4. Results
4.3. Synthesis of aminomethyl derivatives
In the first step, the nitrile group was reduced with sodium borohydride in the presence of a catalytic amount of NiCl2, followed by protection with di-tert-butyl dicarbonate (Boc2O) in methanol (39a-f), furnishing the Boc protetcted product. In the second step, the carbamate product was treated with HCl/EtOAc at room temperature for 17-48 hours to afford the final product 40a-f (Figure 50).
N
Figure 50: Preparation of aminomethyl derivatives
The SSAO activity of compounds 40a cis, 40b cis-trans, 40c cis-trans, 40d cis, 40e cis-trans and 40f cis-cis was tested on the microsomal fraction of rat aorta (Table 7, 8). 4-Phenylbutylamine (4-PBA) was used as the reference substrate [77, 106], while 2-bromoethylamine (2-BEA) was used as selective, irreversible inhibitor [107]. According to the hydrogen peroxide formation assay, compounds 40a cis and 40e cis-trans act as inhibitors of moderate potency as compared to 2-BEA, while compound 40b cis-tans
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behaves as a substrate (dose response curves see Appendix). Deme et al., Arkivoc in press.
Table 7. SSAO inhibition of compounds 40a, c, 40d-f and 2-BEA Compound IC50 (µM)
2-BEA* 0.56 ± 0.12 40a cis 5.0 ± 0.4 40c cis-trans 14.7 ± 1.0
40d cis 17.8 ± 5.2 40e cis-trans 5.1 ± 0.5
40f cis-cis 31.0 ± 6.0
*2-BEA = 2-bromoethylamine
Table 8. SSAO activity of compounds 40b and 4-PBA Compound Km (µM)
4-PBA* 740 ± 55
40b cis-trans 0.94 ± 0.01
*4-PBA = 4-phenylbutylamine
4.4. Synthesis of spirocyclic ring systems
First, we tried to perform the Knoevenagel condensation reaction by the previously applied conditions, which are listed in Table 9 and 10 (Figure 51). The microwave assisted one-pot reaction of compound 35a with ID in water resulted 27%
ring closed product (42a)[20]. In order to improve the yield of the reaction, acid (Ti(O-i-Pr)4) [103] as well as acide and base together (AcOH/NaOEt) [108] catalysts were used. According to the TLC the reaction mixture contained a lot of starting material as well as in the traditional condition (cat. piperidine, EtOH). On the basis of the former results this prompted us to employ another condition, thus the microwave reaction in n-BuOH in the presence of acetic acid after quite short reaction time proved the best (yield 49% (42a), 72% (42b) and 35% (42c) respectively)
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Figure 51: Synthesis of spirocyclic ring systems with indan-1,3-dione and Meldrum’s acid from 2-(dialkylamino)acetophenone derivatives
Table 9: Reaction conditions to the synthesis of spirocyclic derivatives with indan-1,3-dione
*ratio of the diastereomers in the crude product were determined by 1H NMR and/or HPLC. SM: starting material; n.d.: not determined
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The reaction with Meldrum’s acid resulted in lower yields (31% (44a), 15%
(44b), and 18% (44c) respectively) putatively the sensitivity of Meldrum’s acid to the temperature and the sterically more hindered six membered ring in the product. In the case of the compound 44b the traditional condition (cat. piperidine, EtOH, r.t., yield 59%) was proved be the best than the microwave condition in EtOH at 50 °C. Another alternative reaction, in the presence of TiCl4/THF, resulted the ring closed product (44a, 44c), as well[109]. Regarding the stereochemical outcome of these reactions - with ID as well as with Meldrum’s acid, - the cis isomer was formed predominantly. In the case of compound 44a and 44b it was not possible to separate the two diastereomers, therefore the trans isomers were not characterized. In the course of the reaction of 2-(dimethylamino)acetophenone (35c) with ID or Meldrum’s acid after the work-up, the vinyl compounds were isolated in low yield and characterized, as well. In other cases the ring closed product was formed directly.
Table 10: Reaction conditions to the synthesis of spirocyclic derivatives with Meldrum’s acid
R1+R2 Reaction condition Crude
product* Yield (%)
(CH2)3
piperidine, EtOH, rt, 72 h 87:13 24
NaOEt, EtOH, rfx, 43 h n.d. 24
NaOH, MeOH, rfx, 24 h n.d. no product TiCl4/THF, THF, pyridine, rt, 24 h 80:20 26
MW, Meldrum (4 eq), cat. AcOH,
EtOH, 50 °C, 10 h 83:17 31
(CH2)4
piperidine, EtOH, rt, 70 h 64:36 59 MW, Meldrum (4 eq), cat. AcOH,
EtOH, 50 °C, 2.5 h 62:37 15
R1=CH3 R2=H
cat. AcOH, NaOEt, EtOH, rfx, 24 h - lot of SM TiCl4/THF, THF, pyridine, rt, 24 - 5 MW, Meldrum (5.2 eq), cat. AcOH,
EtOH, 45 °C, 9 h - 18
*ratio of the diastereomers in the crude product were determined by 1H NMR and/or HPLC. SM: starting material; n.d.: not determined
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In order to study the rate of the cyclization reaction in the presence of the two different electron withdrawing groups (ID vs. malononitrile), the following competition experiments were executed. The starting 2-(dimethylamino)acetophenone (35a) was reacted with one equivalent ID and one equivalent malononitrile, respectively in n-BuOH in the presence of a catalytic amount of acetic acid under microwave irradiation (Figure 52).
CH3 O
N CH3
CH3 CH3
N CH3
CH3 O O
CH3 N CH3
CH3 CN NC
N CH3 CH3
CN CN
N CH3 CH3
O O
+ + +
ID (1 eq) malononitrile (1 eq)
2 drop AcOH n-BuOH
MW
35a 41a 36a 37a 42a
Figure 52: Competition experiment with malononitrile and ID
First, 170-180 °C was used for 15 minutes, then the temperature was reduced in order to see which product formed the fastest way (Table 11). The reactions were followed by NMR spectroscopic. The amount of the compound 42a (30%) was two times more than the compound 37a (13%) at high temperature, while 32% compound 36a was presented in the reaction mixture. At lower temperature (100 °C) the amount of the product 36a was almost four times more than the product 41a and the ring closed product with ID (42a) was also observed in a few per cent. The ratios of the compound 41a and 36a were less at 50 °C than 100 °C. We performed the reaction without acetic acid as well and after 15 minutes one of the ring closed product appeared (42a) next to the formation of the two vinyl compounds (41a and 36a).
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Table 11: Results of the competition experiments in n-BuOH
Reaction condition Products (%)*
comp. 41a comp. 36a comp. 37a comp. 42a Temp (°C), Time (min)
170-180, 15 1 32 13 30
100, 15 10 38 0 1
50, 15 16 29 0 0
100, 15** 18 42 0 5
100, 30** 11 42 traces 8
*ratio of the crude product was evaluated by 1H NMR
**the reaction was carried out without AcOH
The competition experiment was executed in deuteromethanol (CD3OD) and deuterochloroform (CDCl3) at room temperature without acetic acid catalyst, as well (Table 12). This temperature was not enough for the formation of the ring closed product, but we saw clearly that the formation of compound 36a was much more effective than the formation of compound 41a after a long time.
Table 12: Results of the competition experiments in CD3OD
Reaction condition Products (%)*
comp. 41a comp. 36a comp. 37a comp. 42a 25 °C, Time (h)
1 0 traces 0 0
8 0 17 0 0
5** traces traces 0 0
1 month** traces 80 0 0
*ratio of the crude product was evaluated by 1H NMR
**the reaction was carried out in CDCl3
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