Introduction

Botrytis cinerea and Monilinia fructicola (Winter) Honey are superior phytopathogenic fungi of the ascomycete family. They are responsible for “gray mold” and “brown rot” diseases that attack a wide variety of fruits, vegetables, and field crops around the world (e.g., grapes), causing significant economic losses in the pre- and postharvest stage in the crops [1,2,3,4]. In fact, fungus control is very important in grape-producing countries such as Chile, France, Germany, Italy, South Africa, and the USA, as well as in the wine-producing and exporting countries [5].

In this context, over the past decades, chemical fungicides have been used in pre- and postharvest periods to prevent and control the diseases caused by both of these pathogens. However, the fungi have developed resistance to some conventional fungicides, particularly benzimidazoles and dicarboximides [6]. In fact, these compounds cause severe damage to the environment, human beings, and beneficial microbiota in agriculture and should be replaced with less toxic compounds [7].

Due to the mentioned above, phenolic compounds emerge as a potential source of control of phytopathogen [8,9,10,11,12]. For example, chalcones are natural compounds that belong to the flavonoid family [13]. These compounds have attracted great interest due to their wide range of pharmacological properties, including mainly anti-inflammatory, analgesic, anti-pyretic, anti-mutagenic, and anti-leishmanial properties, the anti-proliferative effect they have on cancer cell lines, and their antifungal effects [14,15,16,17,18,19,20]. Moreover, chalcones have several applications in agriculture, as insect antifeedant [8], antifungal [9], and larvicidal [10] activities have also been demonstrated for chalcone derivatives. For example, several halogenated chalcones (1a–f) have been tested against B. cinerea (see Fig. 1), exhibiting growth inhibition activity values between 28 and 67% at the concentration of 100 µg/mL, [11] while natural flavonoids (2a–i) tested on the same phytopathogen show weak antifungal activity with growth inhibition activity between 2.0 and 37% [12] at the concentration of 40 µg/mL. However, there is currently no information regarding the effect of chalcones and flavonoids on M. fructicola.

Fig. 1
figure 1

Halogenated chalcones and natural flavonoids with an inhibitory effect on B. cinerea

Full size image

A current trend in the discovery and development of highly active compounds is the hybridization of two or more active fragments that may present improved pharmacological activities [21, 22]. Chromanes are small, natural compounds, and fragments of other more complex natural products that are used in this manner. They have attracted intense interest because of their numerous biological activities such as antimicrobial, allergenic, plant growth inhibitory, and anti-herbivore activities and anti-proliferative effects against cancer cell lines [23, 24]. In addition, the saturated derived structure known as dihydrochromane (or tetrahydropyran) is an important structural fragment of the molecules in many biologically active and natural compounds [25, 26], and in particular, antibiotic activity has been identified for this fragment type [27]. As mentioned above, the hybridization of chalcone and dihydrochromane fragments may lead to good B. cinerea and M. fructicola in vitro inhibition growth mycelial activity (see Fig. 2).

Fig. 2
figure 2

Chalcone and dihydrochromane structures hybridization for rational design

Full size image

Since the synthesis of organic compounds focused on obtaining a solution for the control of B. cinerea and M. fructicola postharvest diseases has not been explored in depth, and due to the potential fungicide applications of chalcones, we synthesized ten chalcones (7a–j) and five dihydrochromane–chalcones hybrids (7k–o) that are reported here for the first time. In addition, all compounds were evaluated for antifungal activity against B. cinerea and M. fructicola, and their quantitative structure–activity relationship was studied.

Results and discussion

Chemistry

The compound 1-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)ethanone (3) was isolated from methanolic extract of Senecio graveolens by column chromatography and identified by spectroscopic techniques (IR, NMR, and MS), according to the procedures described in our previous report [28]. Then, compound 3 was converted into 1-(2,2-dimethylchroman-6-yl)ethanone (4) by prenyl cycling the group in acidic media [29]. In this reaction, formic acid was used at room temperature, obtaining a crystalline solid with excellent yield (96%, see Scheme 1). The 1H-NMR spectrum shows two triplet signals at δ = 2.73 and δ = 1.75 ppm (J = 6.7 Hz), each one with two hydrogen atoms corresponding to the benzylic and homobenzylic CH2 of the dihydrochromane skeleton. Similarly, the 13C-NMR spectrum shows a quaternary carbon signal at δ = 75.5 ppm corresponding to the carbon bonded to the oxygen of dihydrochromane and two methyl groups. In addition, the spectroscopic data are consistent with the previous report [30]. The final compounds (7a–o) were synthesized using semisynthetic acetophenone (4) and acetophenones (5a–b) with commercial benzaldehydes (6a–e) by Claisen–Schmidt condensation in alkaline media, showing acceptable to excellent yields (27–99%, see Scheme 1).

Scheme 1
scheme 1

Synthetic step to obtain compound 4 and final compounds 7a–o

Full size image

The structures of the final compounds (7a–o) were determined using spectroscopic evidence (IR, NMR, and MS, see Electronic Supplementary Material spectra S1–S39). Infrared spectra of all compounds show an absorption peak of the typical conjugated carbonyl group (υ ~ 1660 cm−1). In the 1H-NMR, two hydrogen atoms coupling as a doublet downfield are observed (δ ~ 7.78 and 7.42 ppm, J ~ 15.6 Hz), corresponding to β- and α-hydrogen atoms with trans geometry. For known compounds (7a–j), spectroscopic information was compared with previous reports [31,32,33,34,35]. In addition, for the compounds (7k–o) reported for the first time, the 13C-NMR and bidimensional NMR experiments (2D-HSQC and 2D-HMBC) showed two carbon signals of CH at δ ~ 143 and 117 ppm bonded to Hβ and Hα, respectively. Moreover, using the 2D-HMBC experiment, it was shown that this hydrogen (Hβ and Hα)was correlated with the quaternary carbons (δ ~ 130 and 120 ppm) of both aromatic rings and the carbonyl group, confirming the Ar–CO–CH = CH–Ar system that is typical of the chalcone structure. Finally, the complete structural assignment of new compounds was carried out using 2D-HSQC and 2D-HMBC experiments, and mass spectrometry was used to complement this information.

Antifungal evaluation

All compounds were tested in vitro using the radial growth rate assay, and it was found that they inhibit growth compared with the negative control (carrier solvent) [36, 37]. The inhibition concentrations that caused 50% mycelia inhibition growth of B. cinerea and M. fructicola for each compound (7a–o) were calculated. The results for the tested compounds on B. cinerea showed that the activities range between 43.9 and 502.8 µg/mL, while they range between 48.5 and 330.2 µg/mL for M. fructicola. The values obtained for both ascomycetes are summarized in Table 1.

Table 1 Half-inhibition concentration values (IC50) of compounds 7a–o on mycelial growth of B. cinerea and M. fructicola
Full size table

For the set of samples tested against B. cinerea, 7a and 7c compounds have similar activity to that of Captan® (p > 0.05, see Table 1 and Electronic Supplementary Material Fig. S1). The most active compounds (7a and 7c) are small and structurally simple chalcones and show better activity values than the more complex molecules such as the pyrazolo[1,5-a]pyrimidine derivatives reported by Zhang et al. [38] Similarly, comparing our results with other nitrogen heterocycles such as the azoles, 7a and 7c have similar activities to some of the compounds reported by Zhang et al. [39]. In contrast, for M. fructicola, compounds 7a, 7d, 7e, 7f, and 7n have similar activities to that of the positive control Mystic® (p > 0.05); however, there is no information on the effect of chalcone-structure-related compounds on M. fructicola.

Comparing the influence of the methoxy group on ring A (compounds 7f–j) with the hydrogen substituent (7a–e) showed that the methoxy group decreases the activity in B. cinerea and the same effect is shown in M. fructicola (see Table 1). For the new compounds that contain a dihydrochromane fragment (compound 7k–o), the activity on B. cinerea declines for all compounds, while for M. fructicola the effects do not change with the presence of this fragment. The results show that structural modifications of the donor electron group (e.g., OMe) or lipophilic fragment (e.g., dihydrochromane) are not the key features of an increase in the inhibition of activity on B. cinerea and M. fructicola, while an electronegative group (e.g., F, Cl, or Br) could slightly increase this property [11].

Comparing the substituents in ring B and their effect on the mycelial growth inhibition activity on B. cinerea, it was found that the most active substituent is hydrogen (7a, 7f, and 7k), while the 3,4-dioxomethylen fragment decreases mycelial growth inhibition activity (7f and 7o), except in compound 7e. For M. fructicola, the hydrogen substituent increased the mycelial growth inhibition activity (7a and 7f), except in the compound 7n that has a 4-NMe2 fragment, while the substituent in ring B that decreases the activity is the 4-OH fragment (7b, 7g, and 7l). In this sense, several biological activities of chalcones and structurally related compounds have been linked to their substituent on aromatic rings, e.g., the presence of the hydroxyl group affects the mycelial growth inhibition in B. cinerea and may be related to its cell death mechanism on this phytopathogen [40].

On the other hand, compounds 7a, 7e, 7f, and 7k showed high activity in both phytopathogens (IC50 < 90 µg/mL, see Table 1). Compounds 7a, 7f, and 7k have a hydrogen substituent on ring B. Moreover, compound 7c shows selectivity for B. cinerea (threefold higher activity than for M. fructicola, see Table 1), and a similar trend was shown by 7h, while both compounds have the 4-OMe substituent in ring B, the dihydrochromane–chalcone with the same substituent (7m) has no selectivity for B. cinerea.

In addition, only the dihydrochromane–chalcone compound 7n shows selectivity for M. fructicola (more sixfold higher activity than for B. cinerea). However, compounds 7d and 7i that have the dimethylamino group linked to ring B have no specific action on either of the phytopathogens (see Table 1).

The global analysis of the relationship between the structural features and fungicidal activity of the tested compounds is presented in the following section.

Structure–activity relationship study

Quantitative structure–activity relationship (QSAR) studies seek to correlate biological activity (e.g., pIC50) with different physicochemical descriptors of a series of compounds. For this purpose, multilinear equations are sought between the activity (dependent variable) and various physicochemical descriptors (independent variables), or structural parameters (e.g., Free–Wilson descriptors). Therefore, the importance of QSAR equations is that they allow the interpretation of the biological results obtained based on the physicochemical properties and the structure of the molecules, and, on the other hand, they allow the design and prediction of biological activity of new molecules not yet synthesized.

To elucidate the structure–activity relationship of the compounds evaluated against B. cinerea and M. fructicola, a total of 70 descriptors were calculated (see “Materials and methods” section). The formulation of the equations was carried out using a complete training set, as reported in other QSAR works done with a limited number of molecules [41]. The calculations were done in the gaseous phase and in the solvent phase. Multivariate correlations between the descriptors and the biological activity expressed as pIC50 were sought according to the procedures described in our previous reports [35, 42]. For both phytopathogens, the best models were obtained in the gas phase (see Electronic Supplementary Material). The final equations were selected based on the values of q2 and r2, selecting those with the least number of chemical descriptors.

$$ {\mathbf{pIC}}_{50} = 3.899{-}0.050{\mathbf{HA}}^{2} {-}0.468{\mathbf{HD}}{-}1.923{\mathbf{C}}_{5}^{2} $$
(1)
$$ n = \, 12,\;r = \, 0.921,\; \, r^{2} = \, 0.848,\;{\text{ SD}} = \, 0.152,\; \, F = 14.9,\;n1 = \, 3.446 \times 10^{ - 1} ,\; \, n2 = \, 2.309 \times 10^{ - 2} ,\;q^{2} = 0.821 $$

Equation (1) corresponds to the QSAR model for B. cinerea. It is observed that inhibitory activity depends on the number of hydrogen bond acceptor (HA) and donor (HD) atoms. The use of these descriptors in QSAR studies has been indicated to be significant due to their importance in the modes of action of the drugs [43, 44]. The activity shows a nonlinear dependence on the number of hydrogen bond acceptor groups. Therefore, the presence of more than one hydrogen bond acceptor group would significantly reduce the activity. On the other hand, the mycelial growth inhibition activity decreases with the square of the electrical charge on the C5 carbon atom. Using this information and using compound 7c as the template (Fig. 3a), 22 compounds were proposed and the charge at C5 was calculated (see Electronic Supplementary Material Table S5). It was found that fluorine or chlorine atoms bonded to C2, with methyl group bonded to C5 position, decrease the electron population on C5 (see Electronic Supplementary Material Table S5, compounds 7 and 8), Therefore, reducing the atomic charge to a value close to zero in C5 increases the antifungal activity in B. cinerea. Additionally, in the simplest compound (7a), an appropriate atomic charge distribution was achieved with the 3,4-dibromide, 3-fluorine, 3-bromine-4-chlorine, and 3-methyl substitutions (see Electronic Supplementary Material Table S5, compounds 15, 16, 20, and 22).

Fig. 3
figure 3

Electrostatic potential maps of the compounds evaluated in B. cinerea. a Most active compound 7c. b Least active compound 7g

Full size image

Figure 3 shows the electrostatic potential map of compounds 7c and 7g that are the most active and least active compounds of this series of compounds, respectively. It is observed that the insertion of a methoxy group in compound 7c significantly reduces the charge density on the C5 position (green color in 7c, see Fig. 3a) compared to that for 7g (yellow color, see Fig. 3b), which indirectly reduces the red surface on the carbonyl oxygen (see Fig. 3). Therefore, the insertion of electron-donating groups in the para-position with respect to the carbonyl group will be favorable for the activity.

Equation (2) describes the QSAR model for M. fructicola. It is observed that the inhibitory activity depends on the dipole moment (DM) and the electron density on the C1′ carbons and CO. The DM is related to the size and shape of the molecules and to the heterogeneity of the charges on the molecular surface [45, 46]. DM has been previously used in QSAR models to explain the insecticidal activity of spinosyns and spinosoids [47].

$$ {\mathbf{pIC}}_{50} = 1.600 + 0.014{\mathbf{DM}}^{2} + 9.754{\mathbf{C}}_{{1^{\prime } }}^{2} + \, 3.372{\mathbf{CO}}^{2} $$
(2)
$$ n = 13,\; \, r = 0.926,\; \, r^{2} = 0.857, \, \;{\text{SD}} = 0.131,\; \, F = 17.9,\;n1 = 3.068 \times 10^{ - 1} ,\; \, n2 = \, 1.710 \times 10^{ - 2} ,\;q^{2} = \, 0.833 $$

The biological activity shows a more significant dependence on the electric charges on the carbonyl carbon. This suggests that the carbonyl group plays an important role in the mechanism of inhibitory action, possibly through hydrogen bonding with the target, and Michael-type reactions [48]. Figure 4 shows the potential electrostatic maps for the most active (7n) and least active (7b) compounds for M. fructicola. Comparison of these two compounds shows that 7n has a higher electron density on the oxygen atom (red color, Fig. 4a). This is due to the resonance effect of the dihydrochrome system on ring A and the resonance of the dimethylamine group on ring B. This increased polarization of the carbonyl group leads to increased antifungal activity (Eq. 2). On the other hand, the lower negative charge density on the oxygen atom of the carbonyl group in compound 7b (red color, Fig. 4b) leads to a decrease in the carbon polarization of this functional group (green color, Fig. 4b) and, consequently, the antifungal activity of chalcones for M. fructicola is decreased.

Fig. 4
figure 4

Electrostatic potential maps of the compounds evaluated for M. fructicola. a Most active compound 7n. b Least active compound 7b

Full size image

Moreover, for C1′ atomic charge, compound 7n is more negative than 7b (yellow in Fig. 4a and green color in Fig. 4b). Thus, an electron donor substituent on the ortho-position of C1′ will increase the electron density on C1′ and will also increase the antifungal activity of chalcone for M. fructicola (see Electronic Supplementary Material Table S6).

Conclusions

In summary, 15 compounds were synthesized and characterized by classical spectroscopic techniques, of which five are reported here for the first time (7k–o). In addition, all compounds were tested against B. cinerea and M. fructicola, obtaining two and five compounds, respectively, with fungicide activity similar to a commercial control (Captan® and Mystic®, respectively). Using the antifungal activity results, quantitative structure–activity relationship models were developed obtaining two models with good statistical parameters (q2 and r2 higher than 0.8), identifying the key structural features for the design of new molecules with chalcone as the pharmacophoric core.

Materials and methods

General

Melting point was measured using a Fischer Scientific apparatus (Pittsburgh, PA, USA). Infrared spectra were recorded using the Buck Scientific M500 instrument (Norwalk, CT, USA). The recorded range of the IR spectra was 600 cm−1–4000 cm−1, and all samples were examined using ATR (attenuated total reflectance) system. 1H-NMR, 13C-NMR, 2D-HSQC, and 2D-HMBC spectra were recorded using a Bruker Avance 400 digital NMR spectrometer (Berlin, Germany), operating at 400.13 MHz for 1H and 100.6 MHz for 13C. Chemical shifts are reported in δ (ppm downfield from the TMS resonance), and coupling constants (J) are given in Hz. GC–MS was carried out using an Agilent Technologies 6890 instrument (Santa Clara, CA, USA) with automatic ALS and HP MD 5973 mass detector in the splitless mode. The high-resolution mass spectrometry electronic impact (HR-EI-MS) measurements were taken with a VG Autoespect mass spectrometer (Fision, Ipswich, UK).

Plant material and extraction procedure

Senecio graveolens was collected from an area near Chungara Lake at 4500 m.a.s.l. (Chile). The dry plant material (principally flowers, leaves, and stems, total 180 g) was macerated in 95% ethanol (2 × 500 mL) for 72 h, according to the procedures described in our previous reports [28]. The specimen collection is conserved at CODECITE-CIHDE, Arica, Chile.

Chemistry

(4-Hydroxy-3-(3-methylbut-2-enyl)phenyl)ethanone (3)

This compound was separated from dry methanol extract (52.8 g) by column chromatography using EtOAc/hexane (1:9), obtaining a pale yellow solid (1.09 g, 0.6% yield). MP: 95–96 °C. The spectroscopic information (IR, 1H-NMR, and 13C-NMR) and the MS analysis results were consistent with the previous report [28].

1-(2,2-Dimethylchroman-6-yl)ethanone (4)

In a 250-mL round-bottomed flask, prenyl-acetophenone 3 (1.0 g) and formic acid (30 mL) were added. The mixture was stirred at room temperature for 24 h. Finally, the acid was neutralized using Na2CO3 5%. This mixture was extracted with EtOAc (3 × 50 mL), and the organic layer was dried with Na2SO4 and separated by column chromatography using an EtOAc–hexane mixture (2:8) obtaining 0.955 g of colorless solid (96% yield). MP: 91–92 °C. IR: υ/cm−1 2991, 2933, 1665, 1613, 1482, 1339, 1327, 1230, 1193; 1H-NMR (400 MHz, CDCl3): δ 7.65 (1H, d, J = 3.4 Hz, H2), 7.62 (1H, dd, J = 8.4, 3.4 Hz, H6), 6.71 (1H, d, J = 8.4 Hz, H5), 2.73 (2H, t, J = 6.7 Hz, CH2 -C1′), 2.45 (3H, s, CH3CO), 1.75 (2H, t, J = 6.7 Hz, CH2-C2′), 1.27 (6H, s, CH3-C4′ + CH3-C5′). 13C-NMR (100 MHz, CDCl3): δ 197.0 (C=O), 158.6 (C4), 130.5 (C2), 129.3 (C1), 128.3 (C6), 120.7 (C3), 117.2 (C5), 75.5 (C3′), 32.5 (C2′), 26.9 (CH3CO + C4′*), 26.3 (C5′*), 22.3 (C1′). * Interchangeable signals. 1H-NMR, 13C-NMR, and MS analyses results are consistent with the previous report [30].

General procedure for chalcone synthesis (7a–o)

To a dry, 100-mL round-bottomed flask, acetophenone 4 or 5a–b (250 mg, between 1.22 and 2.08 mmol) and commercial benzaldehyde 6a–e (1.2 molar equivalents) were added. Both reagents were solubilized in ethanol (5 mL), a NaOH-saturated solution was added (in 10 mL of ethanol), and the mixture was stirred for 48 h, after which 5% HCl solution was added until pH ~ 7 to end the reaction, and the mixture was extracted with EtOAc (3 x 30 mL). The organic layer was dried with Na2CO3, filtered, and separated with column chromatography using a hexane/EtOAc mixture increased polarity, obtaining compounds 7a–o in yields between 27 and 99%.

(2E)-1,3-diphenylprop-2-en-1-one (7a)

Yellow solid (99% yield). MP: 65–69 °C. IR: υmax/cm−1 3069, 2970, 1682, 1606, 1518, 1420; 1H-NMR (400 MHz, CDCl3): δ 8.03 (2H, d, J = 7.5 Hz, H2′ + H6′), 7.82 (1H, d, J = 15.6 Hz, Hβ), 7.65 (2H, m, H2 + H6), 7.59 (1H, t, J = 7.5, Hz, H4), 7.54 (1H, d, J = 15.6 Hz, Hα), 7.52 (2H, d, J = 7.5 Hz, H3 + H5), 7.42 (3H, m, H3′ + H5′ + H4′); 13C-NMR (100 MHz, CDCl3): δ 190.5 (C = O), 144.8 (Cβ), 138.2 (C1), 134.8 (C1′), 132.7 (C4), 130.5 (C4′), 128.9 + 128.6 + 128.5 + 128.4 (C2 + C3 + C5 + C6 + C2′ + C3′ + C5′ + C6′), 122.1 (Cα). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [35].

(2E)-3-(4-hydroxyphenyl)-1-phenylprop-2-en-1-one (7b)

Orange solid (85% yield). MP: 183–187 °C. IR: υmax/cm−1 3421, 3024, 1647, 1594, 1566, 1513, 1180; 1H-NMR (400 MHz, CDCl3): δ 8.11 (2H, d, J = 8.4 Hz, H2 + H6), 7.75 (1H, d, J = 15.6 Hz, Hβ), 7.71 (2H, d, J = 8.9 Hz, H2′ + C6′), 7.61 (1H, d, J = 7.7 Hz, H4), 7.54 (2H, d, J = 7.7 Hz, H3 + H5), 7.53 (1H, d, J = 15.6 Hz, Hα), 6.92 (2H, d, J = 8.9 Hz, H3′ + H5′); 13C-NMR (100 MHz, CDCl3) δ 189.9 (C=O), 160.8 (C4′), 145.2 (Cβ), 139.4 (C1), 133.3 (C4), 131.5 (C2′ + C6′), 129.4 (C3 + C5), 129.1 (C2 + C6), 127.5 (C1′), 119.6 (Cα), 116.7 (C3′ + C5′). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [35].

(2E)-3-(4-methoxyphenyl)-1-phenylprop-2-en-1-one (7c)

Yellow solid (99% yield). MP: 70–72 °C. IR: υmax/cm−1 3066, 2929, 1662, 1596, 1546, 1511, 1466, 1239, 1214; 1H-NMR (400 MHz, CDCl3): δ 8.01 (2H, d, J = 7.5 Hz, H2 + H6), 7.89 (1H, d, J = 15.6 Hz, Hβ), 7.60 (2H, d, J = 8.7 Hz, H2′ + H6′), 7.57 (1H, t, J = 7.5 Hz, H4), 7.49 (2H, d, J = 7.5 Hz, H3 + H5), 7.42 (1H, d, J = 15.6 Hz, Hα), 6.93 (2H, d, J = 8.7 Hz, H3′ + H5′), 3.85 (3H, s, CH3O-C4′); 13C-NMR (100 MHz, CDCl3): δ 190.5 (C=O), 161.6 (C4′), 144.7 (Cβ), 138.4 (C1), 132.5 (C4), 130.2 (C2′ + C6′), 128.5 (C2 + C6), 128.4 (C3 + C5), 127.5 (C1′), 119.7 (Cα), 114.4 (C3′ + C5′), 55.4 (CH3O-C4′). 1H-NMR, 13C-NMR, and MS analyses are consistent with our previous report [35].

(2E)-3-[4-(dimethylamino)phenyl]-1-phenylprop-2-en-1-one (7d)

Orange solid (82% yield). MP: 109–111 °C. IR: υmax/cm−1 3062, 2966, 1644, 1564, 1532, 1486, 1460, 1228, 1167; 1H-NMR (400 MHz, CDCl3): δ 7.99 (2H, d, J = 7.6 Hz, H2 + H6), 7.79 (1H, d, J = 15.5 Hz, Hβ), 7.55 (2H, d, J = 8.6 Hz, H2′ + H6′), 7.54 (1H, m, H4), 7.48 (2H, d, J = 7.5 Hz, H3 + H5), 7.34 (1H, d, J = 15.5 Hz, Hα), 6.73 (2H, d, J = 8.6 Hz, H3′ + H5′), 3.03 (6H, s, (CH3)2N-C4′); 13C-NMR (100 MHz, CDCl3): δ 190.7 (C=O), 145.7 (Cβ), 138.9 (C1), 132.2 (C2′ + C6′), 130.4 (C4), 128.4 (C2 + C6), 128.3 (C3 + C5), 117.2 (Cα), 112.3 (C3′ + C5′), 110.9 (C1′), 40.4 ((CH3)2N-C4′). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [35].

(2E)-3-(1,3-benzodioxol-5-yl)-1-phenylprop-2-en-1-one (7e)

Pale yellow solid (95% yield). MP: 48–50 °C. IR: υmax/cm−1 3085, 2958, 2920, 1659, 1607, 1578, 1468, 1225; 1H-NMR (400 MHz, CDCl3): δ 8.00 (2H, d, J = 7.6 Hz, H2 + H6), 7.73 (1H, d, J = 15.6 Hz, Hβ), 7.57 (1H, t, J = 7.6 Hz, H4), 7.48 (2H, d, J = 7.6 Hz, H3 + H5), 7.36 (1H, d, J = 15.6 Hz, Hα), 7.16 (1H, s, H2′), 7.11 (1H, d, J = 8.0 Hz, H6′), 6.83 (1H, d, J = 8.0 Hz, H5′), 6.00 (2H, s, OCH2O); 13C-NMR (100 MHz, CDCl3): δ 190.2 (C=O), 149.8 (C4′), 148.3 (C3′), 144.6 (Cβ), 138.3 (C1), 132.5 (C4), 129.2 (C1′), 128.5 (C2 + C6), 128.3 (C3 + C5), 125.1 (C6′), 120.0 (Cα), 108.6 (C2′), 106.6 (C5′), 101.5 (OCH2O). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [35].

(2E)-1-(4-methoxyphenyl)-3-phenylprop-2-en-1-one (7f)

Yellow solid (68% yield). MP: 70–72 °C. IR: υmax/cm−1 3078, 2972, 2954, 1655, 1603, 1558, 1508, 1448, 1241, 1190; 1H-NMR (400 MHz, CDCl3): δ 8.01 (2H, d, J = 7.5 Hz, H2 + H6), 7.89 (1H, d, J = 15.6 Hz, Hβ), 7.60 (2H, d, J = 8.7 Hz, H2′ + H6′), 7.57 (1H, t, J = 7.5 Hz, H4′), 7.49 (2H, d, J = 7.5 Hz, H3 + H5), 7.42 (1H, d, J = 15.6 Hz, Hα), 6.93 (2H, d, J = 8.7 Hz, H3′ + H5′), 3.85 (3H, s, CH3O-C4′); 13C-NMR (100 MHz, CDCl3): δ 190.5 (C=O), 161.6 (C4′), 144.7 (Cβ), 138.4 (C1), 132.5 (C4), 130.2 (C2′ + C6′), 128.5 (C2 + C6), 128.4 (C3 + C5), 127.5 (C1′), 119.7 (Cα), 114.4 (C3′ + C5′), 55.4 (CH3O-C4′). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [34].

(2E)-3-(4-hydroxyphenyl)-1-(4-methoxyphenyl)prop-2-en-1-one (7g)

Yellow solid (62% yield). MP: 184–186 °C. IR: υmax/cm−1 3266, 3086, 2949, 1668, 1605, 1558, 1531, 1229, 1146; 1H-NMR (400 MHz, CDCl3): δ 8.03 (2H, d, J = 8.9 Hz, H2 + H6), 7.77 (1H, d, J = 15.6 Hz, Hβ), 7.56 (2H, d, J = 8.4 Hz, H2′ + H6′), 7.42 (1H, d, J = 15.6 Hz, Hα), 6.98 (2H, d, J = 8.9 Hz, H3 + H5), 6.88 (2H, d, J = 8.4 Hz, H3′ + H5′), 3.89 (3H, s, CH3O-C4′). 13C-NMR (100 MHz, CDCl3): δ 188.9 (C=O), 163.3 (C4), 157.7 (C4′), 143.8 (Cβ), 131.3 (C1), 130.7 (C2 + C6), 130.3 (C2′ + C6′), 128.0 (C1′), 119.7 (Cα), 115.9 (C3′ + C5′), 113.8 (C3 + C5), 55.5 (CH3O-C4′). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [34].

(2E)-3-(4-methoxyphenyl)-1-(4-methoxyphenyl)prop-2-en-1-one (7h)

Pale yellow solid (98% yield). IR: υmax/cm−1 3062, 2945, 2931, 1654, 1590, 1569, 1509, 1457, 1420, 1246, 1212; 1H-NMR (400 MHz, CDCl3): δ 8.03 (2H, d, J = 8.8 Hz, H2 + H6), 7.77 (1H, d, J = 15.6 Hz, Hβ), 7.59 (2H, d, J = 8.7 Hz, H2′ + H6′), 7.42 (1H, d, J = 15.6 Hz, Hα), 6.96 (2H, d, J = 8.8 Hz, H3 + H5), 6.92 (2H, d, J = 8.7 Hz, H3′ + H5′), 3.87 (3H, s, CH3O-C4), 3.83 (3H, s, CH3O-C4′); 13C-NMR (100 MHz, CDCl3): δ 188.6 (C=O), 163.2 (C4), 161.4 (C4′), 143.7 (Cβ), 131.2 (C1), 130.6 (C2 + C6), 130.0 (C2′ + C6′), 127.7 (C1′), 119.4 (Cα), 114.3 (C3 + C5), 113.7 (C3′ + C5′), 55.4 (CH3O-C4), 55.3 (CH3O–C4′). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [34].

(2E)-3-(4-N,N-dimethylaminephenyl)-1-(4-methoxyphenyl)prop-2-en-1-one (7i)

Orange solid (98% yield). MP: 122–124 °C. IR: υmax/cm−1 3079, 2979, 2933, 1648, 1579, 1546, 1522, 1435, 1252, 1231, 1162; 1H-NMR (400 MHz, CDCl3): δ 8.03 (2H, d, J = 8.9 Hz, H2 + H6), 7.78 (1H, d, J = 15.4 Hz, Hβ), 7.55 (2H, d, J = 8.6 Hz, H2′ + H6′), 7.36 (1H, d, J = 15.4 Hz, Hα), 6.97 (2H, d, J = 8.9 Hz, H3 + H5), 6.69 (2H, d, J = 8.6 Hz, H3′ + H5′), 3.88 (3H, s, CH3O-C4), 4.04 (6H, s, (CH3)2N); 13C-NMR (100 MHz, CDCl3): δ 188.9 (C=O), 162.9 (C4), 151.9 (C4′), 144.9 (Cβ), 131.8 (C1), 130.5 (C2 + C4), 130.2 (C2′ + C4′), 122.8 (C1′), 116.4 (Cα), 113.6 (C3 + C5), 111.8 (C3′ + C5′), 55.4 (CH3O-C4), 40.1 (N(CH3)2-C4′)). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [34].

(2E)-3-(1.3-benzodioxol-5-yl)-1-(4-methoxyphenyl)prop-2-en-1-one (7j)

Yellow solid (41% yield). MP: 127–133 °C. IR: υmax/cm−1 3078, 2950, 2921, 1661, 1598, 1510, 1466, 1425, 1251, 1218; 1H-NMR (400 MHz, CDCl3): δ 8.03 (2H, d, J = 8.9 Hz, H2 + H6), 7.73 (1H, d, J = 15.5 Hz, Hβ), 7.39 (1H, d, J = 15.5 Hz, Hα), 7.17 (1H, d, J = 1.5 Hz, H2′), 7.12 (1H, dd, J = 8.0, 1.5 Hz, H6′), 6.98 (2H, d, J = 8.9 Hz, H3 + H5), 6.89 (1H, d, J = 8.0 Hz, H5′), 6.03 (2H, s, OCH2O), 3.89 (s, CH3O-C4). 13C-NMR (100 MHz, CDCl3): δ 188.5 (C=O), 163.3 (C4), 149.7 (C4′), 148.3 (C3′), 143.7 (Cβ), 131.2 (C1), 130.6 (C2 + C6), 129.5 (C1′), 124.9 (C6′), 119.8 (Cα), 113.7 (C3 + C5), 108.6 (C2′), 106.6 (C5′), 101.5 (OCH2O), 55.4 (CH3O-C4). 1H-NMR, 13C-NMR, and MS analyses results are consistent with our previous report [34].

(2E)-1-(2,2-dimethylchroman-6-yl)-3-phenylprop-2-en-1-one (7k)

Pale yellow solid (87% yield). MP: 85–87 °C. IR: υ/cm−1 3050, 2975, 2938, 1659, 1604, 1574, 1495, 1448, 1336, 1258, 1230; 1H-NMR (400 MHz, CDCl3): δ 7.83 (1H, dd, J = 8.2, 2.6 Hz, H6), 7.82 (1H, d, J = 2.6 Hz, H2), 7.80 (1H, d, J = 15.7 Hz, Hβ), 7.64 (1H, m, H4′), 7.56 (1H, d, J = 15.7 Hz, Hα), 7.40 (4H, m, H2′ + H3′ + H5′ + H6′), 6.86 (1H, d, J = 8.2 Hz, H5), 2.85 (2H, t, J = 6.7 Hz, CH2-C1″), 1.85 (2H, t, J = 6.7 Hz, CH2-2″), 1.37 (6H, s, CH3-C4″ + CH3-C5″); 13C-NMR (100 MHz, CDCl3): δ 188.7 (C=O), 158.5 (C4), 143.4 (Cβ), 135.1 (C1′), 130.8 (C5), 130.1 (C2), 128.9 (C2′ + C6′), 128.8 (C4′), 128.4 (C6), 128.3 (C3′ + C5′), 121.9 (C3), 120.9 (C1), 117.3 (Cα), 75.5 (C3″), 32.5 (C2″), 26.9 (C4″ + C5″), 22.3 (C1″). EI-MS (+) m/z 292 [M+] (100%). HR-EI-MS (+) 292.1463 calc, 292.1461 found (Δ = 0.0002).

(2E)-1-(2,2-dimethylchroman-6-yl)-3-(4-hydroxyphenyl)prop-2-en-1-one (7l)

Yellow solid (27% yield). MP: 158-160 °C. IR: υ/cm−1 3226, 2971, 2941, 1647, 1602, 1574, 1512, 1446, 1343, 1321, 1231; 1H-NMR (400 MHz, CDCl3): δ 7.83 (1H, d, J = 1.3 Hz, H2), 7.81 (1H, dd, J = 8.5, 1.3 Hz H6), 7.76 (1H, d, J = 15.5 Hz, Hβ), 7.52 (2H, d, J = 8.4 Hz, H2′ + H6′), 7.42 (1H, d, J = 15.5 Hz, Hα), 6.92 (2H, d, J = 8.4 Hz, H3′ + H5′), 6.85 (1H, d, J = 8.5 Hz, H5), 2.84 (2H, t, J = 6.7 Hz, CH2-C1″), 1.84 (2H, t, J = 6.7 Hz, CH2-C2″), 1.36 (6H, s, CH3-C4″ + CH3-C5″); 13C-NMR (100 MHz, CDCl3): δ 190.0 (C=O), 158.7 (C4 + C4′), 144.5 (Cβ), 132.5 (C2), 131.0 (C3), 130.4 (C2′ + C6′), 128.6 (C1′), 127.3 (C6), 121.0 (C1), 119.2 (C5), 117.4 (Cα), 116.1 (C3′ + C5′), 75.7 (C3″), 32.5 (C2″), 26.9 (C4″ + C5″), 22.4 (C1″). EI-MS (+) m/z 308 [M +] (100%). HR-EI-MS (+) calc 308.1412, found 308.1417 (Δ = − 0.0005).

(2E)-1-(2,2-dimethylchroman-6-yl)-3-(4-methoxyphenyl)prop-2-en-1-one (7m)

Yellow solid (97% yield). MP: 79–81 °C. IR: υ/cm−1 3082, 2975, 1655, 1589, 1510, 1492, 1338, 1318, 1227; 1H-NMR (400 MHz, CDCl3): δ 7.83 (1H, d, J = 1.2 Hz, H2), 7,81 (1H, dd, J = 8.4, 1.2 Hz, H6), 7.77 (1H, d, J = 15.6 Hz, Hβ), 7.60 (2H, d, J = 8.7 Hz, H2′ + H6′), 7.43 (1H, d, J = 15.6 Hz, Hα), 6.93 (2H, d, J = 8.7 Hz, H3′ + H5′), 6.84 (1H, d, J = 8.4 Hz, H5), 3.85 (3H, s, CH3O-C4′), 2.85 (2H, t, J = 6.7 Hz, CH2-C1″), 1.85 (2H, t, J = 6.7 Hz, CH2-C2″), 1.37 (6H, s, CH3-C4″ + CH3-C5″); 13C-NMR (100 MHz, CDCl3): δ 188.9 (C = O), 161.4 (C4′), 158.4 (C4), 143.4 (Cβ), 130.7 (C2), 130.4 (C3), 130.0 (C2′ + C6′), 128.3 (C1′), 127.9 (C6), 120.9 (C1), 119.7 (C5), 117.2 (Cα), 114.3 (C3′ + C5′), 77.5 (C3″), 55.4 (CH3O-C4′), 32.5 (C1″), 26.9 (C2″), 22.3 (C4″ + C5″). EI-MS (+) m/z 322 [M+] (100%). HR-EI-MS (+) 322.1569 calc, 322.1560 found (Δ = 0.0009).

(2E)-3-(4-(dimethylamino)phenyl)-1-(2,2-dimethylchroman-6-yl)prop-2-en-1-one (7n)

Red solid (72% yield). MP: 79-82 °C. IR: υ/cm−1 2977, 2922, 1654, 1592, 1558, 1522, 1434, 1366, 1226, 1163; 1H-NMR (400 MHz, CDCl3): δ 7.83 (1H, dd, J = 8.3, 2.2 Hz, H6), 7.80 (1H, d, J = 2.2 Hz, H2), 7.74 (1H, J = 15.5 Hz, Hβ), 7.54 (2H, d, J = 8.8 Hz, H2′ + H6′), 7.36 (1H, d, J = 15.5 Hz, Hα), 6.83 (1H, d, J = 8.3 Hz, H5), 6.69 (2H, d, J = 8.8 Hz, H3′ + H5′), 3.07 (6H, s, (CH3)2N-C4′), 2.85 (2H, t, J = 6.7 Hz, CH2-C1″), 1.84 (2H, t, J = 6.7 Hz, CH2-C2″), 1.36 (6H, s, CH3-C4″ + CH3-C5″);13C-NMR (100 MHz, CDCl3): δ 189.0 (C=O), 158.0 (C4), 151.8 (C4′), 144.5 (Cβ), 130.8 (C1), 130.5 (C6), 130.1 (C2′ + C6′), 128.1 (C2), 122.0 (C1′), 120.8 (C3), 117.1 (Cα), 116.8 (C5), 111.8 (C3′ + C5′), 75.3 (C3″), 40.0 ((CH3)2N-C4′), 32.5 (C2″), 26.9 (C4″ + C5″), 22.4 (C1″). EI-MS (+) m/z 335 [M +] (100%). HR-EI-MS (+) calc 335.1885, found 335.1895 (Δ = 0.0010).

(2E)-3-(benzo[d] [1, 3] dioxol-5-yl)-1-(2,2-dimethylchroman-6-yl)prop-2-en-1-one (7o)

Yellow solid (42% yield). MP: 158–159 °C. IR: υ/cm−1 3052, 2967, 2941, 1652, 1604, 1576, 1490, 1446, 1360, 1320, 1233; 1H-NMR (400 MHz, CDCl3): δ 7.82 (1H, d, J = 1.9 Hz, H2), 7.80 (1H, dd, J = 8.2, 1.9 Hz, H6), 7.72 (1H, d, J = 15.6 Hz, Hβ), 7.39 (1H, d, J = 15.6 Hz, Hα), 7.17 (1H, s, H2′), 7.12 (1H, d, J = 8.0 Hz, H6′), 6.84 (1H, d, J = 8.2 Hz, H5), 6.83 (1H, d, J = 8.0 Hz, H5′), 6.02 (2H, s, OCH2O), 2,85 (2H, t, J = 6.7 Hz, CH2-C1″), 1.85 (2H, t, J = 6.7 Hz, CH2-C2″), 1.37 (6H, s, CH3-C4″ + CH3-C5″); 13C-NMR (100 MHz, CDCl3): δ 188.7 (C=O), 158.5 (C4), 149.6 (C4′), 148.3 (C3′), 143.4 (Cβ), 130.7 (C1), 130.2 (C1′), 129.6 (C2), 128.3 (C6), 124.9 (C5′), 120.9 (Cα), 120.0 (C3), 117.3 (C6′), 108.6 (C5), 106.2 (C2′), 101.5 (OCH2O), 75.5 (C3″), 32.5 (C2″), 26.9 (C4″ + C5″), 22.4 (C1″). EI-MS (+) m/z 336 [M +] (100%). HR-EI-MS (+) calc 336.1362, found 336.1360 (Δ = 0.0002).

In vitro antifungal activity of synthetic compounds against B. cinerea and M. fructicola

The antifungal activity of the synthesized compounds (7a–o) against B. cinerea and M. fructicola was determined using radial growth rate assay in potato dextrose agar (PDA) growth medium (see Electronic Supplementary Materials Fig S1) [49]. The synthesized compounds were dissolved in an ethanol/water solution and were added to a petri dish containing PDA medium at 50 °C. The final tested concentrations were 12.5, 25, 50, 150, 250, and 500 µg/mL for each compound. A mycelium agar disk (4 mm in diameter) of the pathogen fungi was placed in the center of the PDA plates. PDA medium containing 1% ethanol was considered as the negative control (C−), whereas Captan® and Mystic® 520 SC, commercial fungicides (ANASAC, Bayer), were used as the positive control (C+) at the same concentrations and under the same conditions as the test compounds. B. cinerea was incubated for 3 days at 23 °C, whereas M. fructicola was incubated for 1 week at the same temperature in the dark. Each treatment was replicated three times, and each assay was repeated twice. The diameter of the fungi in the cultures was measured, and the inhibition percentages of mycelial growth for each compound were calculated and compared with the negative control as described in a previous report [50].

From mycelial inhibition percentage values and the concentration (µg/mL), the IC50 value was calculated for each compound using a logarithmic equation fit analysis carried out with Origin 8.0 software.

Statistical analysis

The data were reported as the mean values ± standard deviation (SD). One-way ANOVA and post hoc HSD Tukey tests were used with a confidence level of 0.95. The significant differences between the antifungal activity of each compound with those of Captan® or Mystic® were calculated. These statistical analyses were performed using the Statistical 7.0 software.

Computational details

All compounds (7a–o) were optimized using DFT-B3LYP-6-31G (d,p) level of theory calculations, and the optimized structures were verified by frequency calculations (obtaining no imaginary frequencies) in the gas phase and using the IEFPCM (water) model as the solvent phase. The descriptors obtained from quantum mechanical calculations such as the dipolar moment (DM), atomic charge from the electrostatic potential (C1, C2, C3, C4, C5, C6, C1′, C2′, C3′, C4′, C5′, C6′, Cα, Cβ, CO), highest occupied molecular orbital (HOMO), and lowest unoccupied molecular orbital (LUMO) were obtained directly from the output file, while the chemical potential (µ), hardness (η), softness (S), and electrophilic global index (ω)values were calculated using the following equations.

$$ \mu = \frac{{\left( {E_{\text{LUMO}} + E_{\text{HOMO}} } \right)}}{2} $$
(3)
$$ \eta = \frac{{\left( {E_{\text{LUMO}} - E_{\text{HOMO}} } \right)}}{2} $$
(4)
$$ S = \frac{1}{2\eta } $$
(5)
$$ \omega = \frac{{\mu^{2} }}{2\eta } $$
(6)

In addition, steric and topological descriptors such as molecular weight (MW), lipophilicity index (CLogP), molar refractivity (MR), molecular surface (MS), molecular volume (MV), hydrogen bonding acceptor (HA), hydrogen bonding donor (HD), Balaban index (BI), molecular topological index (MTI), rotatable bonds (RT), topological diameter (TD), and Wiener index (WI) were obtained using molecular mechanics (MM) optimization carried out with the ChemDraw software.

Structure–activity relationship study

The structure–activity relationship study was carried out using multiple linear regressions as described in our previous report with small changes [34, 35]. We developed several regression models using pIC50 (− log10(IC50)) in mol L−1 units as the dependent variable and all descriptors mentioned above in the gas phase and in the solvent phase as independent variables (DM, C1, C2, C3, C4, C5, C6, C1′, C2′, C3′, C4′, C5′, C6′, Cα, Cβ, CO HOMO, LUMO, µ, η, S, ω, MW, CLogP, MR, MS, MV, HA, HD, BI, MTI, RT, TD, and WI) in linear and squared form.

In addition, to avoid random correlations between pIC50 and any descriptor, cross-validation was carried out using the Golbraikh method as described by:

$$ q^{2} = 1 - \frac{{\sum \left( {y_{\text{obs}} - y_{\text{calc}} } \right)^{2} }}{{\sum \left( {y_{\text{obs}} - y_{\text{ave}} } \right)^{2} }} $$
(7)

where yobs is the experimental pIC50, ycal is the pIC50 calculated by the QSAR model, and yave is the average pIC50 of all of the compounds used in the QSAR model. An acceptable value of q2 is equal to or higher than 0.5.

Electronic Supplementary Material: 1H-NMR and 13C-NMR of natural, synthetic, and semisynthetic compounds (spectra S1–34); high-resolution mass spectra of new dihydrochromane–chalcone compounds (7k–o, spectra S35–39); structure–activity models for B. cinerea and M. fructicola in gas and solvent phases (Tables S1–S4); effect of compound 7a at different concentrations on in vitro mycelial growth inhibition of B. cinerea and M. fructicola (Fig S1); Table S5: proposed molecules and their C5 atomic charges based on QSAR model of B. cinerea; Table S6: proposed molecules based on QSAR model of M. fructicola.