English

• 1. INTRODUCTION

  Nitrogen containing heterocycle is an important nucleus in synthetic compounds or natural products because of diver sity activity^[1-5]. Among the nitrogen containing heterocycles, pyridine ring possessed various activityies in synthetic small molecules or natural products, such as plant growth regulatory activities^[6], anti-HIV activity^[7], herbicidal activity^{[8, 9]}, fungicidal activity^{[10, 11]}, anti-inflammatory activity^[12], immunosuppressive activity^[13], antioxidant activity^[14], anticancer activity^[15], and so on. Some of pyridine compounds had been commercialized as pesticides or drugs. 2-Chloro-N-(4΄-chloro-[1, 1΄-biphenyl]-2-yl)nicotinamide, whose commercial name is boscalid (SDH inhibitor), is an famous fungicide to protect crops, vegetables and fruits. In this fungicide, pyridine ring and amide group are the key groups. Urea group also had the substrate of amide, which is always found in synthetic or natural compounds^[16-19]. For instance, the chitin synthetase inhibitor dimilin contains urea group, which is an insect growth regulatory.

  In our previous work, lots of heterocyclic compounds with pesticidal activity or medicinal activity were designed and synthesized^[20-30], including the pyridine derivatives. A series of fluorine substituted pyridine acyl urea derivatives were synthesized from boscalid and some compounds exhibited moderate fungicidal activity^[31]. In this work, the fluorine substituted group was replaced by other non-fluorine substitution (Fig. 1). Some of the title pyridine acyl urea derivatives exhibited good fungicidal activity. The SAR was also studied using molecular docking method.

  Figure 1

  

  Figure 1.  Design method of nicotinic acyl urea derivatives

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  2. EXPERIMENTAL

  2.1 Instruments

  Melting points (M.P.) were measured by an X-4 apparatus and uncorrected. ¹H NMR and ¹³C NMR spectra were tested on a Bruker AV III-500 instrument. ESI-HRMS was measured on a JOEL AccuTOF instrument. Single crystal diffraction was done on a Bruker CCD area detector diffractometer.

  2.2 General procedure

  The intermediates 1, 2 and 3 were synthesized according to the reported work^[32-35].

  Syntheses of target compounds 4a~4r

  To a solution of intermediate 3 (0.36 g, 2 mmol) in dichloromethane (10 mL), the substituted aniline (2.2 mmol) was added dropwise and stirred at 20 ^oC for overnight. TLC monitor and then solvent dichloromethane were removed, and the crude products were purified by using flash chromatograph to give compounds 4a-4r.

  2-Chloro-N-((2, 6-dichlorophenyl)carbamoyl)- nicotinamide 4a

  White solid, yield 71.2%, m.p. 185~187 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 7.25 (t, J = 6.4 Hz, 1H, Ph), 7.40~7.42 (m, 1H, Py), 7.42 (d, J = 6.4 Hz, 2H, Ph), 8.19~8.21 (m, 1H, Py), 8.57~8.59 (m, 1H, Py), 9.26 (s, 1H, NH), 10.09 (s, 1H, NH); HRMS (ESI) for C₁₃H₈C_l3N₃O₂ m/z: calculated, 343.9755; found, 343.9768 [M+H]⁺.

  2-Chloro-N-((3-isopropylphenyl)carbamoyl)- nicotinamide 4b

  White solid, yield 65.6%, m.p. 205~208 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.36 (s, 1H, CH₃), 7.16 (d, J = 6.6 Hz, 2H, Ph), 7.38 (d, J = 6.7 Hz, 2H, Ph), 7.42~7.45 (m, 1H, Py), 8.10~8.12 (m, 1H, Py), 8.60~8.62 (m, 1H, Py), 9.39 (s, 1H, NH), 10.37 (s, 1H, NH); HRMS (ESI) for C₁₄H₁₂ClN₃O₂ m/z: calculated, 290.0691; found, 290.0690 [M+H]⁺.

  2-Chloro-N-((2, 6-dimethoxyphenyl)car- bamoyl)nicotinamide 4c

  White solid, yield 60.5 %, m.p. 138~141 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 1.27 (d, J = 5.6 Hz, 6H, CH₃), 2.88~2.95 (m, 1H, CH), 7.04 (d, J = 6.1 Hz, 1H, Ph), 7.25 (d, J = 6.0 Hz, 1H, Ph), 7.29 (s, 1H, Ph), 7.35 (t, J = 3.6 Hz, 1H, Ph), 7.41~7.44 (m, 1H, Py), 8.08~8.10 (m, 1H, Py), 8.59~8.60 (m, 1H, Py), 9.83 (s, 1H, NH), 10.47 (s, 1H, NH); HRMS (ESI) for C₁₆H₁₆ClN₃O₂ m/z: calculated, 318.1004; found, 318.1017 [M+H]⁺.

  2-Chloro-N-((2-methyl-4-nitrophenyl)carbamoyl)- nicotinamide 4d

  White solid, yield 68.7%, m.p. 239~240 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 3.71 (s, 3H, CH₃), 3.84 (s, 3H, CH₃), 6.64~6.66 (m, 1H, Ph), 7.01 (d, J = 6.8 Hz, 1H, Ph), 7.56~7.58 (m, 1H, Py), 7.86 (s, 1H, Ph), 8.12~8.14 (m, 1H, Py), 8.55~8.56 (m, 1H, Py), 10.81 (s, 1H, NH), 11.44 (s, 1H, NH); HRMS (ESI) for C₁₅H₁₄ClN₃O₄ m/z: calculated, 336.0746; found, 336.0751 [M+H]⁺.

  2-Chloro-N-((2, 4-dichlorophenyl)carbamoyl)- nicotinamide 4e

  White solid, yield 63.4%, m.p. 203~206 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.43 (s, 3H, CH₃), 7.58~7.61 (m, 1H, Py), 8.14~8.17 (m, 2H, Ph), 8.21 (s, 1H, Ph), 8.37~8.38 (m, 1H, Py), 8.57~8.59 (m, 1H, Py), 10.73 (s, 1H, NH), 11.74 (s, 1H, NH); HRMS (ESI) for C₁₄H₁₁ClN₄O₄ m/z: calculated, 335.0542; found, 335.0572 [M+H]⁺.

  2-Chloro-N-(phenylcarbamoyl)nicotinamide 4f

  White solid, yield 55.9%, m.p. 207~210 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 7.47~7.49 (m, 1H, Ph), 7.57~7.60 (m, 1H, Py), 7.74~7.76 (m, 1H, Ph), 8.15~8.17 (m, 1H, Py), 8.31 (d, J = 6.4 Hz, 1H, Ph), 8.57~8.58 (m, 1H, Py), 10.95 (s, 1H, NH), 11.70 (s, 1H, NH); HRMS (ESI) for C₁₃H₈Cl₃N₃O₂ m/z: calculated, 343.9755; found, 343.9775 [M+H]⁺.

  2-Chloro-N-((2-chlorophenyl)car- bamoyl)nicotinamide 4g

  White solid, yield 58.9%, m.p. 166~169 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 7.17 (t, J = 5.9 Hz, 1H, Ph), 7.36 (t, J = 6.4 Hz, 2H, Ph), 7.43~7.45 (m, 1H, Py), 7.51 (d, J = 6.4 Hz, 2H, Ph), 8.11~8.13 (m, 1H, Py), 8.60~8.62 (m, 1H, Py), 9.34 (s, 1H, NH), 10.45 (s, 1H, NH); HRMS (ESI) for C₁₃H₁₀ClN₃O₂ m/z: calculated, 276.0534; found, 276.0527[M+H]⁺.

  N-((3-(tert-butyl)phenyl)carbamoyl)- 2-chloronicotinamide 4h

  White solid, yield 48.9%, m.p. 176~179 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 7.09~7.12 (m, 1H, Ph), 7.31~7.32 (m, 1H, Py), 7.44~7.47 (m, 2H, Ph), 8.19~8.21 (m, 1H, Py), 8.24~8.26 (m, 1H, Ph), 8.61~8.62 (m, 1H, Py), 9.24 (s, 1H, NH), 11.00 (s, 1H, NH); HRMS (ESI) for C₁₃H₉Cl₂N₃O₂ m/z: calculated, 310.0145; found, 310.0162[M+H]⁺.

  N-((2, 6-diethylphenyl)carbamoyl)nicotinamide 4i

  Yellow solid, yield 62.3%, m.p. 183~186 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 1.34 (s, 9H, CH₃), 7.34~7.36 (m, 1H, Py), 7.37 (s, 1H, Ph), 7.40~7.43 (m, 3H, Ph), 8.07~8.09 (m, 1H, Py), 8.59~8.61 (m, 1H, Py), 9.90 (s, 1H, NH), 10.41 (s, 1H, NH); HRMS (ESI) for C₁₇H₁₈ClN₃O₂ m/z: calculated, 332.1160; found, 332.1172[M+H]⁺.

  2-Chloro-N-((4-methyl-2-nitrophenyl)car- bamoyl)nicotinamide 4j

  White solid, yield 61.3%, m.p. 193~195 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 1.24 (t, J = 6.1 Hz, 6H, CH₃), 2.64~2.69 (m, 4H, CH₂), 7.16 (d, J = 6.1 Hz, 2H, Ph), 7.28 (t, J = 6.0 Hz, 1H, Ph), 7.32~7.35 (m, 1H, Py), 8.11~8.13 (m, 1H, Py), 8.54~8.55 (m, 1H, Py), 9.33 (s, 1H, NH), 9.76 (s, 1H, NH); ¹³C NMR (CDCl₃, 125 MHz) δ: 14.93, 24.96, 123.51, 125.68, 127.97, 131.91, 133.05, 138.67, 141.73, 146.49, 151.52, 151.90, 167.86; HRMS (ESI) for C₁₇H₁₈ClN₃O₂ m/z: calculated, 332.1160; found, 332.1157[M+H]⁺.

  2-Chloro-N-((2-methyl-3-nitrophenyl)car- bamoyl)nicotinamide 4k

  White solid, yield 57.1%, m.p. 215~218 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.44 (s, 3H, CH₃), 7.47~7.49 (m, 1H, Py), 7.50 (d, J = 5.6 Hz, 1H, Ph), 8.04 (s, 1H, Ph), 8.29~8.31 (m, 1H, Py), 8.50 (d, J = 6.8 Hz, 1H, Ph), 8.61~8.62 (m, 1H, Py), 8.78 (s, 1H, NH), 12.32 (s, 1H, NH); HRMS (ESI) for C₁₄H₁₁ClN₄O₄ m/z: calculated, 335.0542, found, 335.0572[M+H]⁺.

  Methyl 4-(3-(2-chloronicotinoyl)ureido)benzoate 4l

  White solid, yield 49.2%, m.p. 195~198 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.51 (s, 3H, CH₃), 7.39 (t, J = 6.5 Hz, 1H, Ph), 7.47~7.50 (m, 1H, Py), 7.65 (d, J = 6.2 Hz, 1H, Ph), 8.21~8.23 (m, 1H, Py), 8.26 (d, J = 6.1 Hz, 1H, Ph), 8.63~8.64 (m, 1H, Py), 9.00 (s, 1H, NH), 10.64 (s, 1H, NH); HRMS (ESI) for C₁₄H₁₁ClN₄O₄ m/z: calculated, 335.0542, found, 335.0557[M+H]⁺.

  2-Chloro-N-((2, 5-dichlorophenyl)car- bamoyl)nicotinamide 4m

  White solid, yield 65.5%, m.p. 206~209 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 3.94 (s, 3H, CH₃), 7.46~7.48 (m, 1H, Py), 7.61 (d, J = 6.8 Hz, 2H, Ph), 8.05 (d, J = 6.8 Hz, 2H, Ph), 8.13~8.15 (m, 1H, Py), 8.63~8.65 (m, 1H, Py), 9.28 (s, 1H, NH), 10.70 (s, 1H, NH); ¹³C NMR (CDCl₃, 125MHz) δ: 52.42, 119.66, 123.48, 125.07, 130.88, 131.70, 138.78, 142.39, 146.48, 150.62, 151.66, 166.20, 167.73; HRMS (ESI) for C₁₅H₁₂ClN₃O₄ m/z: calculated, 334.0589; found, 334.0610[M+H]⁺.

  2-Chloro-N-((3, 4-dimethylphenyl)car- bamoyl)nicotinamide 4n

  White solid, yield 65.7%, m.p. 168~171 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.40 (s, 3H, CH₃), 7.09~7.12 (m, 1H, Ph), 7.23 (t, J = 6.2 Hz, 2H, Ph), 7.42~7.44 (m, 1H, Py), 7.91 (t, J = 6.4 Hz, 1H, Ph), 8.13~8.15 (m, 1H, Py), 8.60~8.61 (m, 1H, Py), 9.45 (s, 1H, NH), 10.41 (s, 1H, NH); HRMS (ESI) for C₁₄H₁₂ClN₃O₂ m/z: calculated, 290.0691, found, 290.0648[M+H]⁺.

  2-Chloro-N-((4-phenoxyphenyl)car- bamoyl)nicotinamide 4o

  White solid, yield 67.9%, m.p. 189~191 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.26 (s, 3H, CH₃), 2.27 (s, 3H, CH₃), 7.10 (d, J = 6.5 Hz, 1H, Ph), 7.23 (d, J = 6.4 Hz, 1H, Ph), 7.27 (s, 1H, Ph), 7.40~7.42 (m, 1H, Py), 8.09~8.11 (m, 1H, Py), 8.59~8.61 (m, 1H, Py), 9.49 (s, 1H, NH), 10.35 (s, 1H, NH); HRMS (ESI) for C₁₅H₁₄ClN₃O₂ m/z: calculated, 304.0847; found, 304.0837[M+H]⁺.

  2-Chloro-N-((4-ethylphenyl)car- bamoyl)nicotinamide 4p

  White solid, yield 65.0%, m.p. 172~175 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 7.01~7.03 (m, 4H, Ph), 7.13 (t, J = 6.0 Hz, 1H, Ph), 7.34~7.38 (m, 2H, Ph), 7.43~7.45 (m, 1H, Py), 7.47~7.49 (m, 2H, Ph), 8.12~8.14 (m, 1H, Py), 8.59~8.60 (m, 1H, Py), 9.24 (s, 1H, NH), 10.61 (s, 1H, NH); HRMS (ESI) for C₁₉H₁₄ClN₃O₃ m/z: calculated, 368.0796; found, 368.0775 [M+H]⁺.

  2-Chloro-N-(mesitylcarbamoyl)nicotinamide 4q

  White solid, yield 65.7%, m.p. 133~136 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 1.25 (t, J = 6.1 Hz, 3H, CH₃), 2.63~2.68 (m, 2H, CH₂), 7.18 (d, J = 6.7 Hz, 2H, Ph), 7.42 (d, J = 7.2 Hz, 2H, Ph), 7.43~7.45 (m, 1H, Py), 8.11~8.13 (m, 1H, Py), 8.60~8.62 (m, 1H, Py), 9.29 (s, 1H, NH), 10.37 (s, 1H, NH); ¹³C NMR (CDCl₃, 125 MHz) δ: 16.10, 28.02, 120.46, 123.47, 128.68, 131.79, 135.46, 138.79, 139.91, 146.52, 150.66, 151.60, 167.73; HRMS (ESI) for C₁₅H₁₄ClN₃O₂ m/z: calculated, 304.0847; found, 304.0871[M+H]⁺.

  2-Chloro-N-((2, 5-dichlorophenyl)car- bamoyl)nicotinamide 4r

  Yellow solid, yield 54.9%, m.p. 172~175 ℃; ¹H NMR (CDCl₃, 500 MHz), δ: 2.25 (s, 6H, CH₃), 2.31 (s, 3H, CH₃), 6.93 (s, 2H, Ph), 7.34~7.36 (m, 1H, Py), 8.10~8.12 (m, 1H, Py), 8.54~8.55 (m, 1H, Py), 9.40 (s, 1H, NH), 9.68 (s, 1H, NH); ¹³C NMR (CDCl₃, 125 MHz) δ: 18.49, 20.96, 123.50, 128.87, 130.00, 131.74, 135.26, 136.31, 138.70, 146.51, 151.30, 151.50, 167.69; HRMS (ESI) for C₁₆H₁₆ClN₃O₂ m/z: calculated, 318.1004; found, 318.1007[M+H]⁺.

  2.3 Structure determination

  A colorless crystal suitable for X-ray diffraction study was cultivated in the test tube from EtOH by self-volatilization. A crystal with dimensions of 0.28mm × 0.22mm × 0.14mm was mounted on a Bruker APEX-II CCD diffractometer equipped with a graphite-monochromatic MoKα radiation (λ = 0.71073 Å). Intensity data were collected at 296(2) K by using a multi-scan mode in the range of 5.48≤θ≤60.23° with the following index ranges: –10≤h≤10, –20≤k≤20 and –35≤l≤35. A total of 58508 reflections were collected and 4028 were independent (R_int = 0.0532). The crystal structure was solved by direct methods with SHELXS-97^[44] and refined by full-matrix least-squares refinements based on F² with SHELXL-97. All non-hydrogen atoms were refined anisotropically, and all hydrogen atoms were located in the calculated positions and refined with a riding model. The final refinement converged at R = 0.0442 and wR = 0.1033.

  2.4 Fungicidal activity

  Antifungal activities of 2-chloronicotine acyl urea compounds 4a~4r were determined according to our previous work^[36-38].

  2.5 Molecular docking

  Molecular docking was carried out by using DS 2.5. The active sites were generated from the crystal of SDH (PDB code: 2FBW). The detailed method was according to our previous work^[39-42].

  3. RESULTS AND DISCUSSION

  3.1 Synthesis and spectra analysis

  The synthetic route of acyl urea compounds containing nicotinic motif is illustrated in Scheme 1. In the first step, SOCl₂ is used as solvent and reactant. A gas absorption device was used in this reaction in order to avoid environment pollution. During the second synthesis step, the diluted 2-chloronicotinoyl chloride was dropwise added into ammonia water under an ice bath slowly. Then the Py-CONCO was synthesized easily and used without purification. Finally, the target compounds were gotten as white solid under mild condition.

  Scheme 1

  

  Scheme 1.  Synthesis route of 2-chloro-N-(phenylcarbamoyl)nicotinamide compounds

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  These nicotinic acyl urea compounds have two amino groups, and two single peaks can be found around 9.5 and 10.5 ppm respectively. For the pyridine and benzene rings, these protons are in aromatic 6~8 ppm. For ESI-HR-MS, the measured value is according to the theoretical value within 0.003 errors.

  3.2 Crystal structure

  The compound 2-chloro-N-((2, 5-dichlorophenyl)carbamoyl)nicotinamide 4r was further characterized by X-ray diffraction single crystal analysis and is illustrated in Table 1 and Figs. 2 and 3. As shown in Fig. 2, the phenyl ring (C(8)~C(13)) is nearly in the same plane with pyridine ring (C(1), C(2), C(3), C(4), C(5), N(8)), in which the dihedral angle (θ) is 15.1º with plane equation 6.991x – 0.591y + 7.169z = 3.9984 and 6.214x – 0.467y + 13.226z = 8.0225 respectively, and the largest deviations from the least-squares plane are 0.0062 and 0.0091 nm. The torsion angles of C(2)–C(6)–N(1)–N(2), C(6)–N(1)–C(7)–N(2) and N(1)–C(7)–N(2)–C(8) are 176.43(15)°, –1.2(3)° and –175.91(15)° respectively, which showed the acyl urea bridge is nearly in the same plane. The torsion angles of O(1)–C(6)–N(1)–C(7) and O(2)–C(7)–N(1)–C(6) are –5.3(3)° and 3.5(3)° respectively, which indicated the two C=O groups are in the opposite orientations and nearly in the same plane. Compound 4r has four intermolecular hydrogen bonds and one intramolecular hydrogen bond. The parameters of five hydrogen bonds are listed in Table 2. They are linked together by N–H···N, C–H···Cl, C–H···O and N–H···O. From Fig. 1, the intramolecular N(2)– H(2)···O(1) hydrogen bond formed a six-membered ring in molecule 4r. The hydrogen bonding interactions strengthen the integration of the 3D networks.

  Table 1

  

  Table 1.  Selected Bond Lengths (Å) and Bond Angles (°) for the Title Compound 4r

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  Bond	Dist.		Angle	(°)
  Cl(1)–C(1)	1.7302(16)		C(1)–N(1)–C(7)	128.30(13)
  Cl(2)–C(12)	1.7359(19)		C(7)–N(2)–C(8)	123.34(14)
  N(1)–C(6)	1.358(2)		N(8)–C(1)–Cl(1)	114.20(12)
  O(1)–C(6)	1.217(2)		N(8)–C(1)–C(2)	124.59(14)
  O(2)–C(7)	1.208(2)		O(2)–C(7)–N(1)	119.04(14)
  N(1)–C(7)	1.4060(19)		O(2)–C(7)–N(2)	125.53(15)
  N(2)–C(7)	1.352(2)		O(1)–C(6)–N(1)	124.04(14)
  N(2)–C(8)	1.4055(19)		O(1)–C(6)–N(2)	120.1(2)
  N(8)–C(1)	1.321(2)		C(3)–C(2)–C(1)	116.33(14)
  C(1)–C(2)	1.393(2)		C(8)–C(9)–Cl(3)	119.90(13)
  C(2)–C(3)	1.390(2)		C(8)–C(9)–C(10)	121.01(17)

  Figure 2

  

  Figure 2.  Molecular structure of the title compound 4r

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  Figure 3

  

  Figure 3.  Packing of the title compound 4r

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  Table 2

  

  Table 2.  Hydrogen-bond Parameters (Å) of the Title Compound 4r

  DownLoad: CSV

  D–H···A	d(D–H)	d(H···A)	d(D···A)	∠(DHA)
  N(1)–H(1)···N(8)i	0.86	2.23	3.024(19)	153
  N(2)–H(2)···O(1)	0.86	1.98	2.661(18)	135
  C(4)–H(4)···Cl(2)ii	0.93	2.95	3.868(18)	169
  C(5)–H(5)···Cl(1)iii	0.93	2.98	3.855(17)	157
  C(5)–H(5)···O(2)iv	0.93	2.51	3.196(2)	131
  Symmetry transformations used to generate the equivalent atoms: i –1/2+x, 3/2–y, 1–z; ii y, 1+y, z; iii 1/2–x, 1/2+y, z; iv 1/2+x, 3/2–y, 1–z

  3.3 Fungicidal activity

  Antifungal activities of compounds 4a~4r and positive control fluxapyroxad against Sclerotinia sclerotiorum (SS), Phytophthora infestans (PI), Fusarium oxysporum (FO), Phytophthora capsici (PC), Gibberella zeae (GZ), Rhizoctonia solani (RS), Cercospora arachidicola (CA), Alternaria solani (AS), Physalospora piricola (PP) and Botrytis cinerea (BC) were tested at 50 ppm according to published method^[42], with the results shown in Table 3. The antifungal activity results showed some compounds exhibited good inhibitory against B. Cinerea, G. zeae, P. capsici, and P. piricola. Compounds 4e, 4f and 4i still possessed good inhibitory (> 63%) against B. cinerea, which is the same as fluxapyroxad. Most of the title compounds show good inhibitory against P. piricola, such as compounds 4b, 4d, 4h, 4q, 4r and 4s. For the G. zeae, four compounds (4f, 4g, 4i, and 4q) exhibited good activity (57.1%). But the control displayed weak activity (28.6%) against G. zeae. There is no significant fungicidal activity against A. solani, F. oxysporum, P. infestans, R. solani, and C. arachidicola. For F. oxysporum, only compounds 4g (40%) and 4r (40%) exhibited moderate activity, which is the same as the positive control (29.4%). As the same as C. arachidicola, most of these compounds exhibited low activity (< 40%), which are weaker than the positive control (100%). For the P. capsici, compound 4b (70%) exhibited good activity, while the others exhibited low activity.

  Table 3

  

  Table 3.  Fungicidal Activity of the Title Compounds 4a~4r at 50 μg/mL

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  No.	FO	CA	PP	AI	GZ	SS	BC	RS	PI	PC
  4a	6.7	10.0	11.1	5.6	14.3	11.1	18.2	6.2	5.9	40.0
  4b	20.0	10.0	55.6	22.2	28.6	27.8	27.3	12.3	35.3	70.0
  4c	33.3	20.0	27.8	22.2	14.3	33.3	18.2	12.3	17.6	20.0
  4d	13.3	40.0	55.6	11.1	14.3	38.9	27.3	24.7	11.8	10.0
  4e	6.7	20.0	16.7	22.2	28.6	33.3	63.6	30.9	17.6	40.0
  4f	6.7	20.0	33.3	33.3	57.1	50.0	72.7	30.9	11.8	30.0
  4g	40.0	20.0	16.7	22.2	50.0	11.1	18.2	6.2	5.9	40.0
  4h	6.7	20.0	55.6	22.2	35.7	33.3	45.5	24.7	5.9	40.0
  4i	26.7	40.0	44.4	22.2	57.1	77.8	63.6	43.2	11.8	20.0
  4j	33.3	40.0	38.9	33.3	21.4	33.3	27.3	30.9	5.9	20.0
  4k	6.7	6.0	33.3	11.1	28.6	11.1	18.2	12.3	5.9	10.0
  4l	26.7	10.0	33.3	22.2	28.6	11.1	18.2	6.2	5.9	10.0
  4m	6.7	6.0	22.2	22.2	28.6	11.1	9.1	6.2	5.9	6.0
  4n	26.7	6.0	27.8	22.2	21.4	11.1	9.1	8.6	5.9	20.0
  4o	6.7	6.0	5.6	5.6	42.9	33.3	36.4	24.7	5.9	6.0
  4p	13.3	20.0	55.6	22.2	57.1	27.8	45.5	24.7	5.9	6.0
  4q	40.0	40.0	55.6	44.4	21.4	27.8	18.2	37.0	11.8	30.0
  4r	9.1	33.3	50.0	33.3	16.7	36.6	16.7	5.6	14.4	28.6
  FP	29.4	100	63.6	88.9	28.6	96.4	63.6	88.4	27.3	16.7
  CK	0.0	0.0	0.0	0.0	0.0	0.0	0.0	0.0	0.0	0.0

  3.4 Docking study

  In order to study the mode of action of these compounds, the molecular docking was carried out between compound 4i and the enzyme SDH (PDB: 2FBW) using DS 2.5. The docking results indicated that compound 4i can well occupy active site of SDH (Fig. 4). From Fig. 2, a π-cation interaction exists between compound 4i and Arg 43 amino acid residue of SDH with the distance of 3.9 Å. There is another hydrogen bond (distance 2.0 Å) between the C=O of compound 4i and Trp 173 amino acid residue of SDH. From the docking, the pyridine ring and amide group are key active group in this fungicide, which is the same as the lead compound boscalid.

  Figure 4

  

  Figure 4.  Docking mode of compound 4i and SDH

  DownLoad: Full-Size Img PowerPoint
  
  
• 1. [1]

     Dai, H.; Ge, S. S.; Guo, J.; Chen, S.; Huang, M. L.; Yang, J. Y.; Sun, S. Y.; Ling, Y.; Shi, Y. J. Development of novel bis-pyrazole derivatives as antitumor agents with potent apoptosis induction effects and DNA damage. Eur. J. Med. Chem. 2018, 143, 1066–1076. doi: 10.1016/j.ejmech.2017.11.098

  2. [2]

     Liu, X. H.; Wen, Y. H.; Cheng, L.; Xu, T. M.; Wu, N. J. Design, synthesis, pesticidal activities of pyrimidin-4-amine derivatives bearing a 5-(trifluoromethyl)-1, 2, 4-oxadiazole moiety. J. Agric. Food Chem. 2021, 69, 6968−6980. doi: 10.1021/acs.jafc.1c00236

  3. [3]

     Chu, M. J.; Wang, W.; Ren, Z. L.; Liu, H.; Cheng, X.; Mo, K.; Wang, L.; Tang, F.; Lv, X. H. Discovery of novel triazole-containing pyrazole ester derivatives as potential antibacterial agents. Molecules 2019, 24, 1311. doi: 10.3390/molecules24071311

  4. [4]

     Yu, C. S.; Wang, Q.; Bajsa-Hirschel, J.; Cantrell, C.; Duke, S. O.; Liu, X. H. Synthesis, crystal structure, herbicidal activity and SAR study of novel N-(arylmethoxy)-2-chloronicotinamides derived from nicotinic acid. J. Agric. Food Chem. 2021, 69, 6423−6430. doi: 10.1021/acs.jafc.0c07538

  5. [5]

     Shen, Z. H.; Zhai, Z. W.; Sun, Z. H.; Weng, J. Q.; Tan, C. X.; Liu, X. H. Synthesis, crystal structure and biological activity of 2-chloro-5-(((5-(1-methyl-3-(trifluoromethyl)-1H-pyrazol-4-yl)-4-phenyl-4H-1, 2, 4-triazol-3-yl)thio)methyl)thiazole. Chin. J. Struct. Chem. 2017, 36, 1137–1141.

  6. [6]

     Dai, H.; Liu, J. B.; Tao, W. F.; Miao, W. K.; Fang, J. X.; Wang, Q. M. Synthesis, crystal structure and biological activity of novel 1-(pyridin-2-yl)-2-aryloxy-2-(1H-1, 2, 4-triazol-1-yl)ethanones. Chin. J. Org. Chem. 2016, 36, 393–398. doi: 10.6023/cjoc201509011

  7. [7]

     Pessoa-Mahana, D.; Nunez, A.; Espinosa, C.; Mella-Raipan, J.; Pessoa-Mahana, H. Synthesis of a novel series of 4-arylpiperazinyl derivatives linked to a 2-(pyridin-3-yl)-1H-benzimidazole as new delavirdine analogues. J. Brazil Chem. Soc. 2010, 21, 63–70. doi: 10.1590/S0103-50532010000100011

  8. [8]

     Liu, X. H.; Zhai, Z. W.; Xu, X. Y.; Yang, M. Y.; Sun, Z. H.; Weng, J. Q.; Tan, C. X.; Chen, J. Facile and efficient synthesis and herbicidal activity determination of novel 1, 2, 4-triazolo[4, 3-a]pyridin-3(2H)-one derivatives via microwave irradiation. Bioorg. Med. Chem. Lett. 2015, 25, 5524–5528. doi: 10.1016/j.bmcl.2015.10.064

  9. [9]

     Liu, X. H.; Xu, X. Y.; Tan, C. X.; Weng, J. Q.; Xin, J. H.; Chen, J. Synthesis, crystal structure, herbicidal activities and 3D-QSAR study of some novel 1, 2, 4-triazolo[4, 3-a]pyridine derivatives. Pest Manag. Sci. 2015, 71, 292–301. doi: 10.1002/ps.3804

  10. [10]

      Liu, X. H.; Sun, Z. H.; Yang, M. Y.; Tan, C. X.; Weng, J. Q.; Zhang, Y. G.; Ma, Y. Microwave assistant one pot synthesis, crystal structure, antifungal activities and 3D-QSAR of novel 1, 2, 4-triazolo[4, 3-a]pyridines. Chem. Biol. Drug Des. 2014, 84, 342–347. doi: 10.1111/cbdd.12323

  11. [11]

      Hua, X. W.; Liu, W. R.; Su, Y. Y.; Liu, X. H.; Liu, J. B.; Liu, N. N.; Wang, G. Q.; Jiao, X. Q.; Fan, X. Y.; Xue, C. M.; Liu, Y.; Liu, M. Studies on the novel pyridine sulfide containing SDH based heterocyclic amide fungicide. Pest Manag. Sci. 2020, 76, 2368–2378. doi: 10.1002/ps.5773

  12. [12]

      Chaban, T. I.; Ogurtsov, V. V.; Matiychuk, V. S.; Chaban, I. G.; Demchuk, I. L.; Nektegayev, I. A. Synthesis, anti-inflammatory and antioxidant activities of novel 3H-thiazolo[4, 5-b]pyridines. Acta Chem. Sloven. 2019, 66, 103–111.

  13. [13]

      Fan, C.; Wang, Y. B.; Lu, P.; Xue, X. J.; She, J. X. Synthesis and biological evaluation of novel N-[(7-pyridin-4-yl-2, 3-dihydro-benzofuran-2-yl) methyl]-(4-methyl-1, 2, 3-thiadiazole-5-yl) formamide as a potent immunosuppressant agent. Lett. Drug Des. Discov. 2014, 11, 375–379. doi: 10.2174/15701808113106660084

  14. [14]

      Shehab, W. S.; Abdellattif, M. H.; Mouneir, S. M. Heterocyclization of polarized system: synthesis, antioxidant and anti-inflammatory 4-(pyridin-3-yl)-6-(thiophen-2-yl)pyrimidine-2-thiol derivatives. Chem. Cent. J. 2018, 12, 68. doi: 10.1186/s13065-018-0437-y

  15. [15]

      Chavva, K.; Pillalamarri, S.; Banda, V.; Gautham, S.; Gaddamedi, J.; Yedla, P.; Kumar, C. G.; Banda, N. Synthesis and biological evaluation of novel alkyl amide functionalized trifluoromethyl substituted pyrazolo[3, 4-b]pyridine derivatives as potential anticancer agents. Bioorg. Med. Chem. Lett. 2013, 23, 5893–5895. doi: 10.1016/j.bmcl.2013.08.089

  16. [16]

      Wang, H.; Zhai, Z. W.; Shi, Y. X.; Tan, C. X.; Weng, J. Q.; Han, L.; Li, B. J.; Liu, X. H. Novel trifluoromethylpyrazole acyl thiourea derivatives: synthesis, antifungal activity and docking study. Lett. Drug Des. Discov. 2019, 16, 785–791. doi: 10.2174/1570180815666180704103047

  17. [17]

      Wang, H.; Zhai, Z. W.; Shi, Y. X.; Tan, C. X.; Weng, J. Q.; Han, L.; Li, B. J.; Liu, X. H. Novel trifluoromethylpyrazole acyl urea derivatives: synthesis, crystal structure, fungicidal activity and docking study. J. Mol. Struct. 2018, 1171, 631–638. doi: 10.1016/j.molstruc.2018.06.050

  18. [18]

      Sun, N. B.; Zhai, Z. W.; Tong, J. Y.; Cai, P. P.; He, F. Y.; Han, L.; Liu, X. H. Synthesis, crystal structure and fungicidal activity of 3-(difluoromethyl)-1-methyl-N-((2(trifluoromethyl)phenyl)carbamoyl)-1H-pyrazole-4-carboxamide. Chin. J. Struct. Chem. 2019, 38, 706–712.

  19. [19]

      Jin, T.; Zhai, Z. W.; Han, L.; Weng, J. Q.; Tan, C. X.; Liu, X. H. Synthesis, crystal structure, docking and antifungal activity of a new pyrazole acylurea compound. Chin. J. Struct. Chem. 2018, 37, 1259–1264.

  20. [20]

      Zhao, W.; Shen, Z. H.; Xing, J. H.; Xu, T. M.; Peng, W. L.; Liu, X. H. Synthesis and nematocidal activity of novel pyrazole carboxamide derivatives against meloidogyne incognita. Lett. Drug Des. Discov. 2017, 14, 323–329. doi: 10.2174/1570180813666160930164327

  21. [21]

      Cheng, L.; Zhao, W.; Shen, Z. H.; Xu, T. M.; Wu, H. K.; Peng, W. L.; Liu, X. H. Synthesis, nematicidal activity and docking study of novel pyrazole-4-carboxamide derivatives against meloidogyne incognita. Lett. Drug Des. Discov. 2019, 16, 29–35.

  22. [22]

      Wen, Y. H.; Cheng, L.; Xu, T. M.; Liu, X. H.; Wu, N. J. Synthesis, insecticidal activities and DFT study of pyrimidin-4-amine derivatives containing the 1, 2, 4-oxadiazole motif. Front. Chem. Sci. Eng. 2021, doi: 10.1007/s11705-021-2091-5.

  23. [23]

      Zhao, W.; Shen, Z. H.; Xu, T. M.; Peng, W. L.; Liu, X. H. Synthesis and in vivo nematocidal evaluation of novel 3-(trifluoromethyl)-1H-pyrazole-4-carboxamide derivatives. J. Heterocycl. Chem. 2017, 54, 1751–1756. doi: 10.1002/jhet.2753

  24. [24]

      Fang, Y. M.; Zhang, R. R.; Shen, Z. H.; Wu, H. K.; Tan, C. X.; Weng, J. Q.; Xu, T. M.; Liu, X. H. Synthesis, antifungal activity, and SAR study of some new 6-perfluoropropanyl quinoline derivatives. J. Heterocycl. Chem. 2018, 55, 240–245. doi: 10.1002/jhet.3031

  25. [25]

      Cheng, L.; Zhang, R. R.; Wu, H. K.; Xu, T. M.; Liu, X. H. The synthesis of 6-(tert-butyl)-8-fluoro-2, 3-dimethylquinoline carbonate derivatives and their antifungal activity against pyricularia oryzae. Front. Chem. Sci. Eng. 2019, 13, 369–376. doi: 10.1007/s11705-018-1734-7

  26. [26]

      Zhao, W.; Shen, Z. H.; Xing, J. H.; Xu, T. M.; Peng, W. L.; Liu, X. H. Synthesis, characterization, nematocidal activity and docking study of novel pyrazole-4-carboxamide derivatives. Chin. J. Struct. Chem. 2017, 36, 423–428.

  27. [27]

      Liu, X. H.; Zhao, W.; Shen, Z. H.; Xing, J. H.; Xu, T. M.; Peng, W. L. Synthesis, nematocidal activity and SAR study of novel difluoromethylpyrazole carboxamide derivatives containing flexible alkyl chain moieties. Eur. J. Med. Chem. 2017, 125, 881–889. doi: 10.1016/j.ejmech.2016.10.017

  28. [28]

      Zhao, W.; Shen, Z. H.; Xing, J. H.; Xu, T. M.; Peng, W. L.; Liu, X. H. Synthesis and nematocidal activity of novel pyrazole carboxamide derivatives against meloidogyne incognita. Lett. Drug Des. Discov. 2017, 14, 323–329. doi: 10.2174/1570180813666160930164327

  29. [29]

      Liu, X. H.; Zhao, W.; Shen, Z. H.; Xing, J. H.; Yuan, J.; Yang, G.; Xu, T. M.; Peng, W. L. Synthesis, nematocidal activity and docking study of novel chiral 1-(3-chloropyridin-2-yl)-3-(trifluoromethyl)-1H-pyrazole-4-carboxamide derivatives. Bioorg. Med. Chem. Lett. 2016, 26, 3626–3628. doi: 10.1016/j.bmcl.2016.06.004

  30. [30]

      Liu, X. H.; Fang, Y. M.; Xie, F.; Zhang, R. R.; Shen, Z. H.; Tan, C. X.; Weng, J. Q.; Xu, T. M.; Huang, H. Y. Synthesis and in vivo fungicidal activity of some new quinoline derivatives against riceblast. Pest Manag. Sci. 2017, 73, 1900–1907. doi: 10.1002/ps.4556

  31. [31]

      Zhang, P. P.; Wang, Q.; Min, L. J.; Wu, H. K.; Weng, J. Q.; Tan, C. X.; Zhang, Y. G.; Liu, X. H. Synthesis, crystal structure, fungicidal activity and molecular docking of nicotinic acyl urea derivatives. J. Mol. Struct. 2020, 1205, 127485. doi: 10.1016/j.molstruc.2019.127485

  32. [32]

      Sun, N. B.; Shen, Z. H.; Zhai, Z. W.; Han, L.; Weng, J. Q.; Tan, C. X.; Liu, X. H. Design, synthesis, fungicidal activity and docking study of acyl thiourea derivatives containing pyrazole moiety. Chin. J. Org. Chem. 2017, 37, 2705–2710.

  33. [33]

      Min, L. J.; Wang, Q.; Tan, C. X.; Weng, J. Q.; Liu, X. H. Synthesis, crystal structure and fungicidal activity of 2-chloro-N-(o-tolylcarbamoyl)nicotinamide. Chin. J. Struct. Chem. 2020, 39, 452–458.

  34. [34]

      Liu, X. H.; Wang, Q.; Sun, Z. H.; Wedge, D. E.; Becnel, J. J.; Estep, A. S.; Tan, C. X.; Weng, J. Q. Synthesis and insecticidal activity of novel pyrimidine derivatives containing urea pharmacophore against Aedes aegypti. Pest Manag. Sci. 2017, 73, 953–959. doi: 10.1002/ps.4370

  35. [35]

      Zhang, L. J.; Yang, M. Y.; Sun, Z. H.; Tan, C. X.; Weng, J. Q.; Wu, H. K.; Liu, X. H. Synthesis and antifungal activity of 1, 3, 4-thiadiazole derivatives containing pyridine group drug. Lett. Drug Des. Discov. 2014, 11, 1107–1111. doi: 10.2174/1570180811666140610212731

  36. [36]

      Min, L. J.; Zhai, Z. W.; Shi, Y. X.; Han, L.; Tan, C. X.; Weng, J. Q.; Li, B. J.; Zhang, Y. G.; Liu, X. H. Synthesis and biological activity of acyl thiourea containing difluoromethyl pyrazole motif. Phosphorus Sulfur Silicon Relat. Elem. 2020, 195, 22–28. doi: 10.1080/10426507.2019.1633530

  37. [37]

      Yang, M. Y.; Zhai, Z. W.; Sun, Z. H.; Yu, S. J.; Liu, X. H.; Weng, J. Q.; Tan, C. X.; Zhao, W. G. A facile one-pot synthesis of novel 1, 2, 4-triazolo[4, 3-a]pyridine derivatives containing the trifluoromethyl moiety using microwave irradiation. J. Chem. Res. 2015, 9, 521–523.

  38. [38]

      Zhao, W.; Shen, Z. H.; Xing, J. H.; Yang, G.; Xu, T. M.; Peng, W. L.; Liu, X. H. Synthesis and nematocidal activity of novel 1-(3-chloropyridin-2-yl)-3-(trifluoromethyl)-1H-pyrazole-4-carboxamide derivatives. Chem. Pap. 2017, 71, 921–928. doi: 10.1007/s11696-016-0012-8

  39. [39]

      Zhao, W.; Xing, J. H.; Xu, T. M.; Peng, W. L.; Liu, X. H. Synthesis and in vivo nematocidal evaluation of novel 3-(trifluoromethyl)-1H-pyrazole-4-carboxamide derivatives. Front. Chem. Sci. Eng. 2017, 11, 363–368. doi: 10.1007/s11705-016-1595-x

  40. [40]

      Liu, X. H.; Qiao, L.; Zhai, Z. W.; Cai, P. P.; Cantrell, C. L.; Tan, C. X.; Weng, J. Q.; Han, L.; Wu, H. K. Novel 4-pyrazole carboxamide derivatives containing flexible chain motif: design, synthesis and antifungal activity. Pest Manag. Sci. 2019, 75, 2892–2900. doi: 10.1002/ps.5463

  41. [41]

      Fu, Q.; Cai, P. P.; Cheng, L.; Zhong, L. K.; Tan, C. X.; Shen, Z. H.; Han, L.; Liu, X. H. Synthesis and herbicidal activity of novel pyrazole aromatic ketone analogs as HPPD inhibitor. Pest Manag. Sci. 2020, 76, 868–879. doi: 10.1002/ps.5591

  42. [42]

      Liu, X. H.; Yu, W.; Min, L. J.; Wedge, D. E.; Tan, C. X.; Weng, J. Q.; Wu, H. K.; Cantrell, C. L.; Bajsa-Hischel, J.; Hua, X. W.; Duke, S. O. Synthesis and pesticidal activities of new quinoxalines. J. Agric. Food Chem. 2020, 68, 7324–7332. doi: 10.1021/acs.jafc.0c01042

• 

  Figure 1  Design method of nicotinic acyl urea derivatives

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  Scheme 1  Synthesis route of 2-chloro-N-(phenylcarbamoyl)nicotinamide compounds

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  Figure 2  Molecular structure of the title compound 4r

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  Figure 3  Packing of the title compound 4r

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  Figure 4  Docking mode of compound 4i and SDH

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  Table 1.  Selected Bond Lengths (Å) and Bond Angles (°) for the Title Compound 4r

  Bond	Dist.		Angle	(°)
  Cl(1)–C(1)	1.7302(16)		C(1)–N(1)–C(7)	128.30(13)
  Cl(2)–C(12)	1.7359(19)		C(7)–N(2)–C(8)	123.34(14)
  N(1)–C(6)	1.358(2)		N(8)–C(1)–Cl(1)	114.20(12)
  O(1)–C(6)	1.217(2)		N(8)–C(1)–C(2)	124.59(14)
  O(2)–C(7)	1.208(2)		O(2)–C(7)–N(1)	119.04(14)
  N(1)–C(7)	1.4060(19)		O(2)–C(7)–N(2)	125.53(15)
  N(2)–C(7)	1.352(2)		O(1)–C(6)–N(1)	124.04(14)
  N(2)–C(8)	1.4055(19)		O(1)–C(6)–N(2)	120.1(2)
  N(8)–C(1)	1.321(2)		C(3)–C(2)–C(1)	116.33(14)
  C(1)–C(2)	1.393(2)		C(8)–C(9)–Cl(3)	119.90(13)
  C(2)–C(3)	1.390(2)		C(8)–C(9)–C(10)	121.01(17)

  下载: 导出CSV

  Table 2.  Hydrogen-bond Parameters (Å) of the Title Compound 4r

  D–H···A	d(D–H)	d(H···A)	d(D···A)	∠(DHA)
  N(1)–H(1)···N(8)i	0.86	2.23	3.024(19)	153
  N(2)–H(2)···O(1)	0.86	1.98	2.661(18)	135
  C(4)–H(4)···Cl(2)ii	0.93	2.95	3.868(18)	169
  C(5)–H(5)···Cl(1)iii	0.93	2.98	3.855(17)	157
  C(5)–H(5)···O(2)iv	0.93	2.51	3.196(2)	131
  Symmetry transformations used to generate the equivalent atoms: i –1/2+x, 3/2–y, 1–z; ii y, 1+y, z; iii 1/2–x, 1/2+y, z; iv 1/2+x, 3/2–y, 1–z

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  Table 3.  Fungicidal Activity of the Title Compounds 4a~4r at 50 μg/mL

  No.	FO	CA	PP	AI	GZ	SS	BC	RS	PI	PC
  4a	6.7	10.0	11.1	5.6	14.3	11.1	18.2	6.2	5.9	40.0
  4b	20.0	10.0	55.6	22.2	28.6	27.8	27.3	12.3	35.3	70.0
  4c	33.3	20.0	27.8	22.2	14.3	33.3	18.2	12.3	17.6	20.0
  4d	13.3	40.0	55.6	11.1	14.3	38.9	27.3	24.7	11.8	10.0
  4e	6.7	20.0	16.7	22.2	28.6	33.3	63.6	30.9	17.6	40.0
  4f	6.7	20.0	33.3	33.3	57.1	50.0	72.7	30.9	11.8	30.0
  4g	40.0	20.0	16.7	22.2	50.0	11.1	18.2	6.2	5.9	40.0
  4h	6.7	20.0	55.6	22.2	35.7	33.3	45.5	24.7	5.9	40.0
  4i	26.7	40.0	44.4	22.2	57.1	77.8	63.6	43.2	11.8	20.0
  4j	33.3	40.0	38.9	33.3	21.4	33.3	27.3	30.9	5.9	20.0
  4k	6.7	6.0	33.3	11.1	28.6	11.1	18.2	12.3	5.9	10.0
  4l	26.7	10.0	33.3	22.2	28.6	11.1	18.2	6.2	5.9	10.0
  4m	6.7	6.0	22.2	22.2	28.6	11.1	9.1	6.2	5.9	6.0
  4n	26.7	6.0	27.8	22.2	21.4	11.1	9.1	8.6	5.9	20.0
  4o	6.7	6.0	5.6	5.6	42.9	33.3	36.4	24.7	5.9	6.0
  4p	13.3	20.0	55.6	22.2	57.1	27.8	45.5	24.7	5.9	6.0
  4q	40.0	40.0	55.6	44.4	21.4	27.8	18.2	37.0	11.8	30.0
  4r	9.1	33.3	50.0	33.3	16.7	36.6	16.7	5.6	14.4	28.6
  FP	29.4	100	63.6	88.9	28.6	96.4	63.6	88.4	27.3	16.7
  CK	0.0	0.0	0.0	0.0	0.0	0.0	0.0	0.0	0.0	0.0

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•