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[email protected] Letters in Drug Design & Discovery, 2017, 14, 000-000
1
RESEARCH ARTICLE
Synthesis of Mannich Bases by Two Different Methods and Evaluation of their Acetylcholine Esterase and Carbonic Anhydrase Inhibitory Activities Halise I. Gul1,*, Alkan Demirtas1, Gokbay Ucar1, Parham Taslimi2 and Ilhami Gulcin2,3 1
Ataturk University, Faculty of Pharmacy, Department of Pharmaceutical Chemistry, Erzurum, Turkey; 2Ataturk University, Faculty of Science, Department of Chemistry, Erzurum, Turkey; 3King Saud University, College of Science, Department of Zoology, Riyadh, Saudi Arabia Background: Mannich bases are an important compounds in medicinal chemistry. They have wide range of biological activities including carbonic anhydrase (CA) inhibitory and acetylcholine esterase inhibitory (AChE) activities.
ARTICLE HISTORY
Objective: It was aimed to synthesize Mannich bases, 1-aryl-3-(morpholin-4-yl/piperidin-1-yl)-1propanone hydrochloride, by microwave irradiation and conventional heating methods to compare the methods in terms of reaction times and yields and to investigate their inhibitory effects on AChE enzyme and CA isoenzymes.
Received: July 02, 2016 Revised: October 10, 2016 Accepted: November 18, 2016
Method: Mannich bases were synthesized using conventional heating and microwave irradiation methods under different reaction conditions. Inhibitory effects of the compounds on CA isoenzymes and AChE were evaluated according to literature procedure.
DOI: 10.2174/15701808146661611281206 12
Results: IC50 and Ki values of the compounds were evaluated against hCA I, II and AChE. The compounds had more potent or equal Ki values with the references used. Conclusion: This study makes an important contribution to the Mannich base library in terms of synthetic strategy. According to IC50 or Ki values the compounds 6 in Series A with morpholine and and 15 in Series B with piperidine towards both hCA I and/or II isoenzymes and the compounds 4 in Series A and 11, 13, 14, 15, 16, and 18 in Series B towards AChE seemed the leader compounds of the study.
Keywords: Acetylcholine esterase, carbonic anhydrase, conventional heating, Mannich bases, microwave irradiation. INTRODUCTION Mannich bases are an important group of compounds in medicinal chemistry and they are synthesized by Mannich reaction [1]. Mannich bases have a wide range of biological activities such as carbonic anhydrase inhibitory [2-4], cytotoxic [5-8], anti-inflammatory [9] anticonvulsant [10] and acetylcholine esterase inhibitory [11] activities. The reported mechanisms of Mannich bases for their bioactivities are thiol alkylation [12-15], interaction with enzymes which are important for antioxidant mechanisms [6], inhibition of mitochondrial respiration [16, 17], inhibition of topoisomerase enzyme [18, 19] and the inhibition of tubulin polimerization [20]. Cholinesterases (ChE) are an enzyme family that catalyzes the hydrolysis of acetylcholine (ACh) into choline and acetic acid, an essential process for the restoration of cholinergic neurotransmission. There are two cholinesterase
*Address correspondence to this author at the Department of Pharmaceutical Chemistry, Faculty of Pharmacy, Ataturk University, Erzurum 25240, Turkey; Tel: +90 442 231 5203; Fax: +90 442 231 5201; E-mail:
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types: Acetylcholinesterase (AChE, EC 3.1.1.7) and butyrylcholinesterase (BChE, EC 3.1.1.8). AChE is known to be abundant in the muscle, brain, and erythrocyte membrane, whereas BChE has a higher activity in the liver, intestine, heart, kidney, and lung. They have similar molecular forms and active sites despite being products of different genes on the human chromosomes. Inhibition of hydrolyses of acetylcholine (ACh) and butrylcholine (BCh) by using cholinesterase inhibitors have been considered to increase the level of the ACh and BCh in synapses, which is a useful approach for the treatment of Alzheimer disease (AD). AD is a progressive, chronic, neurodegenerative disorder, characterized by decline in the memory and cognitive abilities. It is estimated that about 6% of the population worldwide aged over 65 is affected. The etiology of AD is still not full known, but the reduced level of acetylcholine and butrylcholine plays a significant role [21]. Although there are many on-going researches for the treatment of AD, only some drugs were approved by FDA such as Tacrine, Donepezil and Rivastigmin. So, there is a need for some new compounds which can be good candidates for the development of new drugs having superior properties. ©2017 Bentham Science Publishers
2 Letters in Drug Design & Discovery, 2017, Vol. 14, No. 5
Carbonic anhydrases (CAs, EC 4.2.1.1) catalyze a very simple but physiologically and biochemically essential reaction in all life kingdoms, followed by the hydration of carbon dioxide (CO2) to bicarbonate (HCO3-) and protons (H+), with a high efficiency. They are present either in eukaryotes or prokaryotes and are encoded by six distinct non-related gene families: alpha (α), beta (β), gamma (γ), delta (δ), zeta (ζ) and eta (η). All human CAs (hCAs) belong to the alpha (α)-class. Until now, sixteen isozymes have been recognized, among these; only twelve are catalytically active (CAs I–IV, CAs VA–VB, CAs VI–VII, CA IX and CAs XII–XIV), whereas, the CA-related proteins (CARPs) VIII, X and XI were formed without any catalytic activity. CA I, and II are cytosolic ones [22, 23]. In the last years, CA isozymes have become an interesting target for the design of inhibitors or activators with biomedical applications and different human diseases such as glaucoma, osteoporosis, neurological disorders, cancer, etc. Originally, CA inhibitors (CAIs) were clinically used mainly as anti-glaucoma diuretics and anti-epileptics, while the novel generation compounds are currently under clinical investigation as antiobesity or anti-tumour drugs and diagnostic tools [22, 24, 25]. However, none of the CAIs in clinical use showed selectivity for a specific isoenzyme. Thus, the development of isoenzyme-specific CAIs inhibitors would be extremely helpful to discover novel classes of drugs lacking of a range of undesired side effects. Synthesis of several chemical structures by microwave irradiation is quite common experimental method especially during the last decades. It has been reported that microwave irradiation method improves the yield of reaction, shortens the reaction time, and provides saving from energy [26] compared to the synthesis by conventional heating method [27-30]. For instance, 3-(3-bromophenyl)-1-{4,6-dimethoxy2-hydroxy-3-[(4-methylpiperazin-1-yl)methyl]phenyl}prop2-en-1-one was synthesized with the yield of 96.5 % in 10 minutes by microwave irradiation while it was synthesized with the yield of 61 % in 1.5 hours by conventional heating method [31]. Although chemical synthesis by microwave irradiation is quite common, there is a limited number of studies on the synthesis of Mannich bases by microwave irradiation [32]. The aims of this study were to determine the most suitable experimental method for the synthesis of some mono Mannich bases having the chemical structure of 1-aryl-3(morpholin-4-yl/piperidin-1-yl)-1-propanone hydrochloride in terms of reaction time and reaction yield by the comparison of the conventional heating method with the microwave irradiation method. Thus, to find out the most suitable method which can be applied for the synthesis of
Gul et al.
new Mannich bases with different chemical structures in further and to make a contribution to the library of Mannich bases with a limited number of studies in which microwave irradiation method applied wasavailable. The Second aim was to investigate the effects of the compounds synthesized on carbonic anhydrase and acetylcholine esterase enzymes, since these enzymes play an important role in the physiology and pathology of several diseases and some types of Mannich bases were reported with CA [2-4] and AChE [11, [21, 40] inhibitory activities. The data obtained may help to find out new drug candidate compound/s for the enzyme activities in question for further studies. MATERIALS AND METHODS Chemistry All the reactions were carried out in CEM Discover Microwave Synthesis Systems, Model 908010 (CEM Matthews, NC, USA). Melting points were determined using an Electrothermal 9100 (IA9100, Bibby Scientific Limited, UK) instrument and were uncorrected. The reactions were monitored by thin layer chromatography (TLC). Synthesis of 1-Aryl-3-(Morpholin-4-yl/Piperidin-1-yl)-1Propanone Hydrochlorides (1-18) by Conventional Heating Method in Glacial Acetic Acid A ketone compound, paraformaldehyde and suitable amine were taken in 1:1.2:1 mol ratios and heated in 10 ml of glacial acetic acid at 120 oC for several times. The reactions were monitored by TLC using CHCl3: MeOH (9:1 or 8:2) as a solvent system. Compounds were purified by crystallisation. Chemical structures of the compounds were confirmed by 1H NMR (See Supplementary File) and their reported melting points. Amine part was changed as morpholine in Fig. ((1) Series A (1-9)) and piperidine in Series B (10-18). Physical data of the compounds obtained by the conventional heating method are reported in Table 1 (See Supplementary File). Synthesis of 1-Aryl-3-(Piperidin-1-yl)-1-Propanone Hydrochlorides (10-18) by Conventional Heating Method in Ethanol at pH=5 A ketone compound, paraformaldehyde and suitable amine were taken in 1:1.2:1 mol ratios and heated in 10 mL of ethanol acidified with HCl (37%) at 100 oC for several times. The reactions were monitored by TLC using CHCl3: MeOH (9:1 or 8:2) as a solvent system. The compounds were purified by crystallisation. Chemical structures of the compounds were confirmed by 1H NMR (See Supplementary File). Amine part was piperidine (10-13, 15, 16, 17).
O Ar
O N
.HCl
Ar
N
.HCl
O A Series (1-9)
B Series (10-18)
Ar: Phenyl (1,10), 4-methylphenyl (2,11), 4-methoxyphenyl (3,12), 4-chlorophenyl (4,13), 4-hydroxyphenyl (5,14), 4-bromophenyl (6,15), 4-fluorophenyl (7,16), 2-thienyl (8,17), 2-furyl (9,18)
Fig. (1). Chemical structures of the compounds 1-18.
CA and AChE Inhibitory Effects of Mannich Bases
Table 1.
Letters in Drug Design & Discovery, 2017, Vol. 14, No. 5
3
Physical data of 1-aryl-3-(morpholin-4-yl/piperidin-1-yl)-1-propanone hydrochlorides by conventional heating method in acetic acide. Ketone
Paraformaldehyde
(mmol)
(mmol)
Amine.HCl (mmol)
Reaction Time (min)
Yield (%)
Crystallisation Solvent
C6H5
12.5
15
12.5
125
39
MeOH
2
4-CH3C6H 4
11.2
13.4
11.2
125
53
CHCl3
3
4-CH3OC6H 4
10
12
10
75
41
MeOH/CHCl3
4
4-ClC6H4
9.7
11.6
9.7
125
21
MeOH/CHCl3
5
4-HOC6H4
11
13.2
11
65
30
MeOH/CHCl3
6
4-BrC6H4
7.5
9
7.5
95
55
MeOH/CHCl3
7
4-FC6H4
10.9
13.1
10.9
175
67
MeOH/CHCl3
8
C4H3 S (2-yl)
11.9
14.3
11.9
145
38
MeOH/CHCl3
9
C4H3O (2-yl)
13.6
16.3
13.6
175
23
MeOH/CHCl3
10
C6H5
12.5
15
12.5
205
46
MeOH/Et2O
11
4-CH3C6H 4
11.2
13.4
11.2
505
34
MeOH/Et2O
12
4-CH3OC6H 4
10
12
10
475
59
MeOH/CHCl3
13
4-ClC6H4
9.7
11.6
9.7
560
23
MeOH
14
4-HOC6H4
11
13.2
11
180
45
MeOH
15
4-BrC6H4
7.5
9
7.5
255
6
MeOH/Et2O
16
4-FC6H4
10.9
13.1
10.9
440
26
MeOH/Et2O
17
C4H3 S (2-yl)
11.9
14.3
11.9
585
34
MeOH/Et2O
18
C4H3O (2-yl)
13.6
16.3
13.6
180
23
MeOH/Et2O
Compounds
Aryl
1
Physical data of the compounds obtained by the conventional heating method in the ethanol at pH=5 are presented in Table 3 (See Supplementary File). Synthesis of 1-Aryl-3-(Morpholin-4-yl/Piperidin-1-yl)-1Propanone Hydrochlorides (1-18) by Microwave Irradiation Method A ketone compound, paraformaldehyde and suitable amine were taken in 1:1.2:1 mol ratios and heated in 10 mL of glacial acetic acid at 120 oC at 70 Watt for series A and B for several times. The reactions were monitored by TLC using CHCl3: MeOH (9:1 or 8:2) as a solvent system. The compounds were purified by crystallisation. Chemical structures of the compounds were confirmed by 1H NMR (data were not present) and their reported melting points. Amine part was morpholine in Series A (1-9) and piperidine in Series B (10-18). Physical data of the compounds obtained by microwave irradiation method are reported in Table 2 (See Supplementary File). BIOLOGICAL ACTIVITY Carbonic anhydrase inhibition and cholinesterase inhibition assays were realised according to the procedures reported in the literatures [2, 4, 23, 33, 34], respectively.
RESULTS AND DISCUSSION Microwave irradiation method decreased the reaction time in both the series, except compound 5. The decrease in the reaction time by microwave irradiation was 1.5-25 times in series A with morpholine, and 3.2-14.6 times in series B with piperidine. The increase in the reaction yield by microwave irradiation was 1.3 times for compound 3, 1.9 times for compound 4, 1.1 times for compounds 9 and 18, 1.4 times for compound 13, and 3.5 times for compound 15. However, the conventional heating method showed 1.1-1.8 times higher yield for compounds 2 (1.2 times), 7 (1.5 times), 8 and 17 (1.1 times), 6 and 11 (1.7 times), 1 (1.8 times), 5 (1.3 times), 10 (2.1 times), 12 (11.8 times), and 16 (1.6 times) than the microwave irradiation method (Table 4). Synthesis of the compounds 10-13, 15-17 were also realised at pH 5 in the ethanol acidified with HCl (37%) instead of glacial acetic acid (Table 3). The reaction time and reaction yields for these compounds were also compared. When the reactions were realised in the acetic acid (Table 1), the reaction time decreased 1.3-3.9 times compared to the reactions with the acidic ethanol medium used for the compounds 10 (3.9 times), 11 (2.1 times), 12, 13, and 14 (1.3 times), and 15 (1.4 times). In other words, the reaction medium with acidic ethanol increased the reaction time 1.33.9 times compared to the one in which the reaction medium
4 Letters in Drug Design & Discovery, 2017, Vol. 14, No. 5
Table 2.
Gul et al.
Physical data of 1-aryl-3-amino (morpholin-4-yl/piperidin-1-yl)-1-propanone hydrochlorides by microwave irradiation method.
Compounds
Aryl
Ketone (mmol)
Para Formaldehyde (mmol)
Amine.HCl (mmol)
Reaction Time (min)
Yield (%)
Crystallisation Solvent
Melting Point (oC)
1
C6H5
12.5
15
12.5
10
22
MeOH
177-178
2
4-CH3C6H 4
11.2
13.4
11.2
5
45
MeOH
206-208
3
4-CH3OC6H 4
10
12
10
10
53
MeOH/CHCl3
211-213
4
4-ClC6H4
9.7
11.6
9.7
55
40
MeOH/CHCl3
211-214
5
4-HOC6H4
11
13.2
11
240
24
MeOH
217-219
6
4-BrC6H4
7.5
9
7.5
60
33
MeOH
215-218
7
4-FC6H4
10.9
13.1
10.9
120
46
MeOH
204-207
8
C4H3 S (2-yl)
11.9
14.3
11.9
80
36
MeOH
220-224
9
C4H3O (2-yl)
13.6
16.3
13.6
80
26
MeOH/CHCl3
207-208
10
C6H5
12.5
15
12.5
65
22
MeOH/Et2O
191-192
11
4-CH3C6H 4
11.2
13.4
11.2
45
20
MeOH/Et2O
177
12
4-CH3OC6H 4
10
12
10
60
5
MeOH/CHCl3
211-212
13
4-ClC6H4
9.7
11.6
9.7
65
32
MeOH
189
14
4-HOC6H4
11
13.2
11
20
45
MeOH
221
15
4-BrC6H4
7.5
9
7.5
60
21
MeOH/Et2O
201-202
16
4-FC6H4
10.9
13.1
10.9
60
16
MeOH/Et2O
189-190
17
C4H3 S (2-yl)
11.9
14.3
11.9
40
31
MeOH/Et2O
200-202
18
C4H3O (2-yl)
13.6
16.3
13.6
40
25
MeOH/Et2O
185-186
o
Reported Compounds (mp, C) [Reference] = 1 (178-179) [35], 2 (224) [36], 3(212-214) [37], 4 (208-209) [38], 5 (215-217) [39], 6 (208-09) [40], 7 (187) [41], 8 (194) [42], 9 (191192) [43], 10 (178-184) [44], 11(174-177) [45], 12 (nr), 13 (188-190) [45], 14 (187-188) [46], 15 (206-207) [36], 16 (166-169) [40], 17 (201-202) [47], 18 (185-186) [48].
Table 3.
Physical data for the compounds 10-18 for which conventional heating method in ethanol at pH=5 was applied.
Compounds
Aryl
Para
Ketone (mmol)
Formaldehyde (mmol)
Amine.HCl (mmol)
Reaction Time (min)
Yield (%)
Crystallisation Solvent
10
C6H5
12.5
15
12.5
805
17
MeOH
11
4-CH3C6H 4
11.2
13.4
11.2
1075
24
MeOH
12
4-CH3OC6H 4
10
12
10
620
29
MeOH
13
4-ClC6H4
9.7
11.6
9.7
750
22
MeOH
14
4-HOC6H4
15
4-BrC6H4
7.5
9
7.5
360
32
MeOH
16
4-FC6H4
10.9
13.1
10.9
590
17
MeOH
17
C4H3 S (2-yl)
11.9
14.3
11.9
455
23
MeOH
18
C4H3O (2-yl)
Not studied
was acetic acid (Table 1 and 3). That is why the acetic acid was found to be a better solvent system for the synthesis and used as the reaction solvent for the synthesis of the designed
Not studied
chemical structures in this study. Comparative results of the reaction yield and times for the studied compounds are presented in Table 4.
CA and AChE Inhibitory Effects of Mannich Bases
Table 4.
Letters in Drug Design & Discovery, 2017, Vol. 14, No. 5
5
Comparative table for the compounds 1-18 for which by conventional heating and microwave irradiation methods were applied. Reaction Time (min)
Compounds
Yield (%)
Conventional
Microwave
Conventional
Microwave
1
125
10
39
22
2
125
5
53
45
3
75
10
41
53
4
125
55
21
40
5
65
240
30
24
6
95
60
55
33
7
175
120
67
46
8
145
80
38
36
9
175
80
23
26
10
205
65
46
22
11
505
45
34
20
12
475
60
59
5
13
560
65
23
32
14
180
20
45
45
15
255
60
6
21
16
440
60
26
16
17
585
40
34
31
18
180
40
23
25
Inhibitory effects of the compounds on carbonic anhydrase isoenzymes hCA I and hCA II are presented in Table 5. IC50 values of the compounds were in the range of 57.22 - 97.27 nM in Series A with mopholine, while they were in the range of 57.75 - 85.41 nM in Series B with piperidine for hCA I isoenzyme. On the other hand, IC50 values of the compounds were in the range of 61.88 - 99.02 nM in Series A and 69.31-86.63 nM in Series B a for hCA II isoenzyme, as shown in Table 5. IC50 values of the reference compound Acetazolamide (AZA) for hCA I and II were 197.30 nM and 115.50 nM, respectively. It means that all the compounds were more effective than AZA for both hCA I and II isoenzymes, since the IC50 values of the compounds were lower than AZA’s. When the effect of substituent on the phenyl ring in Series A with morpholine, was considered by comparing the IC50 values of compounds 2-7 with compound 1, it was noticed that any substituent on the phenyl ring rather than hydrogen was helpful to increase the activity against hCA I by lowering the IC50 value of the compound compared to the IC50 value of hydrogen substituted compound 1. The presence of oxygen atom in compound 3 with methoxy substituent decreased the activity compared to 2 with methyl substituent on the phenyl ring but when the hydroxy group,
which has less strerical hinderence than methoxy substituent, was introduced to the chemical structure in compound 5, it showed similar activity to compound 2 for hCA I. When series B with piperidine was considered, the effect of the substituent on the phenyl ring was not constant for hCAI. Compound 15 with bromine and compound 16 with fluorine had lower IC50 values than 10, which is a nonsubstituted compound and this suggested the increased activity for hCA I. The replacement of the phenyl ring (10) to thiophene (17) or furan (18) decreased the activity by increasing the IC50 value toward hCA I. The most effective compounds for hCA I in terms of the IC50 value were bromine substituted compounds 6 and 15 from series A and B, respectively. Among the compounds having halogen at the para position of the phenyl ring , the order of effectiveness in terms of inhibiting hCA I isoenzyme was as follows: compound 15 (with bromine) > 16 (with fluorine) > 13 (with chlorine) in Series B. It means that the activity was not depending on the electronegativity of the halogen in Series B. Halogen (4, 6, 7) or methoxy (3) substitution increased the activity in series A with morpholine compared to 1 for hCA II. The replacement of the phenyl ring (1) by furan (9) decreased the activity while the replacement of phenyl ring
6 Letters in Drug Design & Discovery, 2017, Vol. 14, No. 5
Table 5.
Gul et al.
Carbonic anhydrase and acetylcholine esterase inhibitory activities of the compounds 1-18. IC50 (nM)
Compounds
KI (nM)
hCA I
r2
hCA II
r2
AChE
r2
hCA I
hCA II
AChE
1
97.27
0.9670
84.94
0.9524
23.89
0.9608
52.58±8.55
64.63±25.86
11.52±5.18
2
74.26
0.9421
89.11
0.9344
17.33
0.9888
60.95±13.51
105.32±35.21
7.36±3.44
3
85.46
0.9655
71.69
0.9655
11.75
0.9698
68.50±21.15
63.05±21.35
5.02±1.28
4
61.63
0.9736
74.97
0.9409
10.19
0.9831
53.07±17.12
85.51±27.67
2.92±0.94
5
75.71
0.9624
97.47
0.9527
18.24
0.9687
46.39±5.89
62.53±16.10
7.18±1.64
6
57.22
0.9884
61.88
0.9391
12.61
0.9913
42.24±7.03
55.85±14.58
6.56±1.92
7
85.24
0.9610
73.24
0.9495
18.19
0.9587
57.61±8.068
74.72±25.97
12.29±4.16
8
71.95
0.9755
84.01
0.9202
16.12
0.9687
67.20±14.53
83.38±36.22
5.35±1.33
9
83.86
0.9658
99.02
0.9485
30.13
0.9623
84.02±25.70
90.97±21.58
13.45±6.44
10
73.10
0.9523
79.66
0.9297
15.75
0.9776
71.53±31.50
95.03±46.27
9.67±3.93
11
85.41
0.9436
75.57
0.9085
13.86
0.9879
80.69±30.95
60.13±13.95
4.11±1.77
12
70.08
0.9727
73.72
0.9275
22.35
0.9601
67.06±20.57
77.96±27.36
10.09±3.84
13
74.31
0.9522
81.53
0.9401
11.18
0.9774
53.72±11.98
80.95±37.27
4.53±1.63
14
75.76
0.9810
77.01
0.9323
14.75
0.9807
76.32±20.72
66.06±20.50
6.65±2.11
15
57.75
0.9905
69.31
0.9181
17.76
0.9858
54.98±13.70
78.06±26.45
6.31±2.09
16
69.22
0.9928
85.56
0.9551
16.49
0.9774
51.03±9.45
85.10±25.73
5.01±2.22
17
78.75
0.9604
83.49
0.9469
15.01
0.9850
98.73±28.22
81.18±19.66
8.09±2.02
18
76.07
0.9658
86.63
0.9615
21.66
0.9797
56.67±13.21
78.62±15.62
6.88±3.04
AZA*
197.30
0.9889
115.50
0.9719
-
-
183.39±19.71
104.60±27.60
-
TAC**
-
-
-
-
43.31
0.9948
-
-
23.29±4.07
*Acetazolamide (AZA) was used as a standard inhibitor for all hCA I-II **Tacrine (TAC) was used as a standard inhibitor for AChE enzyme.
by thiophene did not affect the activity for hCA II. The replacement of hydrogen in 1 by bromine (15), hydroxy (14), methoxy (12), and methyl (11) increased the activity for hCA II in Series B. Among the aromatic rings, phenyl was the best in terms of the IC50 values in both the series. Bromine substituted compounds 6 and 15 were the most effective ones for hCA II as well as in the case of hCA I. When the Ki values were considered, they were in the range of 42.24±7.03 and 84.02±25.70 nM in series A and 51.03±9.45 and 98.73±28.22 nM in series B, while the reference compound AZA had the Ki value of 183.39±19.71 nM for hCA I. This suggests that all the compounds (1-18) with lower Ki values than AZA’s were better inhibitor compared to the reference drug AZA, as shown in Table 5. The most potent compounds for hCA I were 6 (with bromime) in series A, and the compounds which have the similar Ki values in series B, 13 ( with chlorine), 15 (with bromine), and 16 (with fluorine) in terms of the Ki value . On the other hand, the Ki values of the compounds were in the range of 55.85±14.58 and 105.32 ±35.21 nM in series A and 60.13±13.95 and 95.03±46.27 nM in series B for hCA II. All the compounds 1-18, except 2 having the similar Ki
value with AZA, had more potent activity than AZA for hCA II. The most potent compounds for hCA II in terms of the Ki value was bromine substituted compound 6 in Series A and methyl substituted compound 11 in Series B. When the acetylcholine esterase inhibition effects of the compounds were considered, the IC50 values of compounds 1-9 in Series A with morpholine were in the range of 10.19 and 30.13 nM while they were in the range of 11.18 and 22.35 nM for compounds 10-18 in Series B with piperidine, as shown in Table 5. The most effective compounds for acetylcholine esterase inhibition were 4 and 13 having chlorine on the pheny ring in both the series A and B, respectively in terms of the IC50 values. Substitution on the phenyl ring other than hydrogen was a useful modification to increase AChE inhibitory activity in series A for compounds 2-7.The replacement of the phenyl ring 1 by thiophene 8 increased the AChE inhibition activity but the replacement of the phenyl ring 1 to furan 9 decreased the activity in Series A. Different substituent rather than hydrogen on the phenyl ring increased the activity slightly for compounds 11 (with
CA and AChE Inhibitory Effects of Mannich Bases
methyl), 13 (with chlorine), and 14 (with hydroxy) in Series B. The replacement of the phenyl ring 1 by thiophene 8 increased the AChE inhibitory activity (2.2 times) while the replacement of the phenyl ring 1 to furan 9 decreased slightly the activity. All the compounds studied 1-18 were 1.4 - 4.3 times more effective than the reference drug Tacrine (TAC) for the inhibition of AChE in terms of the IC50 Values. When the Ki values of the compounds were considered for AChE, they were in the range of 2.92±0.94 and 13.45±6.44 nM in series A while in the range of 4.11±1.77 and 10.09±3.84 nM in series B. The Ki value of the reference drug TAC was 23.29±4.07 nM. In terms of the AChE inhibition effect, the most effective compounds were compound 4 in series A and compounds 11, 13, 14, 15, 16, and 18 in series B in terms of the Ki values. As conclusion, this study makes an important contribution to the Mannich base library in terms of the synthetic strategy. Acetic acid was found to be a better solvent than ethanol. Microwave irradiation method made a clear improvement in the reaction times. In terms of the reaction yield, there was no best method since the yields differed depending on the compounds synthesized. It follows from the study that compounds 6 and 15 in Series A and B for both hCA I and II isoenzymes in terms of the IC50 or Ki values, and compounds 4 in Series A along with 11, 13, 14, 15, 16, and 18 in Series B in terms of the Ki values for AChE were examined. The leading compounds for in terms of both the IC50 and the Ki values, seems to be chlorine substituted compound 4 in series A and the methyl substituted compound 11 in series B.
Letters in Drug Design & Discovery, 2017, Vol. 14, No. 5 [6] [7]
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CONFLICT OF INTEREST The authors confirm that this article content has no conflict of interest. ACKNOWLEDGEMENTS This research work was supported by Ataturk University Research Found, Erzurum, Turkey (Project No: 2010/166, 2012/74, 2012/75).
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