2013 Volume 61 Issue 5 Pages 572-575
We have developed an efficient synthesis of dichlorodenafil (4), an unapproved sildenafil analogue isolated from dietary supplements. Our sequence employs POCl3-mediated chlorination of readily available chloroacetyl compound 7 followed by selective hydrolysis of the chloro-heterocycle function. Our synthesis confirms the structure of the illegal additive, and will provide regulatory agencies with ready access to authentic standard samples of dichlorodenafil (4) to aid in their mission to protect the public from unapproved and potentially harmful erectile dysfunction (ED) drug analogues that are added to herbal and dietary supplements without providing users with appropriate toxicological or pharmacological information.
Since sildenafil (1) was introduced onto the U.S. market in 1998 for the treatment of erectile dysfunction (ED), other ED drugs such as vardenafil (2) and tadalafil (3) have subsequently been commercialized1,2) (Fig. 1). These synthetic ED medications function by inhibiting the phosphodiesterase type 5 enzyme (PDE-5). For the past several years, unapproved synthetic analogues (mostly based on sildenafil, vardenafil, and tadalafil) have been routinely identified in ‘all-natural’ herbal remedies and dietary supplements.3) These compounds generally exhibit only minor structural differences compared to their approved or commercial counterparts. Based on their structural similarities, it is reasonable to expect these analogues to exhibit similar biological activities. In most cases, however, there is no information available to the public with regard to the toxicological or pharmacological effects of these illegal and unapproved analogues.4) Thus, the presence of analogues in herbal supplements and dietary supplements, especially in an unapproved dosage, could pose a significant risk to public health.
In 2010, the Korea Food and Drug Administration (KFDA) and the Korea Customs Service (KCS) jointly announced the isolation of a new ED drug analogue discovered as an additive in dietary supplements imported by international air mail. Spectroscopic studies established the structure of this analogue as 5-(5-((Z)-1,2-dichlorovinyl)-2-ethoxyphenyl)-1-methyl-3-propyl-1H-pyrazolo[4,3-d]pyrimidin-7(6H)-one (4) as shown in Fig. 2. This new unapproved ED drug analogue possesses a (Z)-dichlorovinyl moiety in place of the sulfonyl-N-methylpiperazine group in sildenafil (1), and was given the name of dichlorodenafil (4). Subsequently, the KFDA amended its Standards and Specification of Food to enable the regulation of dichlorodenafil (4).5)
In order to corroborate the structure (4) that was proposed for dichlorodenafil based on the spectroscopic analysis, and to provide a standard sample to aid in its detection in dietary supplements, we embarked on a synthesis of this substance. To our knowledge, no synthetic approach to dichlorodenafil has been described in the literature to date. We describe in this note an efficient synthesis and confirm that dichlorodenafil possesses structure 4.
As outlined in our synthetic plan (Chart 1), we envisioned that dichlorodenafil (4) could be obtained by selective hydrolysis of chloro-heterocyclic function in 5. Kagan and co-workers reported that upon exposure to 1.1 to 1.3 eq of PCl5, acetophenone produced a complex mixture which included the corresponding dichlorovinyl compound (16%, unspecified E/Z ratio) and chloroacetylbenzene (4%).6) We further envisaged that chloroacetyl 7-chloropyrazolopyrimidinone 6, prepared by ring chlorination7,8) of known chloroacetyl pyrazolopyrimidinone 7, might be elaborated to the desired (Z)-dichlorovinyl compound 5 upon exposure to PCl5 through optimization of the chlorination conditions of Kagan et al. The known chloroacetyl compound 7 could be prepared via Friedel–Crafts acylation of the commercially available pyrazolopyrimidinone intermediate 8.9)
Initially, we pursued a stepwise synthesis of dichlorodenafil (4) as summarized in Chart 2. To this end, known chloroacetyl compound 7 was converted to the corresponding 7-chloropyrazolopyrimidinone 6 by treatment with POCl3 in 88% yield.7,8) We were pleased to find that exposure of the resultant chloroacetyl 7-chloropyrazolopyrimidinone 6 to excess PCl5 (4 equiv.) in refluxing toluene for 12 h furnished a mixture of 5 (66% isolated yield) and 5′ in favor of the desired (Z)-isomer 5. Finally, selective hydrolysis of the chloroheterocycle 5 by treatment with conc.-HCl in t-BuOH at room temperature for 12 h delivered the desired dichlorodenafil (4) in 84% isolated yield.10) In the 1H-NMR spectrum, the appearance of the vinyl proton in (Z)-isomer 4 downfield relative to that in (E)-isomer 4′ as indicated in the Chart 2, and the nOe interaction between the vinyl proton and the aromatic protons [between the H-23 and H-15 and H-23 and H-17] in (Z)-isomer 4 firmly established the configuration of the dichlorovinyl group as (Z) for the major isomer. The spectral data of our synthetic material were identical to those of dichlorodenafil isolated from dietary supplements.
Reaction conditions: (a) POCl3, reflux, 4 h, 88%; (b) PCl5 (4 eq.), toluene, reflux, 12 h, 81%; and (c) conc.-HCl, t-BuOH, rt, 12 h, 84% for 4 and 81% for 4′.
Having accomplished an efficient synthesis and structure confirmation of dichlorodenafil (4), we proceeded to develop a streamlined, two-step preparation by using POCl3 for dichlorovinylation as well as the ring chlorination (Chart 3). To our satisfaction, the chloroacetyl group in 7 was efficiently elaborated to the desired (Z)-dichlorovinyl substituent in refluxing POCl3 for 72 h with concomitant chlorination of the pyrazolopyrimidinone nucleus to yield a mixture of 5 and 5′. Premature interruption of the reaction [POCl3, reflux, 3 h] resulted in the isolation of chloropyrazolopyrimidine 6. The reaction mixture was poured into ice and allowed to stand at room temperature overnight to destroy excess POCl3. It is worth mentioning that when the rate of quenching was hastened by stirring instead of standing overnight, the process then gave varying amounts of starting material 7 due to hydrolysis, which we attributed to a temperature rise during the presumably faster quench. Addition of t-BuOH and stirring the resulting solution at room temperature for 12 h effected selective hydrolysis of the chloroheterocycle to furnish the desired dichlorodenafil (4) in 66% isolated yield along with isomeric 4′ (8%).10)
Reaction conditions: (a) POCl3, reflux, 72 h; (b) t-BuOH/H2O, rt, 12 h.
In summary, we have developed an efficient synthesis of dichlorodenafil (4), an unapproved sildenafil analogue isolated from dietary supplements. Our sequence employs POCl3-mediated chlorination of readily available chloroacetyl compound 7 followed by selective hydrolysis of the chloroheterocycle function. Our synthesis confirms the structure of this illegal additive, and will provide regulatory agencies with ready access to authentic standard samples of dichlorodenafil (4) to aid in their mission to protect the public from unapproved and potentially harmful ED drug analogues that are added to herbal and dietary supplements without providing users with appropriate toxicological and pharmacological information.
Experimental Procedure for Two-Step Synthesis of Dichlorodenafil (4)Chloroacetyl compound 6 (1.42 g, 3.65 mmol) was suspended in POCl3 (15 mL) and heated to reflux for 72 h. After cooling to room temperature, the mixture was poured into crushed ice (150 g), and allowed to stand at room temperature overnight. To the resulting mixture was added t-BuOH (20 mL), and this solution was stirred at room temperature for 12 h. The solution was extracted with CH2Cl2 (30 mL×3), and then, the combined organic layers were successively washed with sat. NaHCO3 and brine, dried over MgSO4, and concentrated in vacuo. The residue was purified by silica gel column chromatography (hexane/ethyl acetate=2/1) to give 4 (0.977 g, 2.40 mmol, 66%) and 4′ (123 mg, 0.302 mmol, 8%) as white solids.
(Z)-7-Chloro-5-(5-(1,2-dichlorovinyl)-2-ethoxyphenyl)-1-methyl-3-propyl-1H-pyrazolo[4,3-d]pyrimidine (8)1H-NMR (400 MHz, CDCl3) δ 7.90 (d, J=2.8 Hz, 1H), 7.57 (dd, J=8.8, 2.8 Hz, 1H), 7.01 (d, J=8.8 Hz, 1H), 6.66 (s, 1H), 4.37 (s, 3H), 4.14 (q, J=6.8 Hz, 2H), 3.05 (t, J=7.6 Hz, 2H), 1.91 (sextet, J=7.6 Hz, 2H), 1.37 (t, J=6.8 Hz, 3H), 1.03 (t, J=7.6 Hz, 3H); 13C-NMR (100 MHz, CDCl3) δ 158.4, 156.6, 146.8, 145.7, 143.2, 135.3, 130.3, 129.2, 128.4, 128.1, 127.8, 114.9, 113.5, 64.9, 38.7, 28.0, 22.3, 14.8, 14.2; IR (neat) 1587, 1522, 1498, 1443, 1255, 1224, 1044, 900, 802, 671; mp 107.6°C; electrospray ionization (ESI)-MS m/z 425 (M+H).
(E)-7-Chloro-5-(5-(1,2-dichlorovinyl)-2-ethoxyphenyl)-1-methyl-3-propyl-1H-pyrazolo[4,3-d]pyrimidine (8′)1H-NMR (400 MHz, CDCl3) δ 8.04 (d, J=2.0 Hz, 1H), 7.67 (dd, J=8.8, 2.0 Hz, 1H), 7.03 (d, J=8.8 Hz, 1H), 6.46 (s, 1H), 4.37 (s, 3H), 4.15 (q, J=6.8 Hz, 2H), 3.05 (t, J=7.6 Hz, 2H), 1.91 (sextet, J=7.6 Hz, 2H), 1.38 (t, J=6.8 Hz, 3H), 1.03 (t, J=7.6 Hz, 3H); 13C-NMR (100 MHz, CDCl3) δ 158.2, 156.7, 146.9, 145.8, 143.2, 132.9, 132.3, 131.7, 127.8, 127.7, 126.7, 113.5, 112.9, 64.8, 38.2, 28.1, 22.3, 14.9, 14.2; IR (neat) 1501, 1467, 1397, 1284, 1262, 1176, 1040, 960, 866, 805; mp 93.7°C; ESI-MS m/z 425 (M+H).
(Z)-5-(5-(1,2-Dichlorovinyl)-2-ethoxylphenyl)-1-methyl-3-propyl-1H-pyrazolo[4,3-d]pyrimidin-7(6H)-one (4)1H-NMR (400 MHz, CDCl3) δ 10.9 (br s, 1H), 8.60 (d, J=2.4 Hz, 1H), 7.62 (dd, J=8.8, 2.8 Hz, 1H), 7.05 (d, J=8.8 Hz, 1H), 6.75 (s, 1H), 4.32 (q, J=6.8 Hz, 2H), 4.28 (s, 3H), 2.95 (t, J=7.6 Hz, 2H), 1.88 (sextet, J=7.6 Hz, 2H), 1.62 (t, J=6.8 Hz, 3H), 1.05 (t, J=7.6 Hz, 3H); 13C-NMR (100 MHz, CDCl3) δ 157.2, 153.9, 147.6, 147.0, 138.6, 134.7, 130.6, 129.7, 129.4, 124.6, 120.6, 116.0, 113.2, 65.8, 38.4, 27.9, 22.5, 14.8, 14.2; IR (neat) 1697, 1494, 1391, 1249, 1158, 1126, 1032, 890, 779, 669; mp 138.5°C; ESI-MS m/z 407 (M+H).
(E)-5-(5-(1,2-Dichlorovinyl)-2-ethoxylphenyl)-1-methyl-3-propyl-1H-pyrazolo[4,3-d]pyrimidin-7(6H)-one (4′)1H-NMR (400 MHz, CDCl3) δ 10.9 (br s, 1H), 8.78 (d, J=2.8 Hz, 1H), 7.72 (dd, J=8.8, 2.4 Hz, 1H), 7.07 (d, J=8.8 Hz, 1H), 6.54 (s, 1H), 4.34 (q, J=7.2 Hz, 2H), 4.27 (s, 3H), 2.94 (t, J=7.6 Hz, 2H), 1.88 (sextet, J=7.6 Hz, 2H), 1.62 (t, J=7.2 Hz, 3H), 1.03 (t, J=7.6 Hz, 3H); 13C-NMR (100 MHz, CDCl3) δ 157.0, 154.0, 147.7, 147.0, 138.8, 133.0, 132.3, 131.6, 128.2, 124.7, 120.3, 114.6, 112.8, 65.8, 38.4, 28.0, 22.5, 14.9, 14.3; IR (neat) 1687, 1561, 1492, 1319, 1244, 1157, 1035, 920, 812, 752, 682; mp 170.5°C; ESI-MS m/z 407 (M+H).
This work was supported by the NRF grant funded by the Korea Food and Drug Administration (Grant No. 12162KFDA101).
