Ultrasound-assisted regioselective synthesis of aminomethylated daidzein derivatives via Mannich reaction

1 Introduction

Daidzein (7,4′-dihydroxyisoflavone) is one of the most abundant isoflavonoids found in legumes, especially soybeans. It has been used for the prevention of cardiovascular disease, lessening the symptoms of the menopause and protection against osteoporosis [1–3]. It was also shown to inhibit proliferation of different cancer-derived cell lines in culture and reduce the multiplicity, but not incidence, of mammary tumors in rats [4, 5]. One of the causes of its effect on cell growth may be inhibition of specific protein kinases [6, 7]. Furthermore, daidzein is also known to be an alcohol-dehydrogenase (ADH) inhibitor [8]. However, due to the low solubility of daidzein in water, its absorption upon oral administration is quite limited, which makes it difficult to be used as a natural additive in foodstuff or in medicine. In order to improve the adsorption and bioavailability of daidzein, a series of daidzein derivatives have been synthesized through different procedures in recent years. Through a convenient Atherton-Todd reaction, Chen and Xiao synthesized a series of 7-O-phosphorylated daidzein derivatives [9, 10]. Using 2,2,2-trifluoro-N-(p-methoxyphenyl)acetamidates as the glycosyl donors, 7-O-glycosides of daidzein derivatives have also been synthesized by a glycosylation procedure [11, 12]. Employing a key protecting group strategy, daidzein 4′-sulfate, daidzein 7-sulfate and daidzein 4′,7-disulfate were synthesized regiospecifically [13]. Despite a number of daidzein derivatives having been synthesized through different procedures, most of the derivatization focused on the reaction of the substituent at the 7- and/or 4′-position of daidzein, and reactions occurring directly at 8-position or other positions of daidzein are quite rare.

The Mannich reaction has been reported to be an efficient method for the introduction of a basic (alkylamino)methyl moiety to construct various nitrogen-containing compounds in pharmaceuticals featuring better water solubility and bioavailability [14]. Various drugs obtained from the Mannich reaction were found to be more effective and less toxic than parent compounds [15, 16]. Through the Mannich reaction, many nitrogen-containing flavonoid analogues have been synthesized as potential anticancer agents [17, 18]. However, the wide application of this useful reaction is limited due to some drawbacks, such as lower yield and relatively long reaction time.

Ultrasound irradiation has been demonstrated as an alternative energy source for organic reactions ordinarily accomplished by heating [19, 20]. Many homogeneous and heterogeneous reactions can be conducted smoothly by ultrasonic activation to provide improved yields and increased selectivities [21–23]. Our group has observed that the reaction rate was notably accelerated under the ultrasound irradiation during the synthesis of chrysin derivatives linked with 1,2,3-triazoles [24]. Here, we report an improved procedure for the synthesis of nitrogen-containing daidzein derivatives through a Mannich reaction under ultrasound irradiation. A series of 7,4′-dihydroxy-8-aminomethyl-isoflavone and 7,4′-dihydroxy-3′,8-bis(aminomethyl)-isoflavone derivatives were regioselectively obtained through Mannich reaction under ultrasound irradiation.

2 Results and discussion

The A ring of daidzein, especially positions C-6 and C-8, might be more reactive toward electrophiles than the C ring (Fig. 1). Other positions in daidzein, such as the C-3′ or C-5′ of the C ring could also potentially be involved in such Mannich reactions. Previous studies on the reaction of daidzein with electrophiles under a variety of conditions have shown that mixtures often result through substitution at different positions, including C-6 and C-8 of the A ring and C-3′ or C-5′ of the C ring [25, 26]. In order to acquire a pure, single product, one frequently adopted approach is to protect one or both phenolic hydroxyl groups prior to the reaction. However, such an indirect approach usually requires multi-step transformations. Obviously, it could be of great value to find a regioselevtive Mannich reaction, which would allow the addition of extended functionalized aminomethyl groups to daidzein without the need for protecting groups. Although the 4′- and 7-phenolic hydroxyl groups in daidzein are chemically similar, fine differences exist in their chemical electronic environments. We expected the C-8 of daidzein to be the most reactive site for electrophillic reactions. Optimization of appropriate Mannich conditions under the help of ultrasound irradiation may provide a highly regioselective aminomethylation of daidzein.

Fig. 1:

Structure of daidzein (1) and possible sites for the Mannich reaction.

Initially, the aminomethylation reaction of daidzein (1), formaldehyde (2), and dipropylamine (3a) under ultrasound irradiation (150 W, 45 KHz, 65 °C) in DMF was used to test the regioselectivity and the isolated product was identified as the 8-aminomethylated daidzein (4a; Scheme 1). As the formation of two regioisomers 6-aminomethylated and 3′- (or 5′-) aminomethylated daidzein derivatives seemed possible, unambiguous structure determination of the obtained product was quite crucial here. The structure of the resulting 8-aminomethylated daidzein was characterized by NMR and mass data. The mass spectrum of 4a revealed a peak at m/z = 368.1854 [M+H]⁺, indicating the formation of mono-aminomethylated daidzein derivative. The ¹H NMR spectra of the isolated product showed the absence of the signal at δ = 6.88 ppm for 8-proton of daidzein. Moreover, the ¹³C NMR spectra showed that the signal of C-8 for 4a was shifted downfield to δ = 108.7 ppm compared to δ = 102.5 ppm for that of daidzein. Therefore, NMR and MS data of the product provided strong evidence for the formation of regioisomer 4a and ruled out the other alternative regioisomers.

Scheme 1:

The model reaction for optimization of the reaction conditions.

The reaction condition was further optimized to improve the yield of 4a. Factors such as solvent, ultrasonic power, frequency, temperature, and reaction time were investigated. Solvents including EtOH, iPrOH, DMF, tBuOH, EtOH-H₂O, iPrOH-H₂O, DMF-H₂O, tBuOH-H₂O (1:2, v/v), and H₂O were used, and the result showed that tBuOH-H₂O (1:2, v/v) was the best reaction solvent. In view of the different solubility of daidzein (1) and aminomethylated daidzein (4a) in tBuOH or H₂O, the ratio of tBuOH and H₂O was further optimized. When the ratio of tBuOH-H₂O is lower than 4:10, the product yields increased as the ratio of tBuOH-H₂O increased. A further increase of the ratio of tBuOH-H₂O led to decrease the product yields. This might be tentatively attributed to the formation of bis(aminomethylated) daidzein derivatives or other byproducts. In order to acquire a pure, single Mannich product, tBuOH-H₂O (2:10, v/v) was chosen as solvent. The power (Table 1, entries 1–3) and irradiation frequency (Table 1, entries 2, 4, and 5) of ultrasound irradiation were investigated and the best result was obtained with the power 150 W and irradiation frequency 45 KHz (Table 1, entry 2). Investigation of the reaction temperature showed that raising the reaction temperature increased the yield (Table 1, entries 2, 6–9), and the product was obtained with a higher yield at 65 °C, (Table 1, entry 2). The proper reaction time was found to be 2 h (Table 1, entries 2, 10–13). The optimal molar ratio of daidzein and dipropylamine (3a) was 1:1, and the yield of 4a decreased when an excess 3a was used. A reaction time longer than 2 h and usage of excess 3a led to decreased yields, this might be attributed to the formation of bis(aminomethylated) daidzein derivatives. Compared with magnetic stirring (Table 1, entries 14–15), the use of ultrasound irradiation leads to faster reaction and higher yield.

Under the optimized reaction conditions, the scope and generality of this protocol for the synthesis of regioselective 8-aminomethylated daidzein derivatives was tested. As shown in Scheme 2, using primary amines, secondary amines, and heterocyclic amines as reactants, the corresponding 8-aminomethylated daidzein derivatives 4a–g were obtained in excellent yields. All the reactions were carried out under similiar conditions in the absence of ultrasound irradiation (Scheme 2), and the results showed that the yields decreased markedly within the same time. Thus, ultrasound showed a beneficial effect on the synthesis of 8-aminomethylated daidzein derivatives and improved the yields of the products significantly.

Scheme 2:

Synthesis of 7,4′-dihydroxy-8-aminomethyl-isoflavone derivatives. Reaction conditions: (a) the molar ratio of daidzein/amine was 1:1, tBuOH-H₂O=2:10 (v/v), 65 °C, 2 h; footnotes: (b) the yield of the reaction with heating/stirring but without ultrasound; (c) the yield of the reaction with ultrasound irradiation (150 W, 45 Hz).

On the basis of the successful regioselective synthesis of 8-aminomethylated daidzein derivatives under unltrasound irradiation, we extended our study to the synthesis of bis(aminomethylated) daidzein derivatives under similar conditions. Structural analysis indicated that C-6 of the A-ring and C-3′(5′) are both the second reactive position. Using tBuOH-H₂O (1:1, v/v) as the co-solvent, daidzein (1), formaldehyde (2), and dipropylamine (3a) as the reactants (molar ratio of daidzein/dipropylamine, 1:2), under ultrasound (150 W, 45 KHz) at 65 °C for 5 h, the reaction produced 3′,8-bis(aminomethylated) daidzein derivative (5a). The corresponding 3′,8-bis(aminomethylated) daidzein derivatives 5h and 5i were also synthesized under similar conditions (Scheme 3). Increasing the amount of amines was beneficial for the formation of bis(aminomethylated) daidzein derivatives.

Scheme 3:

Synthesis of 7,4′-dihydroxy-3′,8-bis(aminomethyl)-isoflavone derivatives. Reaction conditions: (a) the molar ratio of daidzein/amine was 1:2, tBuOH-H₂O=1:1 (v/v), 65 °C, 5 h; footnotes: (b) the yield of the reaction with heating/stirring but without ultrasound; (c) the yield of the reaction with ultrasound irradiation (150 W, 45 Hz).

The structures of 3′,8-bis(aminomethylated) daidzein derivatives 5a, 5h and 5i are elucidated on the basis of HRMS, ¹H NMR, ¹³C NMR and 2D NMR (DEPT, ¹H-¹H COSY, HSQC and HMBC spectra). For example, the mass spectrum of 5a reveals a peak corresponding to its molecular ion at m/z = 481.3058 [M+H]⁺. In the ¹H NMR spectrum of compound 5a, two doublet signals at δ = 7.88 and 6.82 ppm are attributed to 5-H and 6-H, indicating that 6-position of daidzein is not the second aminomethylated position. The signals at δ = 7.29 (m, 2H) and 6.74 (d, 1H) ppm were attributed to three hydrogen atoms of C ring of 5a, which shows that 3′ or 5′ position of 5a was aminomethylated. Structure elucidation was further achieved on the basis of the ¹H-¹³C connectivities (HMBC). In the ¹H-¹³C correlation spectra of 5a, the methylene protons at δ = 4.07 ppm showed correlation with C-7 (164.5 ppm), C-8 (108.7 ppm) and C-9 (155.2 ppm), and the methylene protons at δ = 3.74 showed correlation with C-2′ (129.7 ppm), C-3′ (122.7 ppm) and C-4′ (157.8 ppm), confirming that the methylene groups are linked to C-8 and C-3′, respectively, which is in accordance with the structure of 5a.

3 Conclusion

In summary, an ultrasonic-assisted protocol for the regioselective synthesis of novel 7,4′-dihydroxy-8-aminomethyl-isoflavone derivatives and 7,4′-dihydroxy-3′,8-bis(aminomethyl)-isoflavone derivatives utilizing Mannich reaction is reported. The regioselectivity was rationalized and confirmed by NMR spectroscopic data.

4 Experimental section

5 Supplementary information

Pictures of the ¹H and ¹³C NMR spectra of 4a–4j, 4′a, 4′h, 4′i, as well as two-dimensional NMR spectra of 4′a, are given as supplementary information (DOI: 10.1515/znb-2015-0068).