.95; H, 4.47; N, 34.60.

2-Chloro-4,6-bis(4-methyl-1-piperazinyl)-1,3,5-triazine Trihydrochloride (5g·3HCl)

Entry 12: To a solution of 1 (1.0 mmol) in dry tetrahydrofuran (THF, 6 mL) was added 1-methylpiperazine (gH, 3.0 mmol) with stirring at room temperature. After removal of the precipitated solid, 1 M HCl/EtOH was added to the filtrate, and filtration of the resulting material gave product 5g·3HCl in 20% yield as colorless crystals.

5g·3HCl: ¹H-NMR (CDCl₃) δ: 2.35 (6H, s, CH₃), 2.73 (8H, m, H3, 5), 3.22 (8H, m, H2, 6). ¹³C-NMR (CDCl₃) δ: 43.37 (C2, 6), 45.71 (CH₃), 51.50 (C3, 5), 164.34 (C2′, 4′), 169.57 (C6′). Positive-ion FAB-MS m/z: 312 (M+H⁺). HR-FAB-MS m/z: 312.1710 (Calcd for C₁₃H₂₃N₇Cl for 312.1703).

2,2′-(1,4-Piperazinediyl)bis[4,6-di(4-methyl-1-piperazinyl)-1,3,5-triazine] (Ag)

Entry 13: To a solution of gH (33.0 mmol) in AcOH (70 mL) was added 1 (1.84 g, 10.0 mmol), and the resulting mixture was refluxed for 1 d. The precipitated solids were collected and washed with hot water (20 mL×2). The obtained precipitate was dissolved in 10% NaOH (20 mL) and the resulting solution was extracted with CH₂Cl₂ (40 mL×2). The organic layer was washed with brine (30 mL) and dried over MgSO₄. After removal of the solvent, Ag (46%) was obtained as a pale yellow solid. Recrystallization from EtOH–H₂O gave the dimeric TAZ derivative Ag as a white powder.

Ag: mp 255–257°C decn. (from EtOH–H₂O). ¹H-NMR (CDCl₃) δ: 2.31 (12H, s, CH₃), 2.40 (16H, t, J=5.0 Hz, H3′, 5′), 3.78a) (8H, s, Ha), 3.80a) (16H, m, H2′, 6′). ¹³C-NMR (CDCl₃) δ: 43.04 (C2′, 6′), 43.14 (Ca), 46.23 (CH₃), 55.00 (C3′, 5′), 165.39 (C4, 6), 165.53 (C2). Positive-ion FAB-MS m/z: 637 (M+H⁺). HR-FAB-MS m/z: 637.4651 (Calcd for C₃₀H₅₃N₁₆Cl for 637.4639).

General Procedure for Stepwise Substitution of TCTAZ with Alcohols and Amines

(Step 1) To a solution of 1 (5–20 mmol) in dry acetone (7–60 mL) were added an alcohol (ROH) and collidine. After stirring for 1–3 h at 0°C or at room temperature, the precipitated collidine·HCl was removed by filtration. Evaporation of the solvent gave crude alkoxy-chloro-TAZ (7 or 8). (Step 2) This crude product was dissolved in dry dioxane (7–60 mL), and an amine with or without DIPEA was added. Then stirring was continued at room temperature, or the resulting mixture was subjected to MW at 150–160°C (50 W) under the conditions shown in Table 2. After evaporation of the solvent, the residue was purified by chromatography or recrystallization. A typical protocol for the preparation of compound 3ds (Entry 14) is shown below in detail.

1,1′-(6-Heptoxy-1,3,5-triazine-2,4-diyl)bis[(piperidin-4-yl)methanol] (3ds), [1-[4,6-Bis(heptyloxy)-1,3,5-triazin-2-yl]piperidin-4-yl]methanol (4dss), and 2,4,6-Tris(heptyloxy)-1,3,5-triazine (9sss) [37068-43-2]²⁴⁾

Entry 14: (Step 1) To a solution of 1 (1.02 g, 5.5 mmol) in dry acetone (7 mL) were added sH (0.843 g, 7.25 mmol) and collidine (0.69 g, 5.7 mmol) at 0°C. After stirring for 1 h at 0°C, the resulting white precipitated salt (collidine·HCl) was removed by filtration. After evaporation of the solvent, a dark orange solid was obtained. (Step 2) This material was dissolved in dry dioxane (5 mL), and dH (2.53 g, 22.0 mmol) was added, and then the resulting mixture was subjected to MW at 160°C (50 W) for 10 min with stirring. After evaporation of the solvent, the residue was purified by flash chromatography (CH₂Cl₂ : 95%EtOH : 28%NH₃=95 : 4.5 : 0.5→93 : 6.5 : 0.5) to give 3ds (1.69 g, 73% yield) as a white solid, 4dss (93 mg, 4% yield), and 9sss (23 mg, 1% yield).

3ds: mp 122–123°C (from CH₃CN). IR (KBr) cm⁻¹: 3420 (OH of alcohol), 1355, 1050 (C–N of t-amine). ¹H-NMR (CDCl₃) δ: 0.88 (3H, t, J=7.0 Hz, H7‴), 1.18 (4H, m, H3′β, 5′β), 1.29 (6H, m, H5‴, 6‴, 4‴), 1.40 (2H, m, H3‴), 1.47 (2H, br s, OH), 1.76 (8H, m, H2‴, 4′α, 3′α, 5′α), 2.80 (4H, dt, J=12.8, 2.1 Hz, H2′β, 6′β) 3.51 (4H, d, J=5.8 Hz, H1) 4.26 (2H, t, J=6.9 Hz, H1‴), 4.79 (4H, br d, J=12.8 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 14.06 (C7‴), 22.58 (C6‴), 25.96 (C3‴), 28.57 (C3′, 5′), 28.93, 29.07 (C2‴, 4‴), 31.75 (C5‴), 39.12 (C4′), 43.24 (C2′, 6′), 66.57 (C1‴), 67.69 (C1), 165.82 (C2″, 4″), 170.91 (C6″). Positive-ion FAB-MS m/z: 422 (M+H⁺). HR-FAB-MS m/z: 422.3141 (Calcd for C₂₂H₄₀N₅O₃: 422.3131). Anal. Calcd for C₂₂H₃₉N₅O₃: C, 62.68; H, 9.32; N, 16.61. Found: C, 62.45; H, 9.15; N, 16.60.

4dss: mp 68–70°C (from CH₃CN). IR (KBr) cm⁻¹: 3365 (OH of alcohol), 1250, 1130, 1045 (C–O alcohol and ether). ¹H-NMR (CDCl₃) δ: 0.88 (6H, t, J=7.0 Hz, H7‴), 1.20 (2H, m, H3′β, 5′β), 1.25–1.35 (12H, m, H4‴, 5‴, 6‴), 1.41 (4H, m, H3‴), 1.60 (1H, br s, OH), 1.73–1.83 (7H, m, H2‴, 4′α, 3′α, 5′α), 2.86 (2H, dt, J=13.1, 2.1 Hz, H2′β, 6′β), 3.52 (2H, d, J=6.1 Hz, H1), 4.31 (4H, t, J=6.9 Hz, H1‴), 4.81 (2H, dm, J=13.1 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 14.04 (C7‴), 22.57 (C6‴), 25.87 (C3‴), 28.53, 28.79, 29.00 (C3′, 5′, 2‴, 4‴), 31.73 (C5‴), 38.95 (C4′), 43.55 (C2′, 6′), 67.49 (C1), 67.49 (C1‴), 166.39 (C2″, 4″), 171.90 (C6″). Positive-ion FAB-MS m/z: 423 (M+H⁺). HR-FAB-MS m/z: 423.3339 (Calcd for C₂₃H₄₃N₄O₃: 423.3335). Anal. Calcd for C₂₃H₄₂N₄O₃: C, 65.37; H, 10.02; N, 13.26. Found: C, 65.43; H, 10.05; N, 13.08.

9sss: Positive-ion FAB-MS m/z: 424 (M+H⁺). HR-FAB-MS m/z: 424.3538 (Calcd for C₂₄H₄₆N₃O₃: 424.3539). ¹H-NMR (CDCl₃) δ: 0.88 (9H, t, J=7.0 Hz, H7′), 1.25–1.35 (18H, m, H5′, 6′, 4′), 1.43 (6H, m, H3′), 1.78 (6H, m, H2′), 4.38 (6H, t, J=6.7 Hz, H1′). ¹³C-NMR (CDCl₃) δ: 13.96 (C7′), 22.53 (C6′), 25.75 (C3′), 28.64 (C2′), 28.91 (C4′), 31.69 (C5′), 68.43 (C1′), 173.15 (C2, 4, 6).

Entry 15: (Step 1) The reaction was carried out in the same manner as that described above procedure (Entry 14, Step 1) except for the ratio of reagents [alcohol (sH) and collidine]. (Step 2) The residue was dissolved in dry acetonitrile, and dH and DIPEA were added, and then the resulting mixture was stirred at room temperature for 6 h. After evaporation of the solvent, the residue was purified by flash chromatography (n-hexane : EtOAc=70 : 30→60 : 40) to give 3ds (1%) as a white solid and 4dss (47%). Analytical samples of 3ds and 4dss were obtained by recrystallization from acetonitrile as colorless crystals.

2,4-Dichloro-6-(heptyloxy)-1,3,5-triazine (7s) [107392-85-8]²³⁾

Entry 16: In a manner similar to that for preparation of compound 3ds (Entry 14, Step 1), the reaction of 1 (22 mmol), sH (29 mmol) and collidine (23 mmol) was carried at 0°C for 1 h. After removal of the salt (collidine·HCl), ice-water (40 mL) was added to the filtrate and the mixture was extracted with CH₂Cl₂ (20 mL×3). The organic layer was dried, and filtrated. The filtrate was evaporated and the residue was purified by open column chromatography (CH₂Cl₂) to give 7s (70%) as a colorless oil and 9sss (1%) as a white solid.

7s: ¹H-NMR (CDCl₃) δ: 0.89 (3H, t, J=7.0 Hz, H7′), 1.31 (4H, m, H5′, 6′), 1.35 (2H, m, H4′), 1.44 (2H, m, H3′), 1.82 (2H, m, H2′), 4.49 (2H, t, J=6.6 Hz, H1′). ¹³C-NMR (CDCl₃) δ: 13.99 (C7′), 22.51 (C6′), 25.55 (C3′), 28.27 (C2′), 28.77 (C4′), 31.60 (C5′), 70.68 (C1′), 171.05 (C1), 172.45 (C3, 5). Positive-ion FAB-MS m/z: 264 (M+H⁺). HR-FAB-MS m/z: 264.0618 (Calcd for C₁₀H₁₆Cl₂N₃O: 264.0670).

Reactions of 7s with Amines

The reaction of 7s with various nucleophiles (dH, eH, hH, and iH) under the conditions shown in Table 2 afforded compound 3ds (85%), 3es (92%), 3hs (69%), and 3is (37%), respectively (see Entries 17–20). A typical protocol for the preparation of 3ds is shown below in detail. Physical and spectroscopic data of these compounds are shown below.

Entry 17: A solution of 7s (12.0 mmol) and dH (75.7 mmol) in dry dioxane (15 mL) was subjected to MW with stirring. The obtained product was separated by flash chromatography (n-hexane : EtOAc=40 : 60→20 : 80) to give 3ds (85%) as a white solid.

1,1′-[6-(Heptyloxy)-1,3,5-triazine-2,4-diyl]bis(piperidin-3-ol) (3es²⁷⁾)

Entry 18: Colorless crystals, mp 126–127°C (CH₃CN). IR (KBr) cm⁻¹: 3420 (OH of alcohol), 1375, 1510 (C–N of t-amine), 1105 (C–O). ¹H-NMR (pyridine-d₅) δ: 0.83 (3H, t, J=7.0, H7″), 1.15–1.25 (6H, m, H4″, 5″, 6″), 1.38 (2H, m, H3″), 1.50 (2H, m, H5α or β), 1.65–1.85 (6H, m, H2″, 4α or β, 5β or α), 2.13 (2H, m, H4β or α), 3.19 (2H, m, H6α or β), 3.38 (2H, m, H2α or β), 3.96 (2H, m, H3), 4.36 (2H, br s, H1″), 4.50 (2H, br s, H6β or α), 4.87 (2H, br d, J=10.4 Hz, H2β or α), 4.90 (2H, br s, OH). ¹³C-NMR (pyridine-d₅) δ: 14.11 (C7″), 22.71 (C6″), 23.53 (C5), 26.29 (C3″), 29.20, 29.42 (C2″, 4″), 31.88 (C5″), 34.28 (C4), 43.65 (C6), 51.23 (C2), 66.28, 66.36 (C1″, 3), 166.82 (C2′, 4′), 171.63 (C6′). Positive-ion FAB-MS m/z: 394 (M+H⁺). HR-FAB-MS m/z: 394.2822 (Calcd for C₂₀H₃₆N₅O₃: 394.2818). Anal. Calcd for C₂₀H₃₅N₅O₃: C, 58.89; H, 9.04; N, 17.17. Found: C, 58.95; H, 9.04; N, 17.11.

3,3′-[1,1′-(6-(Heptyloxy)-1,3,5-triazine-2,4-diyl)bis(piperidine-4,1-diyl)]diphenol (3hs)

Entry 19: White solid, mp 79–80°C. IR (KBr) cm⁻¹: 3400 (O–H of ArOH), 1575, 1515 (C–N of t-amine). ¹H-NMR (CDCl₃) δ: 0.86 (3H, t, J=6.9 Hz, H7‴), 1.26 (6H, m, H4‴, 5‴, 6‴), 1.39 (2H, m, H3‴), 1.61 (4H, m, H3′, 5′), 1.73 (2H, m, H2‴), 1.85 [6H, br d, J=12.5 Hz, H3′, 5′ (4H), OH (2H)], 2.68 (2H, br t, J=12.5 Hz, H4′), 2.86 (4H, t, J=12.5 Hz, H2′β, 6′β), 4.28 (2H, t, J=6.9 Hz, H1‴), 4.90 (4H, br d, J=13.1 Hz, H2′α, 6′α), 5.74 (1H, br s, H₂O), 6.66 (2H, s, H2), 6.67 (2H, d, J=7.6 Hz, H4), 6.74 (2H, d, J=7.6 Hz, H6), 7.14 (2H, t, J=7.6 Hz, H5). ¹³C-NMR (CDCl₃) δ: 14.02 (C7‴), 22.57 (C6‴), 25.95 (C3‴), 28.95 (C2‴ or C4‴), 29.05 (C4‴ or C2‴), 31.74 (C5‴), 33.08 (C3′, 5′), 42.96 (C4′), 44.10 (C2′, 6′), 66.73 (C1‴), 113.37 (C4), 113.81 (C2), 119.07 (C6), 129.58 (C5), 147.76 (C1), 156.02 (C3), 165.80 (C2″, 4″), 170.99 (C6″). Positive-ion FAB-MS m/z: 546 (M+H⁺). HR-FAB-MS m/z: 546.3441 (Calcd for C₃₂H₄₄N₅O₃: 546.3444). Anal. Calcd for C₃₂H₄₃N₅O₃·0.5H₂O: C, 69.29; H, 7.99; N, 12.62. Found: C, 69.16; H, 8.01; N, 12.43.

4,4′-[6-(Heptyloxy)-1,3,5-triazine-2,4-diyl]bis[2-[2-(1-piperazinyl)-ethoxy]ethanol] (3is)

Entry 20: Yellow oil. IR (KBr) cm⁻¹: 3375 (OH of alcohol), 1570, 1525 (C–N of t-amine), 1120 (C–O). ¹H-NMR (CDCl₃) δ: 0.88 (3H, t, J=7.0 Hz, H7‴), 1.29 (6H, m, H4‴, 5‴, 6‴), 1.40 (2H, m, H3‴), 1.74 (2H, m, H2‴), 2.54 (8H, t, J=5.0 Hz, H3′, 5′), 2.62 (4H, t, J=5.2 Hz, H5), 2.93 (2H, br s, OH), 3.61 (4H, t, J=4.6 Hz, H2), 3.68 (4H, t, J=5.2 Hz, H4), 3.71 (4H, t, J=4.6 Hz, H1), 3.83 (8H, t, J=5.0 Hz, H2′, 6′), 4.25 (2H, t, J=6.9 Hz, H1‴). ¹³C-NMR (CDCl₃) δ: 14.00 (C7‴), 22.55 (C6‴), 25.94 (C3‴), 28.90 (C2‴ or C4‴), 29.02 (C4‴ or C2‴), 31.73 (C5‴), 42.93 (C2′, 6′), 53.20 (C3′, 5′), 57.99 (C5), 61.91 (C1), 66.69 (C1‴), 67.75 (C4), 72.39 (C2), 165.97 (C2″, 4″), 170.95 (C6″). Positive-ion FAB-MS m/z: 540 (M+H⁺). HR-FAB-MS m/z: 540.3881 (Calcd for C₂₆H₅₀N₇O₅: 540.3873). Anal. Calcd for C₂₆H₄₉N₇O₅·0.2H₂O: C, 57.48; H, 9.16; N, 18.05. Found: C, 57.38; H, 9.37; N, 18.15.

1,1′-[6-(1,3-Benzodioxol-5-yloxy)-1,3,5-triazine-2,4-diyl]bis-4-piperidinemethanol (3dt)

Entry 21: This compound was prepared in a manner similar to that for compound 3ds (Entry 14). Purification by flash chromatography (CH₂Cl₂ : EtOH=95 : 5→93 : 7) gave 3dt (74%) as a yellow solid. An analytical sample was obtained by recrystallization from propionitrile as colorless crystals: mp 165–166°C (from C₂H₅CN). IR (KBr) cm⁻¹: 1585, 1505 (C–N of t-amine) 1035 (C–O of alcohol). ¹H-NMR (CDCl₃) δ: 1.18 (4H, m, H3′β, 5′β), 1.52 (2H, br s, OH), 1.75 (6H, m, H4′α, 3′α, 5′α), 2.78 (4H, t, J=12.5 Hz, H2′β, 6′β), 3.50 (4H, d, J=6.1 Hz, H1), 4.68 (4H, br s, H2′α, 6′α), 5.97 (2H, s, H2‴), 6.62 (1H, d, J=2.4, 8.5 Hz, H6‴), 6.70 (1H, d, J=2.4 Hz, H4‴), 6.74 (1H, d, J=8.5 Hz, H7‴). ¹³C-NMR (CDCl₃) δ: 28.53 (C3′, 5′), 39.06 (C4′), 43.34 (C2′, 6′), 67.61 (C1), 101.39 (C2‴), 104.28 (C4‴), 107.44 (C7‴), 114.29 (C6‴), 144.35 (C7a‴), 147.23 (C3a‴ or 5‴), 147.45 (C5‴ or 3a‴), 165.89 (C2″, 4″), 171.20 (C6″). Positive-ion FAB-MS m/z: 444 (M+H⁺). HR-FAB-MS m/z: 444.2251 (Calcd for C₂₂H₃₀N₅O₅: 444.2247). Anal. Calcd for C₂₂H₂₉N₅O₅: C, 59.58; H, 6.59; N, 15.79. Found: C, 59.57; H, 6.70; N, 15.95.

1-(4,6-Dimethoxy-1,3,5-triazine-2-yl)-4-piperidinemethanol (4dpp), 1-[4-(1,3-Benzodioxol-5-yloxy)-6-methoxy-1,3,5-triazine-2-yl]-4-piperidinemethanol (4dpt), 2-(1,3-Benzodioxol-5-yloxy)-4,6-dimethoxy-1,3,5-triazine (9 ppt), and 2,4-Bis(1,3-benzodioxol-5-yloxy)-6-methoxy-1,3,5-triazine (9ptt)

Entry 22: (Step 1) To a solution of 1 (4.06 g, 22 mmol) in MeOH (60 mL) were added tH (2.76 g, 20 mmol) and NaOH (3.20 g, 80 mmol) in water (40 mL) at room temperature, and the resulting mixture was stirred for 2 h. The separated solid was filtrated. (Step 2) The mixture of this solid and dH (1.15 g, 100 mmol) in dry dioxane (30 mL) was subjected to MW with stirring. After evaporation of the solvent, CHCl₃ and MeOH (both 30 mL) were added to the residue and stirred vigorously. The separated solid was removed by filtration and then the solvent was evaporated. Purification of the residue by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=997 : 2.7 : 0.3→970 : 27 : 3) gave 9 ppt (1.24 g, 21%) as a white solid, 9ptt (1.86 g, 22%) as a white solid, 4dpt (638 mg, 8% yield) as a pale yellow solid, and 4dpp (1.11 g, 20% yield) as a white solid. An analytical sample 9 ppt was obtained by recrystallization from MeOH.

4dpp: mp 116–118°C. IR (KBr) cm⁻¹: 3335 (OH of alcohol), 1360, 1130 (C–N of t-amine). ¹H-NMR (CDCl₃) δ: 1.2 (2H, m, H3′β, 5′β), 1.63 (1H, br s, OH), 1.8 (3H, m, H3′α, 4′α, 5′α), 2.88 (2H, dt, J=13.1, 2.7 Hz, H2′β, 6′β), 3.53 (2H, d, J=6.1 Hz, H1), 3.95 (6H, s, MeO), 4.82 (2H, dm, J=13.1 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 28.49 (C3′, 5′), 38.91 (C4′), 43.59 (C2′, 6′), 54.40 (MeO), 67.40 (C1), 166.32 (C2″), 172.27 (C4″, 6″). Positive-ion FAB-MS m/z: 255 (M+H⁺). HR-FAB-MS m/z: 255.1454 (Calcd for C₁₁H₁₉N₄O₃: 255.1457). Anal. Calcd for C₁₁H₁₈N₄O₃: C, 51.96; H, 7.13; N, 22.03. Found: C, 51.97; H, 7.17; N, 21.79.

4dpt: ¹H-NMR (CDCl₃) δ: 1.19 (2H, m, H3′β, 5′β), 1.75–1.85 (3H, m, H3′α, 4′α, 5′α), 2.86 (2H, m, H2′β, 6′β), 3.52 (2H, d, J=6.1 Hz, H1), 3.90 (3H, s, OMe), 4.65 (1H, d, J=13.1 Hz, H2′α or 6′α), 4.81 (1H, d, J=13.1 Hz, H6′α or 2′α), 5.98 (2H, s, H2‴), 6.61 (1H, dd, J=8.5, 2.4 Hz, H6‴), 6.68 (1H, d, J=2.4 Hz, H4‴), 6.76 (1H, d, J=8.5 Hz, H7‴). ¹³C-NMR (CDCl₃) δ: 28.42 (C3′, 5′), 38.76 (C4′), 43.64 (C2′, 6′), 54.50 (MeO), 67.25 (C1), 101.53 (C2‴), 103.96 (C4‴), 107.69 (C7‴), 114.08 (C6‴), 144.89 (C7a‴), 146.50 (C5‴), 147.70 (C3a‴), 166.30 (C2″), 172.22 (C4″ or C6″), 172.38 (C6″ or C4″). Positive-ion FAB-MS m/z: 361 (M+H⁺). HR-FAB-MS m/z: 361.1516 (Calcd for C₁₇H₂₁N₄O₅: 361.1512).

9ppt: ¹H-NMR (CDCl₃) δ: 4.01 (6H, s, OCH₃), 6.00 (2H, s, H2), 6.62 (1H, dd, J=8.2, 2.4 Hz, H6), 6.68 (1H, d, J=2.4 Hz, H4), 6.79 (1H, d, J=8.2 Hz, H7). ¹³C-NMR (CDCl₃) δ: 55.47 (OCH₃), 101.75 (C2), 103.71 (C4), 107.96 (C7), 113.91 (C6), 145.51 (C7a), 146.05 (C5), 148.03 (C3a), 173.55 (C4′, 6′), 174.00 (C2′). Positive-ion FAB-MS m/z: 278 (M+H⁺). HR-FAB-MS m/z: 278.0775 (Calcd for C₁₂H₁₂N₃O₅: 278.0777).

9ptt: mp 156–157°C (from MeOH). IR (KBr) cm⁻¹: 1355, 1170 (C–N of t-amine). ¹H-NMR (CDCl₃) δ: 3.96 (3H, s, OCH₃), 5.99 (4H, s, H2), 6.61 (2H, dd, J=8.5, 2.4 Hz, H6), 6.66 (2H, d, J=2.4 Hz, H4), 6.78 (2H, d, J=8.5 Hz, H7). ¹³C-NMR (CDCl₃) δ: 55.63 (OCH₃), 101.78 (C2), 103.68 (C4), 107.98 (C7), 113.88 (C6), 145.58 (C7a), 145.99 (C5), 148.04 (C3a), 173.81 (C6′) 176.26 (C2′, 4′). Positive-ion FAB-MS m/z: 384 (M+H⁺). HR-FAB-MS m/z: 384.0829 (Calcd for C₁₈H₁₄N₃O₇: 384.0832). Anal. Calcd for C₁₈H₁₃N₃O₇: C, 56.40; H, 3.42; N, 10.96. Found: C, 56.18; H, 3.59; N, 11.09.

[1,1′-[6-(4-Hydroxyphenoxy)-1,3,5-triazine-2,4-diyl]bis(piperidine-4,1-diyl)]dimethanol (3du), [1,1′,1″,1‴-[6,6′-[1,4-Phenylenebis(oxy)]-bis(1,3,5-triazine-6,4,2-triyl)]tetrakis(piperidine-4,1-diyl)]tetramethanol (Bdu) and [1,1′-[6-[4-[[4-[4-(Hydroxymethyl)piperidin-1-yl]-6-(4-hydroxyphenoxy)-1,3,5-triazin-2-yl]oxy]phenoxy]-1,3,5-triazine-2,4-diyl]-bis(piperidine-4,1-diyl)]dimethanol (Cdu)

Entry 23: This compound was prepared in a manner similar to that for compound 3ds (Entry 14). Purification of products by flash chromatography (CH₂Cl₂ : EtOH=90 : 10→85 : 15→80 : 20) gave 3du (29%) as a brown solid, Bdu (9%) as a white solid, and Cdu (3%) as a brown solid.

3du: mp 228–229°C. IR (KBr) cm⁻¹: 3400 (OH of alcohol), 1580, 1500 (C–N of t-amine), 1200, 1035 (C–O). ¹H-NMR (DMSO-d₆) δ: 1.01 (4H, m, H3′β, 5′β), 1.62 (2H, m, H4′α), 1.64 (4H, m, H3′α, 5′α), 2.50 (4H, s, H2′β, 6′β), 3.25 (4H, t, J=5.6 Hz, H1), 4.4 (2H, br s, H2′α, 6′α), 4.41 (2H, t, J=5.3 Hz, C1-OH), 4.57 (2H, br s, H2′α, 6′α), 6.73 (2H, d, J=9.0 Hz, H3‴, 5‴), 6.92 (2H, d, J=9.0 Hz, H2‴, 6‴), 9.25 (1H, s, ArOH). ¹³C-NMR (DMSO-d₆) δ: 28.26 (C3′, 5′), 38.48 (C4′), 42.77 (C2′, 6′), 65.54 (C1), 115.14 (C3‴, 5‴), 122.32 (C2‴, 6‴), 144.52 (C1‴), 154.02 (C4‴), 165.34 (C2″, 4″), 170.83 (C6″). Positive-ion FAB-MS m/z: 416 (M+H⁺). HR-FAB-MS m/z: 416.2303 (Calcd for C₂₁H₃₀N₅O₄: 416.2303). Anal. Calcd for C₂₁H₂₉N₅O₄: C, 60.71; H, 7.04; N, 16.86. Found: C, 60.45; H, 7.27; N, 16.57.

Bdu: mp 249–251°C. IR (KBr) cm⁻¹: 3385 (OH of alcohol), 1580, 1490 (C–N of t-amine), 1195, 1040 (C–O).¹H-NMR (DMSO-d₆) δ: 1.01 (8H, br s, H3′β, 5′β), 1.62 (12H, m, H3′α, 4′α, 5′α), 2.77 (8H, br s, H2′β, 6′β), 3.25 (8H, t, J=5.5 Hz, H1), 4.42 (4H, t, J=5.5 Hz, OH), 4.4 (4H, br s, H2′α, 6′α), 4.59 (4H, br s, H2′α, 6′α), 7.14 (4H, s, H2‴, 3‴, 5‴, 6‴). ¹³C-NMR (DMSO-d₆) δ: 28.26 (C3′, 5′), 38.45 (C4′), 42.80 (C2′, 6′), 65.51 (C1), 122.07 (C2‴, 3‴), 148.85 (C1‴, 4‴), 165.29 (C2″, 4″), 170.51 (C6″). Positive-ion FAB-MS m/z: 721 (M+H⁺). HR-FAB-MS m/z: 721.4150 (Calcd for C₃₆H₅₃N₁₀O₆: 721.4144).

Cdu: ¹H-NMR (DMSO-d₆) δ: 1.0 (6H, m, H3, 5 in A, B), 1.65 (9H, m, H4, 3, 5 in A, B), 2.75 (6H, m, H2, 6 in A, B), 3.25 (6H, br s, CH₂OH), [4.4 (m) and 4.6 (m) (6H, H2, 6 in A, B)], 4,43 (3H, br s, CH₂OH), 6.76 (2H, d, J=9.0 Hz, H3, 5 in E), 6.98 (2H, d, J=9.0 Hz, H2, 6 in E), 7.19 (4H, m, H2, 3, 5, 6 in F), 9.37 (1H, br s, ArOH). ¹³C-NMR (DMSO-d₆) δ: 28.10 (C3, 5 in A), 28.26 (C3, 5 in B), 38.13 (C4 in A), 38.46 (C4 in B), 43.08, 43.12 (C2, 6 in A, B), 65.30 (CH₂OH in A), 65.53 (CH₂OH in B),115.37 (C3, 5 in E), 122.02 (C2, 3, 5, 6 in F), 122.23 (C2, 6 in E), 144.04 (C1 in E), 148.36, 149.35 (C1, 4 in F), 154.61 (C4 in E), 165.28 (C2, 4 in D), 165.75 (C2 in C), 170.48, 171.75, 172.15 (C4, 6 in C, C6 in D). Positive-ion FAB-MS m/z: 716 (M+H⁺). HR-FAB-MS m/z: 716.3520 (Calcd for C₃₆H₄₆N₉O₇: 716.3522).

Antiviral Activity Assay and Cytotoxicity of Target Compounds

The antiviral activities of synthesized compounds were measured by using a plaque reduction assay²⁵⁾ as described in our previous paper.³⁾ Results for antiviral activity (EC₅₀) and cytotoxicity (IC₅₀) with Vero cells are summarized in Table 3.

Acknowledgment

Appreciation is expressed to Ms. Airi Koga for skillful technical assistance.

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• 27) We used a racemic 3-hydroxylated piperizine derivative (eH) as a starting material in the synthesis of C₃- or C_S-symmetric type target TAZ derivatives. The obtained symmetric products (2e, 3es) showed very simple symmetric ¹³C-NMR in DMSO-d₆ or pyridine-d₅, indicating little difference with respect to the signals assignable to symmetric trisubstituted TAZs. From a stereochemical viewpoint, however, the obtained C₃-type product (2e) can be considered to be a mixture of four stereoisomers, i.e., two C₃-symmetric molecules that have the same absolute configuration regarding three introduced chiral piperidines (R,R,R or S,S,S) in the molecules and a set of two enantiomers (R,R,S and S,S,R). The trisubstituted symmetric TAZ derivative (3es) that has two chiral piperidines in the molecule is also expected to be a mixture of three stereoisomers, i.e., a C_S-symmetrical meso-compound having different absolute configurations (R and S) with reference to the two chiral carbons and two enantiomeric molecules that have the same absolute configuration regarding two introduced piperidine rings in each molecule. The obtained diastereomeric mixture exhibited very simple symmetrical ¹³C-NMR in pyridine-d₅, showing little difference magnetically with respect to all of the signals assignable to trisubstituted TAZ. Since these chiral compounds (2e and 3es) as a mixture of stereoisomers showed no significant anti-HSV-1 activity or cytotoxicity at a concentration of less than 100 µM, we did not carry out further detailed experiments for isolation or determination of the absolute configurations of these stereoisomers. We have already reported some examples of the formation of theoretical stereoisomers from a reaction using a racemic reagent as the starting material for the construction of similar symmetrical molecules (see ref. 5).