# 2,4-Dichlorophenoxyacetic Acid Derived Schiff Base and Its Lanthanide(III) Complexes: Synthesis, Characterization, Spectroscopic Studies, and Plant Growth Activity

Authors: Ganesh N. Naik; Raghavendra P. Bakale; Aishakhanam H. Pathan; Shashikala G. Ligade; Shreenivas A. Desai; Kalagouda B. Gudasi
Journal: Journal of Chemistry
 (2013)

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## Abstract

2,4-Dichlorophenoxyacetic acid derived Schiff base (HL) and its lanthanide [La(III), Pr(III), Nd(III), Sm(III), Eu(III), Gd(III), Dy(III), Y(III)] complexes were synthesized and characterized by various spectroscopic (1H, 13C, DEPT and 2D HMQC NMR, FT-IR, UV-Vis, and mass) techniques and other analytical methods. HL exhibits “E” and “Z” isomerism and was confirmed by variable temperature 1H NMR studies. The spectral and analytical data reveals the bidentate coordination of HL to lanthanide(III) ion, through carboxylic acid group via deprotonation. Fluorescence spectrum of europium complex shows bands at 578, 592, and 612 nm assignable to D05→F07, D05→F17, and D05→F27, respectively. Auxin activity of HL and lanthanum(III) complex on wheat seeds (Triticum durum) was measured at different concentrations. The percentage germination, root length, and shoot length were recorded. An enhancement in the plant growth activity of the ligand was observed on complexation and the best activity was observed at 10−6 M concentration.


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## Body

## 1. Introduction

Rare earth elements are being used in agriculture as micronutrients and fertilizers [1–4]. These are found to have nitrogen fixing capacity, to enhance activity of hydrolytic enzymes, to promote seed germination, to strengthen photosynthetic rate, and to reduce water loss in plants [5, 6]. Recent experiments indicate that lanthanide salts could accelerate the seed germination, increases chlorophyll content, and improve root growth [7, 8]. These elements also found to have significant effect on the physiological and biochemical reactions in plant growth and development [9, 10]. The effect of lanthanide(III) complexes on the growth of plants has been an important topic in agricultural field. There are only few reports which explain the auxin effect of lanthanide complexes on plant growth [11, 12].Indole-3 acetic acid, naphthalene acetic acid, and 2,4-dichlorophenoxyacetic acid are the well known plant growth regulating hormones, widely used in agricultural field [13, 14].  The derivatives of these hormones were also found to have plant growth regulating and other biological activities [13, 15, 16].Literature survey reveals that chelation of lanthanide metal ions with plant growth promoting auxins synergistically increases the plant growth [12]. Keeping this in view, we have synthesized 2,4-dichlorophenoxy acetic acid derived Schiff base and its lanthanide(III) complexes and screened for their auxin activity on wheat seeds. The work is focused on the synthesis, characterization, spectroscopic investigation, thermal, fluorescent behavior, and plant growth promoting activity of a series of rare earth complexes of 2,4-dichlorophenoxy acetic acid derived Schiff base (HL).


## 2. Experimental

### 2.1. Instrumentation and Materials

Elemental analyses (C, H, and N) were performed using Leco Model TruSpec CHNS analyzer. The percentage of metal content was determined according to the literature procedure [17] with xylenol orange as the indicator and EDTA as the chelating agent. SHIMADZU LCMS 2010A spectrometer is used to know the mass. Molar conductivities in DMSO (10−3M) at room temperature were measured using an Elico conductivity bridge having platinum electrode. Melting points were determined in an open capillary on a Gallenkamp melting point apparatus and are uncorrected. IR spectra were recorded in a KBr matrix using an Impact-410 Nicolet (USA) FT-IR spectrometer in 4000–400 cm−1 range. The 1H, 13C, DEPT, 2D HMQC NMR, and variable temperature 1H NMR were recorded in DMSO-d6 solvent on BRUKER AV-500 MHz High Resolution Multinuclear FT-NMR spectrometer using SiMe4 as an internal standard at δ = 0 ppm. Thermogravimetry (TG) and differential thermal analysis (DTA) were run in nitrogen atmosphere in a temperature range of 20°C to 1000°C at a heating rate of 10°C min−1 using a Perkin-Elmer (Pyris Diamond) analyzer. UV-Visible spectra were recorded on a CARY 50 Bio UV-Visible spectrophotometer in 200–1100 nm range in DMSO solvent. Fluorescence spectrum was measured on F-7000 FL spectrophotometer. Plant growth activity was performed in a dual chamber seed germinator at 25°C temperature and 70±5% humidity with proper illumination.2,4-Dichlorophenoxyacetic acid and 2-formyl phenoxyacetic acid were obtained from Sigma Aldrich. Lanthanide(III) oxides were procured from Indian Rare Earths Ltd, India. Other chemicals were obtained from s.d. Fine Chemicals, India, and used as received. Solvents were purified by standard methods [18]. Lanthanide nitrates were prepared by dissolving the corresponding oxide (99.99%,) in 50% HNO3 followed by evaporation of the excess acid. All the compounds were routinely checked by thin-layer chromatography (TLC) on aluminum-backed silica gel plates. The wheat seeds (Triticum durum) DWR-2006 were collected from University of Agricultural Sciences (UAS), Dharwad, India.


### 2.2. Synthesis

#### 2.2.1. Synthesis of 2-{[2-(2, 4-Dichloro-phenoxy)-Acetyl]-Hydrazonomethyl}-Phenoxy)-Acetic Acid (HL)

A mixture of  2,4-dichlorophenoxyacetic acid (1) (12 g, 54.28 mmol) and concentrated H2SO4 (4 mL) in dry methanol (50 mL) was refluxed for 15 hrs to yield the 2,4-dichlorophenoxy acetic acid methyl ester (2). Product was separated and purified. To the methanolic solution of 2, (8 g, 34.03 mmol), hydrazine hydrate (3 g, 44.08 mmol) was added slowly and stirred for 30 min. The 2,4-dichlorophenoxyacetic acid hydrazide separated (3) was filtered and washed with methanol. The hydrazide (3) (7 gm, 29.77 mmol) was stirred with 2-formyl phenoxy acetic acid (4) (5.36 g, 29.77 mmol) in THF (75 mL) for 30 min and the product formed was filtered and washed thoroughly with THF to obtain 2-{[2-(2,4-dichloro-phenoxy)-acetyl]-hydrazonomethyl}-phenoxy)-acetic acid (HL) (5). Yield: 90%, M.P: 233–235°C.1H NMR (DMSO-d6, ppm): 4.82, 4.90 (s, 2H, C2H/C2′H), 4.80, 5.31 (s, 2H, C11H/C11′H), 6.97, 7.01 (d, 1H, J = 9.4 Hz, C4H/C4′H), 7.03, 7.08 (d, 1H, J = 6.4 Hz, C17H/C17′H), 7.11, 7.17 (d, 1H, J = 6.4 Hz, C16H/C16′H), 7.32–7.41 (m, 2H, C14H/C14′H and C5H/C5′H), 7.57, 7.61 (d, 1H, J = 5 Hz, C7H/C7′H), 7.83–7.90 (m, 1H, C6H/C6′H), 8.42, 8.65 (s, 1H, C9H/C9′H), 11.72, 11.80 (1H, NH/N′H), 13.16 (s, 1H, OH).13C NMR (DMSO-d6, ppm): 64.62, 64.75 (C2/C2′), 64.67, 66.91 (C11/C11′), 112.47, 112.84 (C4/C4′), 114.97, 115.12 (C17/C17′), 116.15, 117.13 (C8/C8′), 121.01, 121.03 (C16/C16′), 122.02, 122.20 (C13/C13′), 124.38, 124.94 (C6/C6′), 125.58, 125.77 (C15/C15′), 127.75, 127.96 (C5/C5′), 129.12, 129.30 (C7/C7′), 131.15, 131.44 (C14/C14′), 139.00, 143.00 (C9/C9′), 152.54, 152.73 (C12/C12′), 156.08, 156.20 (C3/C3′), 163.28, 168.12 (C10/C10′), 169.88, 169.94 (C1/C1′).


#### 2.2.2. General Procedure for the Synthesis of Lanthanide Complexes

The complexes were prepared by treating HL with freshly prepared lanthanide(III) nitrates in presence of triethyl amine according to literature procedure [19] with slight modifications. The mixture of HL (0.5 gm, 1.26 mmol) and triethyl amine (0.14 g, 1.38 mmol) was taken in THF (50 mL) and stirred till HL completely dissolves. To this solution, freshly prepared Ln(NO3)3 (0.42 mmol) (Ln = La, Pr, Nd, Sm, Eu, Gd, Dy, Y) was added slowly under stirring and was further refluxed for 2 hrs. Solvent was evaporated under reduced pressure. The solid obtained was washed with water and THF several times. The product was dried under vacuum. Yield: 60–70%.1H NMR of lanthanum(III) complex (DMSO-d6, ppm): 4.70 (s, 2H, C2H), 5.28 (s, 2H, C11H), 6.92 (d, 1H, J = 9.2 Hz, C4H), 7.10 (d, 1H, J = 6.3 Hz, C16H), 7.25 (d, 1H,  J = 6.3 Hz, C17H), 7.39 (m, 1H, C5H), 7.40 (s, 1H, C14H), 7.56 (d, 1H, J = 9.2 Hz, C7H), 7.86–7.90 (m, 1H, C6H), 8.41 (s, 1H, C9H), 11.64 (1H, NH).13C NMR lanthanum(III) complex (DMSO-d6, ppm): 66.91 (C11), 67.55 (C2), 112.47 (C4), 115.28 (C17), 117.13 (C8), 122.13 (C13), 122.25 (C16), 124.30 (C15), 126.92 (C6), 127.96 (C5), 129.12 (C7), 131.90 (C14), 138.50 (C9), 153.00 (C12), 156.08, (C3), 164.30 (C10), 176.46 (C1).


### 2.3. Protocol for the Plant Growth Activity on Wheat Seeds

DWR-2006 (Triticum durum), a local variety of wheat seeds developed at University of Agricultural Sciences, Dharwad, India, was selected to investigate the growth activities of the synthesized compounds. The solutions of HL, lanthanum(III) complex, La(NO3)3, and 2,4-dichlorophenoxyacetic acid were prepared by dissolving them in minimum quantity of DMSO (~1 mL) and further diluted with distilled water to obtain the solutions of 1 × 10−5M, 1 × 10−6M and, 1×  10−7M concentrations. Seed germination experiments were carried out according to the literature method [20] with slight modifications. Healthy wheat seeds were selected and washed with distilled water before soaking in test solutions. Hundred seeds were soaked in test solutions of particular concentration for 3 minutes and then arranged on the moistened specially prepared germination paper placed on polythene sheet. One more moistened germination paper was placed over the seeds and loosely rolled. Each roll was labeled clearly and kept upright in the seed germinator and maintained at 25°C temperature and 70±5% humidity, with proper illumination. The conditions were maintained for eight days. After eight days, percentage germination was calculated. Further, 10 seeds were randomly selected from each roll for root and shoot length measurements. The experiments were performed in triplicate.


#### 2.3.1. Statistical Analysis

The results are evaluated with SPSS (Statistical Package for Social Science, Windows, version 17.0) packed program. One way ANOVA with Duncan statistical analysis with significance atP<0.05 was used for data analysis.


## 2.1. Instrumentation and Materials

Elemental analyses (C, H, and N) were performed using Leco Model TruSpec CHNS analyzer. The percentage of metal content was determined according to the literature procedure [17] with xylenol orange as the indicator and EDTA as the chelating agent. SHIMADZU LCMS 2010A spectrometer is used to know the mass. Molar conductivities in DMSO (10−3M) at room temperature were measured using an Elico conductivity bridge having platinum electrode. Melting points were determined in an open capillary on a Gallenkamp melting point apparatus and are uncorrected. IR spectra were recorded in a KBr matrix using an Impact-410 Nicolet (USA) FT-IR spectrometer in 4000–400 cm−1 range. The 1H, 13C, DEPT, 2D HMQC NMR, and variable temperature 1H NMR were recorded in DMSO-d6 solvent on BRUKER AV-500 MHz High Resolution Multinuclear FT-NMR spectrometer using SiMe4 as an internal standard at δ = 0 ppm. Thermogravimetry (TG) and differential thermal analysis (DTA) were run in nitrogen atmosphere in a temperature range of 20°C to 1000°C at a heating rate of 10°C min−1 using a Perkin-Elmer (Pyris Diamond) analyzer. UV-Visible spectra were recorded on a CARY 50 Bio UV-Visible spectrophotometer in 200–1100 nm range in DMSO solvent. Fluorescence spectrum was measured on F-7000 FL spectrophotometer. Plant growth activity was performed in a dual chamber seed germinator at 25°C temperature and 70±5% humidity with proper illumination.2,4-Dichlorophenoxyacetic acid and 2-formyl phenoxyacetic acid were obtained from Sigma Aldrich. Lanthanide(III) oxides were procured from Indian Rare Earths Ltd, India. Other chemicals were obtained from s.d. Fine Chemicals, India, and used as received. Solvents were purified by standard methods [18]. Lanthanide nitrates were prepared by dissolving the corresponding oxide (99.99%,) in 50% HNO3 followed by evaporation of the excess acid. All the compounds were routinely checked by thin-layer chromatography (TLC) on aluminum-backed silica gel plates. The wheat seeds (Triticum durum) DWR-2006 were collected from University of Agricultural Sciences (UAS), Dharwad, India.


## 2.2. Synthesis

### 2.2.1. Synthesis of 2-{[2-(2, 4-Dichloro-phenoxy)-Acetyl]-Hydrazonomethyl}-Phenoxy)-Acetic Acid (HL)

A mixture of  2,4-dichlorophenoxyacetic acid (1) (12 g, 54.28 mmol) and concentrated H2SO4 (4 mL) in dry methanol (50 mL) was refluxed for 15 hrs to yield the 2,4-dichlorophenoxy acetic acid methyl ester (2). Product was separated and purified. To the methanolic solution of 2, (8 g, 34.03 mmol), hydrazine hydrate (3 g, 44.08 mmol) was added slowly and stirred for 30 min. The 2,4-dichlorophenoxyacetic acid hydrazide separated (3) was filtered and washed with methanol. The hydrazide (3) (7 gm, 29.77 mmol) was stirred with 2-formyl phenoxy acetic acid (4) (5.36 g, 29.77 mmol) in THF (75 mL) for 30 min and the product formed was filtered and washed thoroughly with THF to obtain 2-{[2-(2,4-dichloro-phenoxy)-acetyl]-hydrazonomethyl}-phenoxy)-acetic acid (HL) (5). Yield: 90%, M.P: 233–235°C.1H NMR (DMSO-d6, ppm): 4.82, 4.90 (s, 2H, C2H/C2′H), 4.80, 5.31 (s, 2H, C11H/C11′H), 6.97, 7.01 (d, 1H, J = 9.4 Hz, C4H/C4′H), 7.03, 7.08 (d, 1H, J = 6.4 Hz, C17H/C17′H), 7.11, 7.17 (d, 1H, J = 6.4 Hz, C16H/C16′H), 7.32–7.41 (m, 2H, C14H/C14′H and C5H/C5′H), 7.57, 7.61 (d, 1H, J = 5 Hz, C7H/C7′H), 7.83–7.90 (m, 1H, C6H/C6′H), 8.42, 8.65 (s, 1H, C9H/C9′H), 11.72, 11.80 (1H, NH/N′H), 13.16 (s, 1H, OH).13C NMR (DMSO-d6, ppm): 64.62, 64.75 (C2/C2′), 64.67, 66.91 (C11/C11′), 112.47, 112.84 (C4/C4′), 114.97, 115.12 (C17/C17′), 116.15, 117.13 (C8/C8′), 121.01, 121.03 (C16/C16′), 122.02, 122.20 (C13/C13′), 124.38, 124.94 (C6/C6′), 125.58, 125.77 (C15/C15′), 127.75, 127.96 (C5/C5′), 129.12, 129.30 (C7/C7′), 131.15, 131.44 (C14/C14′), 139.00, 143.00 (C9/C9′), 152.54, 152.73 (C12/C12′), 156.08, 156.20 (C3/C3′), 163.28, 168.12 (C10/C10′), 169.88, 169.94 (C1/C1′).


### 2.2.2. General Procedure for the Synthesis of Lanthanide Complexes

The complexes were prepared by treating HL with freshly prepared lanthanide(III) nitrates in presence of triethyl amine according to literature procedure [19] with slight modifications. The mixture of HL (0.5 gm, 1.26 mmol) and triethyl amine (0.14 g, 1.38 mmol) was taken in THF (50 mL) and stirred till HL completely dissolves. To this solution, freshly prepared Ln(NO3)3 (0.42 mmol) (Ln = La, Pr, Nd, Sm, Eu, Gd, Dy, Y) was added slowly under stirring and was further refluxed for 2 hrs. Solvent was evaporated under reduced pressure. The solid obtained was washed with water and THF several times. The product was dried under vacuum. Yield: 60–70%.1H NMR of lanthanum(III) complex (DMSO-d6, ppm): 4.70 (s, 2H, C2H), 5.28 (s, 2H, C11H), 6.92 (d, 1H, J = 9.2 Hz, C4H), 7.10 (d, 1H, J = 6.3 Hz, C16H), 7.25 (d, 1H,  J = 6.3 Hz, C17H), 7.39 (m, 1H, C5H), 7.40 (s, 1H, C14H), 7.56 (d, 1H, J = 9.2 Hz, C7H), 7.86–7.90 (m, 1H, C6H), 8.41 (s, 1H, C9H), 11.64 (1H, NH).13C NMR lanthanum(III) complex (DMSO-d6, ppm): 66.91 (C11), 67.55 (C2), 112.47 (C4), 115.28 (C17), 117.13 (C8), 122.13 (C13), 122.25 (C16), 124.30 (C15), 126.92 (C6), 127.96 (C5), 129.12 (C7), 131.90 (C14), 138.50 (C9), 153.00 (C12), 156.08, (C3), 164.30 (C10), 176.46 (C1).


## 2.2.1. Synthesis of 2-{[2-(2, 4-Dichloro-phenoxy)-Acetyl]-Hydrazonomethyl}-Phenoxy)-Acetic Acid (HL)

A mixture of  2,4-dichlorophenoxyacetic acid (1) (12 g, 54.28 mmol) and concentrated H2SO4 (4 mL) in dry methanol (50 mL) was refluxed for 15 hrs to yield the 2,4-dichlorophenoxy acetic acid methyl ester (2). Product was separated and purified. To the methanolic solution of 2, (8 g, 34.03 mmol), hydrazine hydrate (3 g, 44.08 mmol) was added slowly and stirred for 30 min. The 2,4-dichlorophenoxyacetic acid hydrazide separated (3) was filtered and washed with methanol. The hydrazide (3) (7 gm, 29.77 mmol) was stirred with 2-formyl phenoxy acetic acid (4) (5.36 g, 29.77 mmol) in THF (75 mL) for 30 min and the product formed was filtered and washed thoroughly with THF to obtain 2-{[2-(2,4-dichloro-phenoxy)-acetyl]-hydrazonomethyl}-phenoxy)-acetic acid (HL) (5). Yield: 90%, M.P: 233–235°C.1H NMR (DMSO-d6, ppm): 4.82, 4.90 (s, 2H, C2H/C2′H), 4.80, 5.31 (s, 2H, C11H/C11′H), 6.97, 7.01 (d, 1H, J = 9.4 Hz, C4H/C4′H), 7.03, 7.08 (d, 1H, J = 6.4 Hz, C17H/C17′H), 7.11, 7.17 (d, 1H, J = 6.4 Hz, C16H/C16′H), 7.32–7.41 (m, 2H, C14H/C14′H and C5H/C5′H), 7.57, 7.61 (d, 1H, J = 5 Hz, C7H/C7′H), 7.83–7.90 (m, 1H, C6H/C6′H), 8.42, 8.65 (s, 1H, C9H/C9′H), 11.72, 11.80 (1H, NH/N′H), 13.16 (s, 1H, OH).13C NMR (DMSO-d6, ppm): 64.62, 64.75 (C2/C2′), 64.67, 66.91 (C11/C11′), 112.47, 112.84 (C4/C4′), 114.97, 115.12 (C17/C17′), 116.15, 117.13 (C8/C8′), 121.01, 121.03 (C16/C16′), 122.02, 122.20 (C13/C13′), 124.38, 124.94 (C6/C6′), 125.58, 125.77 (C15/C15′), 127.75, 127.96 (C5/C5′), 129.12, 129.30 (C7/C7′), 131.15, 131.44 (C14/C14′), 139.00, 143.00 (C9/C9′), 152.54, 152.73 (C12/C12′), 156.08, 156.20 (C3/C3′), 163.28, 168.12 (C10/C10′), 169.88, 169.94 (C1/C1′).


## 2.2.2. General Procedure for the Synthesis of Lanthanide Complexes

The complexes were prepared by treating HL with freshly prepared lanthanide(III) nitrates in presence of triethyl amine according to literature procedure [19] with slight modifications. The mixture of HL (0.5 gm, 1.26 mmol) and triethyl amine (0.14 g, 1.38 mmol) was taken in THF (50 mL) and stirred till HL completely dissolves. To this solution, freshly prepared Ln(NO3)3 (0.42 mmol) (Ln = La, Pr, Nd, Sm, Eu, Gd, Dy, Y) was added slowly under stirring and was further refluxed for 2 hrs. Solvent was evaporated under reduced pressure. The solid obtained was washed with water and THF several times. The product was dried under vacuum. Yield: 60–70%.1H NMR of lanthanum(III) complex (DMSO-d6, ppm): 4.70 (s, 2H, C2H), 5.28 (s, 2H, C11H), 6.92 (d, 1H, J = 9.2 Hz, C4H), 7.10 (d, 1H, J = 6.3 Hz, C16H), 7.25 (d, 1H,  J = 6.3 Hz, C17H), 7.39 (m, 1H, C5H), 7.40 (s, 1H, C14H), 7.56 (d, 1H, J = 9.2 Hz, C7H), 7.86–7.90 (m, 1H, C6H), 8.41 (s, 1H, C9H), 11.64 (1H, NH).13C NMR lanthanum(III) complex (DMSO-d6, ppm): 66.91 (C11), 67.55 (C2), 112.47 (C4), 115.28 (C17), 117.13 (C8), 122.13 (C13), 122.25 (C16), 124.30 (C15), 126.92 (C6), 127.96 (C5), 129.12 (C7), 131.90 (C14), 138.50 (C9), 153.00 (C12), 156.08, (C3), 164.30 (C10), 176.46 (C1).


## 2.3. Protocol for the Plant Growth Activity on Wheat Seeds

DWR-2006 (Triticum durum), a local variety of wheat seeds developed at University of Agricultural Sciences, Dharwad, India, was selected to investigate the growth activities of the synthesized compounds. The solutions of HL, lanthanum(III) complex, La(NO3)3, and 2,4-dichlorophenoxyacetic acid were prepared by dissolving them in minimum quantity of DMSO (~1 mL) and further diluted with distilled water to obtain the solutions of 1 × 10−5M, 1 × 10−6M and, 1×  10−7M concentrations. Seed germination experiments were carried out according to the literature method [20] with slight modifications. Healthy wheat seeds were selected and washed with distilled water before soaking in test solutions. Hundred seeds were soaked in test solutions of particular concentration for 3 minutes and then arranged on the moistened specially prepared germination paper placed on polythene sheet. One more moistened germination paper was placed over the seeds and loosely rolled. Each roll was labeled clearly and kept upright in the seed germinator and maintained at 25°C temperature and 70±5% humidity, with proper illumination. The conditions were maintained for eight days. After eight days, percentage germination was calculated. Further, 10 seeds were randomly selected from each roll for root and shoot length measurements. The experiments were performed in triplicate.


### 2.3.1. Statistical Analysis

The results are evaluated with SPSS (Statistical Package for Social Science, Windows, version 17.0) packed program. One way ANOVA with Duncan statistical analysis with significance atP<0.05 was used for data analysis.


## 2.3.1. Statistical Analysis

The results are evaluated with SPSS (Statistical Package for Social Science, Windows, version 17.0) packed program. One way ANOVA with Duncan statistical analysis with significance atP<0.05 was used for data analysis.


## 3. Results and Discussions

The Schiff base (HL) was prepared as shown in Scheme1. The structure of HL was confirmed by IR, NMR, and mass spectral analysis. All the complexes were obtained in moderate to good yields (60–70%) by reacting HL and lanthanide(III) nitrates in THF in 3 : 1 (ligand to metal) molar ratio. The complexes are soluble in DMSO and DMF. The analytical results given in Table 1 agree with the suggested formula of complexes. Elemental analysis, FT-IR, 1H, 13C, 2D HMQC NMR, TGA/DSC, and UV-Visible spectroscopy were used to characterize the complexes. The elemental analyses indicate 3 : 1 (ligand to metal) stoichiometry. The lower molar conductance values of the complexes in DMSO at 10−3M concentration suggest their nonelectrolytic nature and are given in Table 1.Table 1
Elemental analysis and molar conductivity measurements.
Compounds
Color
Calculated (found)*
C
H
N
M
Molar conductance**
HL
White
51.40 (51.42)
3.55 (3.56)
7.05 (6.98)
—
—
[La(L)3·2H2O]
Yellow
44.82 (44.81)
3.39 (3.39)
6.15 (6.13)
10.16 (10.56)
5.2
[Pr(L)3·2H2O]
Yellow
44.76 (44.75)
3.39 (3.40)
6.14 (6.13)
10.30 (10.35)
7.8
[Nd(L)3·3H2O]
Light pink
44.17 (44.68)
3.27 (3.26)
6.06 (6.10)
10.40 (11.03)
6.3
[Sm(L)3·2H2O]
Yellow
44.45 (44.42)
3.36 (3.37)
6.10 (6.11)
10.91 (11.12)
9.8
[Eu(L)3·2H2O]
Yellow
44.40 (44.43)
3.36 (3.35)
6.09 (6.10)
11.01 (10.08)
7.1
[Gd(L)3·3H2O]
Off  white
43.76 (43.69)
3.24 (3.26)
6.00 (6.03)
11.23 (11.56)
4.6
[Dy(L)3·3H2O]
Yellow
43.59 (43.55)
3.23 (3.22)
5.98 (6.05)
11.56 (12.05)
8.5
[Y(L)3·2H2O]
Yellow
46.53 (46.31)
3.52 (3.53)
6.38 (6.39)
06.75 (06.86)
4.5
*The values in the parenthesis are calculated ones.
*
*Ohm−1 cm2 mol−1.Scheme 1
Synthetic route for the preparation of HL.


### 3.1. IR Spectral Studies

The characteristic IR bands of HL and Ln(III) complexes are compiled in Table2. In the IR spectrum of HL, the broad band observed at 3290 cm−1 was assigned to υ(OH) group of carboxylic acid. The appearance of this frequency at a slightly lower wave number is due to the involvement of OH group in intermolecular hydrogen bonding with oxygen atom of amide υ(C=O) functional group [21]. The HL shows a very strong absorption band at 1736 cm−1 assigned to the υ(C=O) of the carboxylic acid. The bands at 1659 cm−1 and 1609 cm−1 were assigned to amide υ(C=O) and υ(C=N), respectively.Table 2
FT-IR analysis of HL and[Ln(L)3·nH2O].
Compounds
υ (OH)
υ (CO)(amide)
υ (CO)(carboxylic)
υ
asy (COO−)
υ
s (COO−)
Δ
n (υasy−υs)
υ  (C=N)
υ (H2O)
HL
3290
1659
1736
n.o*
n.o
n.o
1609
n.o
NaL
n.o
1692
n.o
1603
1377
226
1611
3460
[La(L)3·2H2O]
n.o
1692
n.o
1590
1372
218
1609
3448
[Pr(L)3·2H2O]
n.o
1690
n.o
1564
1348
216
1613
3450
[Nd(L)3·3H2O]
n.o
1695
n.o
1574
1362
212
1612
3435
[Sm(L)3·2H2O]
n.o
1689
n.o
1574
1358
216
1608
3448
[Eu(L)3·2H2O]
n.o
1691
n.o
1581
1370
211
1610
3450
[Gd(L)3·3H2O]
n.o
1692
n.o
1573
1355
218
1613
3421
[Dy(L)3·3H2O]
n.o
1696
n.o
1562
1346
216
1608
3425
[Y(L)3·2H2O]
n.o
1693
n.o
1581
1368
213
1613
3448
*n.o: not observed.The absence of band due to carboxylic OH group in the spectra of all lanthanide (III) complexes suggests the coordination to the metal ion via deprotonation. The band due toυ(C=O) of carboxylic acid of HL was absent in all the lanthanide(III) complexes, indicating the participation of carbonyl oxygen in coordination to the metal ion [22–24]. Whereas the two new characteristic bands appeared on complexation in the ranges of 1590–1562 cm−1 and 1372–1346 cm−1 were assigned to asymmetric and symmetric stretching frequencies of carboxylate ion, respectively. The difference between υasy(COO-) and υsy(COO-) frequencies in all complexes were found to be less than the difference observed in the sodium salt of HL. This implies that carboxylic acid group of HL has coordinated to metal ion in bidentate fashion via deprotonation as suggested by Deacon [22, 25–28].A strong band at 1659 cm−1 in the spectrum of uncoordinated ligand is assigned to amide υ(C=O) and has shifted to higher wave number in all complexes. This increase in frequency of amide υ(C=O) functional group may be due to the breaking of hydrogen bond present in HL on complexation. Azomethine nitrogen υ(C=N) has not suffered any change on complexation indicating its noninvolvement in coordination.The presence of a broad band in the region 3460–3421 cm−1 was attributed to the  υ(O-H) of coordinated water molecules in all complexes [29]. This was further confirmed by the appearance of a weak nonligand band in the region 831–856 cm−1, assignable to rocking mode of coordinated water molecule [30]. The presence of coordinated water molecules was further confirmed by thermal studies.


### 3.2. NMR Spectroscopy

#### 3.2.1. NMR Spectral Studies of HL

The NMR spectrum of HL shows double set of signals for all the protons and carbons due to its existence in E and Z isomeric forms arising due to the restricted rotation along the (–HC=N–) functional group. From the1H NMR data, the ratio of E and Z isomers was approximately found to be 70 : 30 with E isomer as the predominant one over Z isomer.The isomeric structure of HL and its numbering are presented in Figure1. The 1H NMR spectrum is given in Figure 2. The detailed assignments for both the isomers are given in the experimental section. The singlet observed at 13.16 ppm was assigned to OH proton of carboxylic acid. Two singlets observed at 11.72 and 11.80 ppm were assigned to amide NH  and NH′ protons, respectively. The C11H  and C11′H protons present adjacent to carbonyl functional group were observed at 4.80 and 5.31 ppm, respectively. The C2H and C2′H protons present adjacent to carboxylic functional group were observed at 4.82 and 4.90 ppm, respectively. The azomethine protons C9H and C9′H were observed at 8.42 and 8.65 ppm, respectively [31]. The aromatic protons were observed between 6.99–7.90 ppm.Figure 1
Isomeric structures of HL.Figure 2
1H NMR spectrum of HL.1H-NMR analysis is supported by the 13C and DEPT NMR spectral analysis for the confirmation of E and Z isomerism in HL. The 13C spectrum of HL is given in Figure 3 and its DEPT NMR is presented at Figure 4. DEPT NMR at 135° pulse assisted the assignment of 13C resonance. The primary and tertiary carbons phased up and secondary carbons phased down, while signals for quaternary carbons and other carbons with no attached protons found absent in DEPT NMR were observed at 169.88 and 169.94 ppm, respectively. These quaternary carbons were absent in DEPT-135 NMR spectrum. Carbon signals at 116.15, 117.13 (C8/C8′), 122.02, 122.20 (C13/C13′), 125.58, 125.77 (C15/C15′), 152.54, 152.73 (C12/C12′), 163.28, 168.12 (C10/C10′), 156.08, 156.20 (C3/C3′), and 169.88, 169.94 (C1/C1′) were absent in DEPT-135 NMR spectrum which confirms their assignment for quaternary carbons. In 13C NMR spectrum, the methyne carbons C9  and  C9′ were resonated at 139.00 and 143.00 ppm, respectively, and were phased up in the DEPT-135 NMR spectrum. The signals for CH carbons in DEPT NMR were observed at 112.47, 112.84 (C4/C4′), 114.97, 115.12 (C17/C17′), 121.01, 121.03 (C16/C16′), 124.38, 124.94 (C6/C6′), 127.75, 127.96 (C5/C5′), 129.12, 129.30 (C7/C7′), 131.15, 131.44 (C14/C14′) and 139.00, 143.00 (C9/C9′) phased up in spectrum are in accordance with 13C NMR spectral assignments. The C11 and C11′ carbons were observed at 64.67 and 66.91 ppm, respectively. The signals for C2 and C2′ were observed at 64.62 and 64.75 ppm, respectively. The signals for methylene carbons C11, C11′, C2, and C2′ were phased down in the DEPT NMR spectrum. In 13C NMR the resonances for aromatic carbons were observed in the range of 112.47–131.90 ppm.Figure 3
13C NMR spectrum of HL.Figure 4
DEPT NMR spectrum of HL.All assignments were further studied by 2D HMQC NMR spectral analysis and the spectrum is given in Figure5. The spectrum correlates the directly bonded 1H and 13C NMR resonances. The resonance observed in 1H NMR spectrum at 13.16 ppm, 11.72, and 11.80 were assigned to OH, NH, and NH′ protons, respectively, which are devoid of any attached carbon signals, and were confirmed in 2D HMQC NMR spectral analysis.Figure 5
2D HMQC NMR spectrum of HL.Isomeric Investigation of HL by Variable Temperature NMR. It is well known that, in case of E and Z isomers, as the temperature increases, interconversion of two isomers also increases due to the increase in rotation along C8-C9 bond and one of them will become predominant over the other isomer at higher temperature [32]. To study the interconvertion phenomenon, variable temperature 1H NMR analyses were undertaken [33] in the temperature range from 298 K to 363 K and the spectrum is given in Figure 6. At room temperature (298 K), the resonances for both E and Z isomers are well distinguishable. As the temperature is increased (308 K), the intensities of signals assigned to Z isomer have comparatively decreased. At 338 K, signals were reduced to more than half of their original intensity. It implies that increase in temperature causes the interconvertion of two isomers. At 348 K, the signals observed at 8.65 ppm (C9′H), 7.61 ppm (C7′H), and 5.31 ppm (C11′H) for Z isomer have completely disappeared and at the same time, the remaining signals have also suffered a significant decrease in their intensities. At 363 K, almost a single set of signals was observed.Figure 6
Variable temperature1HNMR spectra of HL.


#### 3.2.2. NMR Spectral Studies of Lanthanum(III) Complex

In the spectrum of the lanthanum(III) complex shown at Figure7, only one set of signals was observed for each proton and carbon, indicating the presence of HL in only one form. The resonance at 13.16 ppm assigned to carboxylic acid proton in HL has completely disappeared in lanthanum(III) complex indicating the coordination of HL through carboxylic group via deprotonation [22]. The carboxylic acid coordination was supported by downfield shift of CH2  protons adjacent to carboxylic acid. The C11H, C9H, and NH signals observed at 4.80, 8.42, and 11.72 ppm, respectively in the spectrum of HL have now been observed at 4.70, 8.41, and 11.64 ppm, respectively, in the NMR spectrum of lanthanum(III) complex. This observation confirms that the carbonyl oxygen and azomethine nitrogen were not involved in coordination. Significant changes were not observed in the chemical shift values of aromatic protons after complexation.Figure 7
1H NMR spectrum of [La (L)3·2H2O].13C NMR spectrum further supports the mode of coordination of HL. The spectrum is given in Figure 8. The carbonyl carbon C1 of carboxylic acid observed at 169.88 ppm in HL has shifted downfield to 176.46 ppm in the complex suggesting the involvement of carboxylic acid in coordination [22]. It was further supported by downfield shift of C2 carbon resonance from 64.62 ppm to 67.55 ppm on complexation. Small change was observed in the chemical shift values of carbonyl carbon C10 on complexation. This indicates breakdown of hydrogen bond after complexation [34].Figure 8
13C NMR spectrum of [La (L)3·2H2O].The13C NMR spectral assignment was further confirmed by DEPT spectral analysis and is presented in Figure 9. The CH2  carbons observed at 66.91 ppm assigned to C11 carbon  and 67.55 ppm to C2 carbon were found phased down in the spectrum. The signals at 112.47 (C4), 115.28 (C17), 122.25 (C16), 126.92 (C6), 127.96 (C5), 129.12 (C7), 131.90 (C14), and 138.50 (C9) are due to CH carbons and were found phased up in the spectrum. The resonances at 117.13 (C8), 122.13 (C13), 124.30 (C15), 153.00 (C12), 156.08, (C3), 164.30 (C10), and 176.46 (C1) are the quaternary carbons, which were absent in DEPT spectrum.Figure 9
DEPT NMR spectrum of [La (L)3·2H2O].The assignments were further studied by 2D HMQC NMR spectrum. The spectrum is given in Figure10. The resonance observed at 11.64 ppm was devoid of any attached carbon signal confirming their assignment to NH proton.Figure 10
2D HMQC spectrum of [La (L)3·2H2O].


### 3.3. Thermal Analysis

The thermal stability data of Ln(III) complexes (where, Ln = La, Pr, Nd, Sm, Eu, Gd, Dy, Y) are compiled in Table3. As a representative, the thermogram of La(III) complex is shown in Figure 11. Under nitrogen atmosphere, the complex undergoes a three-step decomposition. The first weight loss found between 137.03°C to 210.02°C corresponds to the loss of coordinated water molecules. In La(III), Pr(III), Sm(III), Eu(III), and Y(III) complexes, the weight loss corresponds to two coordinated water molecules while in Nd(III), Gd(III), and Dy(III) complexes, the weight loss corresponds to three coordinated water molecules. The second weight loss between 365.71°C to 510.60°C is due to the loss of three ligand molecules in all complexes. Third step weight losses correspond to the formation of residue. The observed weight losses match with calculated values for the metal content [35].Table 3
Thermal decomposition data of[Ln(L)3·nH2O].
Complex
Temperature range (°C)
Weight loss in %
Molecule decomposed
Found
Calculated
[La(L)3·2H2O]
137.03–148.31
2.60
2.64
2H2O
378.71–488.46
87.38
87.35
3L
488.46–1000
10.02
10.01
Residue
[Pr(L)3·2H2O]
152.03–173
2.66
2.64
2H2O
375.71–510.60
87.35
87.23
3L
510.60–1000
9.99
10.13
Residue
[Nd(L)3·3H2O]
155.33–181.03
3.90
3.91
3H2O
376.71–485.32
86.13
86.03
3L
485.32–1000
9.97
10.06
Residue
[Sm(L)3·2H2O]
142.03–170.02
2.59
2.62
2H2O
365.71–483.40
86.50
86.53
3L
483.40–1000
10.91
10.85
Residue
[Eu(L)3·2H2O]
162.51–210.02
2.65
2.62
2H2O
370.71–484
86.40
86.46
3L
484–1000
10.95
10.92
Residue
[Gd(L)3·3H2O]
168.20–203.72
3.82
3.86
3H2O
369–477
85.11
85.14
3L
477–1000
11.07
11.10
Residue
[Dy(L)3·3H2O]
156.03–183.66
3.87
3.84
3H2O
386.01–495.48
84.72
84.68
3L
495.48–1000
11.41
11.48
Residue
[Y(L)3·2H2O]
156–173
2.72
2.71
2H2O
374.07–488
89.55
89.59
3L
488–1000
7.73
07.70
ResidueFigure 11
Thermogram of [La (L)3·2H2O].


### 3.4. ESI Mass Analysis

The ESI mass spectrum of HL is given in Figure12. The molecular ion peak m/z = 419 corresponds to sodium adduct of HL. The mass spectra of complexes are in good agreement with the expected molecular weights. All the molecular weights are adducting with the proton. The representative spectrum of lanthanum complex is given in Figure 13. In the given mass spectrum of La(III) complex, the molecular ion peak m/z = 1358 due to [M-H]+ ion.Figure 12
Mass spectrum of HL.Figure 13
ESI mass spectrum of [La (L)3·2H2O].


### 3.5. Electronic Spectra

The electronic spectra of HL and its lanthanide(III) complexes were recorded in DMSO at room temperature. The two absorption maxima at 278 nm and 318 nm in the spectrum of HL were assigned toπ→π* transitions of carbonyl oxygen and azomethine (–C=N–) moiety. No significant changes were observed in these bands on complexation.


### 3.6. Emission Spectra

The emission spectrum of Eu(III) complex was recorded in the solid state in the range of 300–700 nm by selective excitation wavelength at 318 nm. The three emission peaks observed were at 578, 592, and 612 nm. The bands at 578, 592, and 612 nm were assigned toD05→F07, D05→F17, and D05→F27, respectively. The intensity of 612 nm band was found to be more intense than other two bands [36, 37].From the spectral and analytical data, the general molecular formula for complexes is given as [Ln(L)3·nH2O]. The tentative structure for the complexes is given in Figure 14.Figure 14
Tentative structure of [Ln (L)3·nH2O].


### 3.7. Plant Growth Activity

The effect of HL and its La(III) complex at different concentrations in the growth of root and shoot of the germinated wheat seeds was analyzed with statistical analysis and data are compiled in Table4. Among the three concentrations used, that is, 1 × 10−5, 1 × 10−6, and 1 ×10−7M, the growth of root and shoot is more at 1 × 10−6M concentration. The growth activity exhibited by HL (Group 5) and La(III) complex (Group 6) is compared with that of control (Group 1), solvent (Group 2), standard auxin (Group 3), and metal salt (Group 4). At 1 × 10−5M concentration, the percentage germination is less, while it is good and almost equal at 1 × 10−6 and 1 ×10−7M concentrations. This indicates that, at 10−6M concentration, both HL and La(III) complex have more growth promoting activity than the standard auxin used.Table 4
Plant growth activity on wheat seeds.
Sl. no.
Treatments
Germination percentage
Root length(cm)
Shoot length(cm)
1 × 10−5 M
1
Group 1**
100
18.60 ± 1.89fghi
16.14 ± 0.56bc*
2
Group 2
97.60
18.08 ± 1.95ghi
16.03 ± 0.53bc
3
Group 3
80.00
17.87 ± 2.88hi
15.55 ± 1.67c
4
Group 4
85.00
20.97 ± 2.37cd
16.31 ± 1.48bc
5
Group 5
76.00
16.82 ± 1.29i
13.09 ± 1.91e
6
Group 6
80.50
14.77 ± 0.92j
13.56 ± 2.47de
1 × 10−6 M
1
Group 1
98.50
18.04 ± 0.92ghi
16.55 ± 2.03bc
2
Group 2
98.00
17.91 ± 2.63hi
16.28 ± 1.63c
3
Group 3
98.60
21.35 ± 1.30c
17.06 ± 2.22bc
4
Group 4
98.00
20.05 ± 2.14cdefg
16.84 ± 2.15bc
5
Group 5
100
25.70 ± 1.25b
18.10 ± 2.23b
6
Group 6
98.50
28.28 ± 0.99a
20.06 ± 1.22a
1 × 10−7 M
1
Group 1
100
18.90 ± 2.51efgh
15.98 ± 2.04bc
2
Group 2
99.00
18.82 ± 3.08efghi
16.17 ± 2.94bc
3
Group 3
98.00
19.32 ± 0.73defgh
15.27 ± 2.52cd
4
Group 4
96.00
20.26 ± 2.37cdef
16.92 ± 1.77bc
5
Group 5
98.50
20.74 ± 1.83cde
16.48 ± 3.17bc
6
Group 6
98.00
21.78 ± 2.63c
16.24 ± 1.13bc
*Values are the mean of triplicates. Mean values followed by different letters are significantly different within a row or column at 5% probability level; that is, α = 0.05 (P<0.05) according to Duncan statistics.
*
*Group 1: water, Group 2: DMSO and water mixture, Group 3: 2,4-D, Group 4: lanthanum nitrate, Group 5: HL, Group 6: La (III) complex.


#### 3.7.1. Shoot Length

At 1× 10−5M concentration, surprisingly Groups 5 and 6 have shown a decreased activity while at 1 × 10−7M, a significant change was observed. But at 1 × 10−6M, a significant enhancement in shoot length is observed on complexation (Group 6). The graph is given in Figure 15.Figure 15
Comparison of shoot length Group 1 = Water, Group 2 = DMSO and Water mixture, Group 3 = 2,4-D, Group 4 = Lanthanum nitrate, Group 5 = HL, Group 6 = La(L)3·2H2O.


#### 3.7.2. Root Length

At 1×10−5M concentration, Group 4 has shown significant increase in root length compared to Groups 3, 5, and 6. At 1 × 10−6M, Groups 5 and 6 have shown significant enhancement in root length compared to even Group 3 (the commercial auxin). When compared among the Groups 5 and 6, the activity has enhanced on complexation (Group 6). A similar observation is made at 1 × 10−7M concentration, but the extent of enhancement is less compared to that at 1 × 10−6M. The comparative graph is given in Figure 16.Figure 16
Comparison of root length. Group 1 = Water, Group 2 = DMSO and Water mixture, Group 3 = 2,4-D, Group 4 = Lanthanum nitrate, Group 5 = HL, Group 6 = La(L)3·2H2O.


## 3.1. IR Spectral Studies

The characteristic IR bands of HL and Ln(III) complexes are compiled in Table2. In the IR spectrum of HL, the broad band observed at 3290 cm−1 was assigned to υ(OH) group of carboxylic acid. The appearance of this frequency at a slightly lower wave number is due to the involvement of OH group in intermolecular hydrogen bonding with oxygen atom of amide υ(C=O) functional group [21]. The HL shows a very strong absorption band at 1736 cm−1 assigned to the υ(C=O) of the carboxylic acid. The bands at 1659 cm−1 and 1609 cm−1 were assigned to amide υ(C=O) and υ(C=N), respectively.Table 2
FT-IR analysis of HL and[Ln(L)3·nH2O].
Compounds
υ (OH)
υ (CO)(amide)
υ (CO)(carboxylic)
υ
asy (COO−)
υ
s (COO−)
Δ
n (υasy−υs)
υ  (C=N)
υ (H2O)
HL
3290
1659
1736
n.o*
n.o
n.o
1609
n.o
NaL
n.o
1692
n.o
1603
1377
226
1611
3460
[La(L)3·2H2O]
n.o
1692
n.o
1590
1372
218
1609
3448
[Pr(L)3·2H2O]
n.o
1690
n.o
1564
1348
216
1613
3450
[Nd(L)3·3H2O]
n.o
1695
n.o
1574
1362
212
1612
3435
[Sm(L)3·2H2O]
n.o
1689
n.o
1574
1358
216
1608
3448
[Eu(L)3·2H2O]
n.o
1691
n.o
1581
1370
211
1610
3450
[Gd(L)3·3H2O]
n.o
1692
n.o
1573
1355
218
1613
3421
[Dy(L)3·3H2O]
n.o
1696
n.o
1562
1346
216
1608
3425
[Y(L)3·2H2O]
n.o
1693
n.o
1581
1368
213
1613
3448
*n.o: not observed.The absence of band due to carboxylic OH group in the spectra of all lanthanide (III) complexes suggests the coordination to the metal ion via deprotonation. The band due toυ(C=O) of carboxylic acid of HL was absent in all the lanthanide(III) complexes, indicating the participation of carbonyl oxygen in coordination to the metal ion [22–24]. Whereas the two new characteristic bands appeared on complexation in the ranges of 1590–1562 cm−1 and 1372–1346 cm−1 were assigned to asymmetric and symmetric stretching frequencies of carboxylate ion, respectively. The difference between υasy(COO-) and υsy(COO-) frequencies in all complexes were found to be less than the difference observed in the sodium salt of HL. This implies that carboxylic acid group of HL has coordinated to metal ion in bidentate fashion via deprotonation as suggested by Deacon [22, 25–28].A strong band at 1659 cm−1 in the spectrum of uncoordinated ligand is assigned to amide υ(C=O) and has shifted to higher wave number in all complexes. This increase in frequency of amide υ(C=O) functional group may be due to the breaking of hydrogen bond present in HL on complexation. Azomethine nitrogen υ(C=N) has not suffered any change on complexation indicating its noninvolvement in coordination.The presence of a broad band in the region 3460–3421 cm−1 was attributed to the  υ(O-H) of coordinated water molecules in all complexes [29]. This was further confirmed by the appearance of a weak nonligand band in the region 831–856 cm−1, assignable to rocking mode of coordinated water molecule [30]. The presence of coordinated water molecules was further confirmed by thermal studies.


## 3.2. NMR Spectroscopy

### 3.2.1. NMR Spectral Studies of HL

The NMR spectrum of HL shows double set of signals for all the protons and carbons due to its existence in E and Z isomeric forms arising due to the restricted rotation along the (–HC=N–) functional group. From the1H NMR data, the ratio of E and Z isomers was approximately found to be 70 : 30 with E isomer as the predominant one over Z isomer.The isomeric structure of HL and its numbering are presented in Figure1. The 1H NMR spectrum is given in Figure 2. The detailed assignments for both the isomers are given in the experimental section. The singlet observed at 13.16 ppm was assigned to OH proton of carboxylic acid. Two singlets observed at 11.72 and 11.80 ppm were assigned to amide NH  and NH′ protons, respectively. The C11H  and C11′H protons present adjacent to carbonyl functional group were observed at 4.80 and 5.31 ppm, respectively. The C2H and C2′H protons present adjacent to carboxylic functional group were observed at 4.82 and 4.90 ppm, respectively. The azomethine protons C9H and C9′H were observed at 8.42 and 8.65 ppm, respectively [31]. The aromatic protons were observed between 6.99–7.90 ppm.Figure 1
Isomeric structures of HL.Figure 2
1H NMR spectrum of HL.1H-NMR analysis is supported by the 13C and DEPT NMR spectral analysis for the confirmation of E and Z isomerism in HL. The 13C spectrum of HL is given in Figure 3 and its DEPT NMR is presented at Figure 4. DEPT NMR at 135° pulse assisted the assignment of 13C resonance. The primary and tertiary carbons phased up and secondary carbons phased down, while signals for quaternary carbons and other carbons with no attached protons found absent in DEPT NMR were observed at 169.88 and 169.94 ppm, respectively. These quaternary carbons were absent in DEPT-135 NMR spectrum. Carbon signals at 116.15, 117.13 (C8/C8′), 122.02, 122.20 (C13/C13′), 125.58, 125.77 (C15/C15′), 152.54, 152.73 (C12/C12′), 163.28, 168.12 (C10/C10′), 156.08, 156.20 (C3/C3′), and 169.88, 169.94 (C1/C1′) were absent in DEPT-135 NMR spectrum which confirms their assignment for quaternary carbons. In 13C NMR spectrum, the methyne carbons C9  and  C9′ were resonated at 139.00 and 143.00 ppm, respectively, and were phased up in the DEPT-135 NMR spectrum. The signals for CH carbons in DEPT NMR were observed at 112.47, 112.84 (C4/C4′), 114.97, 115.12 (C17/C17′), 121.01, 121.03 (C16/C16′), 124.38, 124.94 (C6/C6′), 127.75, 127.96 (C5/C5′), 129.12, 129.30 (C7/C7′), 131.15, 131.44 (C14/C14′) and 139.00, 143.00 (C9/C9′) phased up in spectrum are in accordance with 13C NMR spectral assignments. The C11 and C11′ carbons were observed at 64.67 and 66.91 ppm, respectively. The signals for C2 and C2′ were observed at 64.62 and 64.75 ppm, respectively. The signals for methylene carbons C11, C11′, C2, and C2′ were phased down in the DEPT NMR spectrum. In 13C NMR the resonances for aromatic carbons were observed in the range of 112.47–131.90 ppm.Figure 3
13C NMR spectrum of HL.Figure 4
DEPT NMR spectrum of HL.All assignments were further studied by 2D HMQC NMR spectral analysis and the spectrum is given in Figure5. The spectrum correlates the directly bonded 1H and 13C NMR resonances. The resonance observed in 1H NMR spectrum at 13.16 ppm, 11.72, and 11.80 were assigned to OH, NH, and NH′ protons, respectively, which are devoid of any attached carbon signals, and were confirmed in 2D HMQC NMR spectral analysis.Figure 5
2D HMQC NMR spectrum of HL.Isomeric Investigation of HL by Variable Temperature NMR. It is well known that, in case of E and Z isomers, as the temperature increases, interconversion of two isomers also increases due to the increase in rotation along C8-C9 bond and one of them will become predominant over the other isomer at higher temperature [32]. To study the interconvertion phenomenon, variable temperature 1H NMR analyses were undertaken [33] in the temperature range from 298 K to 363 K and the spectrum is given in Figure 6. At room temperature (298 K), the resonances for both E and Z isomers are well distinguishable. As the temperature is increased (308 K), the intensities of signals assigned to Z isomer have comparatively decreased. At 338 K, signals were reduced to more than half of their original intensity. It implies that increase in temperature causes the interconvertion of two isomers. At 348 K, the signals observed at 8.65 ppm (C9′H), 7.61 ppm (C7′H), and 5.31 ppm (C11′H) for Z isomer have completely disappeared and at the same time, the remaining signals have also suffered a significant decrease in their intensities. At 363 K, almost a single set of signals was observed.Figure 6
Variable temperature1HNMR spectra of HL.


### 3.2.2. NMR Spectral Studies of Lanthanum(III) Complex

In the spectrum of the lanthanum(III) complex shown at Figure7, only one set of signals was observed for each proton and carbon, indicating the presence of HL in only one form. The resonance at 13.16 ppm assigned to carboxylic acid proton in HL has completely disappeared in lanthanum(III) complex indicating the coordination of HL through carboxylic group via deprotonation [22]. The carboxylic acid coordination was supported by downfield shift of CH2  protons adjacent to carboxylic acid. The C11H, C9H, and NH signals observed at 4.80, 8.42, and 11.72 ppm, respectively in the spectrum of HL have now been observed at 4.70, 8.41, and 11.64 ppm, respectively, in the NMR spectrum of lanthanum(III) complex. This observation confirms that the carbonyl oxygen and azomethine nitrogen were not involved in coordination. Significant changes were not observed in the chemical shift values of aromatic protons after complexation.Figure 7
1H NMR spectrum of [La (L)3·2H2O].13C NMR spectrum further supports the mode of coordination of HL. The spectrum is given in Figure 8. The carbonyl carbon C1 of carboxylic acid observed at 169.88 ppm in HL has shifted downfield to 176.46 ppm in the complex suggesting the involvement of carboxylic acid in coordination [22]. It was further supported by downfield shift of C2 carbon resonance from 64.62 ppm to 67.55 ppm on complexation. Small change was observed in the chemical shift values of carbonyl carbon C10 on complexation. This indicates breakdown of hydrogen bond after complexation [34].Figure 8
13C NMR spectrum of [La (L)3·2H2O].The13C NMR spectral assignment was further confirmed by DEPT spectral analysis and is presented in Figure 9. The CH2  carbons observed at 66.91 ppm assigned to C11 carbon  and 67.55 ppm to C2 carbon were found phased down in the spectrum. The signals at 112.47 (C4), 115.28 (C17), 122.25 (C16), 126.92 (C6), 127.96 (C5), 129.12 (C7), 131.90 (C14), and 138.50 (C9) are due to CH carbons and were found phased up in the spectrum. The resonances at 117.13 (C8), 122.13 (C13), 124.30 (C15), 153.00 (C12), 156.08, (C3), 164.30 (C10), and 176.46 (C1) are the quaternary carbons, which were absent in DEPT spectrum.Figure 9
DEPT NMR spectrum of [La (L)3·2H2O].The assignments were further studied by 2D HMQC NMR spectrum. The spectrum is given in Figure10. The resonance observed at 11.64 ppm was devoid of any attached carbon signal confirming their assignment to NH proton.Figure 10
2D HMQC spectrum of [La (L)3·2H2O].


## 3.2.1. NMR Spectral Studies of HL

The NMR spectrum of HL shows double set of signals for all the protons and carbons due to its existence in E and Z isomeric forms arising due to the restricted rotation along the (–HC=N–) functional group. From the1H NMR data, the ratio of E and Z isomers was approximately found to be 70 : 30 with E isomer as the predominant one over Z isomer.The isomeric structure of HL and its numbering are presented in Figure1. The 1H NMR spectrum is given in Figure 2. The detailed assignments for both the isomers are given in the experimental section. The singlet observed at 13.16 ppm was assigned to OH proton of carboxylic acid. Two singlets observed at 11.72 and 11.80 ppm were assigned to amide NH  and NH′ protons, respectively. The C11H  and C11′H protons present adjacent to carbonyl functional group were observed at 4.80 and 5.31 ppm, respectively. The C2H and C2′H protons present adjacent to carboxylic functional group were observed at 4.82 and 4.90 ppm, respectively. The azomethine protons C9H and C9′H were observed at 8.42 and 8.65 ppm, respectively [31]. The aromatic protons were observed between 6.99–7.90 ppm.Figure 1
Isomeric structures of HL.Figure 2
1H NMR spectrum of HL.1H-NMR analysis is supported by the 13C and DEPT NMR spectral analysis for the confirmation of E and Z isomerism in HL. The 13C spectrum of HL is given in Figure 3 and its DEPT NMR is presented at Figure 4. DEPT NMR at 135° pulse assisted the assignment of 13C resonance. The primary and tertiary carbons phased up and secondary carbons phased down, while signals for quaternary carbons and other carbons with no attached protons found absent in DEPT NMR were observed at 169.88 and 169.94 ppm, respectively. These quaternary carbons were absent in DEPT-135 NMR spectrum. Carbon signals at 116.15, 117.13 (C8/C8′), 122.02, 122.20 (C13/C13′), 125.58, 125.77 (C15/C15′), 152.54, 152.73 (C12/C12′), 163.28, 168.12 (C10/C10′), 156.08, 156.20 (C3/C3′), and 169.88, 169.94 (C1/C1′) were absent in DEPT-135 NMR spectrum which confirms their assignment for quaternary carbons. In 13C NMR spectrum, the methyne carbons C9  and  C9′ were resonated at 139.00 and 143.00 ppm, respectively, and were phased up in the DEPT-135 NMR spectrum. The signals for CH carbons in DEPT NMR were observed at 112.47, 112.84 (C4/C4′), 114.97, 115.12 (C17/C17′), 121.01, 121.03 (C16/C16′), 124.38, 124.94 (C6/C6′), 127.75, 127.96 (C5/C5′), 129.12, 129.30 (C7/C7′), 131.15, 131.44 (C14/C14′) and 139.00, 143.00 (C9/C9′) phased up in spectrum are in accordance with 13C NMR spectral assignments. The C11 and C11′ carbons were observed at 64.67 and 66.91 ppm, respectively. The signals for C2 and C2′ were observed at 64.62 and 64.75 ppm, respectively. The signals for methylene carbons C11, C11′, C2, and C2′ were phased down in the DEPT NMR spectrum. In 13C NMR the resonances for aromatic carbons were observed in the range of 112.47–131.90 ppm.Figure 3
13C NMR spectrum of HL.Figure 4
DEPT NMR spectrum of HL.All assignments were further studied by 2D HMQC NMR spectral analysis and the spectrum is given in Figure5. The spectrum correlates the directly bonded 1H and 13C NMR resonances. The resonance observed in 1H NMR spectrum at 13.16 ppm, 11.72, and 11.80 were assigned to OH, NH, and NH′ protons, respectively, which are devoid of any attached carbon signals, and were confirmed in 2D HMQC NMR spectral analysis.Figure 5
2D HMQC NMR spectrum of HL.Isomeric Investigation of HL by Variable Temperature NMR. It is well known that, in case of E and Z isomers, as the temperature increases, interconversion of two isomers also increases due to the increase in rotation along C8-C9 bond and one of them will become predominant over the other isomer at higher temperature [32]. To study the interconvertion phenomenon, variable temperature 1H NMR analyses were undertaken [33] in the temperature range from 298 K to 363 K and the spectrum is given in Figure 6. At room temperature (298 K), the resonances for both E and Z isomers are well distinguishable. As the temperature is increased (308 K), the intensities of signals assigned to Z isomer have comparatively decreased. At 338 K, signals were reduced to more than half of their original intensity. It implies that increase in temperature causes the interconvertion of two isomers. At 348 K, the signals observed at 8.65 ppm (C9′H), 7.61 ppm (C7′H), and 5.31 ppm (C11′H) for Z isomer have completely disappeared and at the same time, the remaining signals have also suffered a significant decrease in their intensities. At 363 K, almost a single set of signals was observed.Figure 6
Variable temperature1HNMR spectra of HL.


## 3.2.2. NMR Spectral Studies of Lanthanum(III) Complex

In the spectrum of the lanthanum(III) complex shown at Figure7, only one set of signals was observed for each proton and carbon, indicating the presence of HL in only one form. The resonance at 13.16 ppm assigned to carboxylic acid proton in HL has completely disappeared in lanthanum(III) complex indicating the coordination of HL through carboxylic group via deprotonation [22]. The carboxylic acid coordination was supported by downfield shift of CH2  protons adjacent to carboxylic acid. The C11H, C9H, and NH signals observed at 4.80, 8.42, and 11.72 ppm, respectively in the spectrum of HL have now been observed at 4.70, 8.41, and 11.64 ppm, respectively, in the NMR spectrum of lanthanum(III) complex. This observation confirms that the carbonyl oxygen and azomethine nitrogen were not involved in coordination. Significant changes were not observed in the chemical shift values of aromatic protons after complexation.Figure 7
1H NMR spectrum of [La (L)3·2H2O].13C NMR spectrum further supports the mode of coordination of HL. The spectrum is given in Figure 8. The carbonyl carbon C1 of carboxylic acid observed at 169.88 ppm in HL has shifted downfield to 176.46 ppm in the complex suggesting the involvement of carboxylic acid in coordination [22]. It was further supported by downfield shift of C2 carbon resonance from 64.62 ppm to 67.55 ppm on complexation. Small change was observed in the chemical shift values of carbonyl carbon C10 on complexation. This indicates breakdown of hydrogen bond after complexation [34].Figure 8
13C NMR spectrum of [La (L)3·2H2O].The13C NMR spectral assignment was further confirmed by DEPT spectral analysis and is presented in Figure 9. The CH2  carbons observed at 66.91 ppm assigned to C11 carbon  and 67.55 ppm to C2 carbon were found phased down in the spectrum. The signals at 112.47 (C4), 115.28 (C17), 122.25 (C16), 126.92 (C6), 127.96 (C5), 129.12 (C7), 131.90 (C14), and 138.50 (C9) are due to CH carbons and were found phased up in the spectrum. The resonances at 117.13 (C8), 122.13 (C13), 124.30 (C15), 153.00 (C12), 156.08, (C3), 164.30 (C10), and 176.46 (C1) are the quaternary carbons, which were absent in DEPT spectrum.Figure 9
DEPT NMR spectrum of [La (L)3·2H2O].The assignments were further studied by 2D HMQC NMR spectrum. The spectrum is given in Figure10. The resonance observed at 11.64 ppm was devoid of any attached carbon signal confirming their assignment to NH proton.Figure 10
2D HMQC spectrum of [La (L)3·2H2O].


## 3.3. Thermal Analysis

The thermal stability data of Ln(III) complexes (where, Ln = La, Pr, Nd, Sm, Eu, Gd, Dy, Y) are compiled in Table3. As a representative, the thermogram of La(III) complex is shown in Figure 11. Under nitrogen atmosphere, the complex undergoes a three-step decomposition. The first weight loss found between 137.03°C to 210.02°C corresponds to the loss of coordinated water molecules. In La(III), Pr(III), Sm(III), Eu(III), and Y(III) complexes, the weight loss corresponds to two coordinated water molecules while in Nd(III), Gd(III), and Dy(III) complexes, the weight loss corresponds to three coordinated water molecules. The second weight loss between 365.71°C to 510.60°C is due to the loss of three ligand molecules in all complexes. Third step weight losses correspond to the formation of residue. The observed weight losses match with calculated values for the metal content [35].Table 3
Thermal decomposition data of[Ln(L)3·nH2O].
Complex
Temperature range (°C)
Weight loss in %
Molecule decomposed
Found
Calculated
[La(L)3·2H2O]
137.03–148.31
2.60
2.64
2H2O
378.71–488.46
87.38
87.35
3L
488.46–1000
10.02
10.01
Residue
[Pr(L)3·2H2O]
152.03–173
2.66
2.64
2H2O
375.71–510.60
87.35
87.23
3L
510.60–1000
9.99
10.13
Residue
[Nd(L)3·3H2O]
155.33–181.03
3.90
3.91
3H2O
376.71–485.32
86.13
86.03
3L
485.32–1000
9.97
10.06
Residue
[Sm(L)3·2H2O]
142.03–170.02
2.59
2.62
2H2O
365.71–483.40
86.50
86.53
3L
483.40–1000
10.91
10.85
Residue
[Eu(L)3·2H2O]
162.51–210.02
2.65
2.62
2H2O
370.71–484
86.40
86.46
3L
484–1000
10.95
10.92
Residue
[Gd(L)3·3H2O]
168.20–203.72
3.82
3.86
3H2O
369–477
85.11
85.14
3L
477–1000
11.07
11.10
Residue
[Dy(L)3·3H2O]
156.03–183.66
3.87
3.84
3H2O
386.01–495.48
84.72
84.68
3L
495.48–1000
11.41
11.48
Residue
[Y(L)3·2H2O]
156–173
2.72
2.71
2H2O
374.07–488
89.55
89.59
3L
488–1000
7.73
07.70
ResidueFigure 11
Thermogram of [La (L)3·2H2O].


## 3.4. ESI Mass Analysis

The ESI mass spectrum of HL is given in Figure12. The molecular ion peak m/z = 419 corresponds to sodium adduct of HL. The mass spectra of complexes are in good agreement with the expected molecular weights. All the molecular weights are adducting with the proton. The representative spectrum of lanthanum complex is given in Figure 13. In the given mass spectrum of La(III) complex, the molecular ion peak m/z = 1358 due to [M-H]+ ion.Figure 12
Mass spectrum of HL.Figure 13
ESI mass spectrum of [La (L)3·2H2O].


## 3.5. Electronic Spectra

The electronic spectra of HL and its lanthanide(III) complexes were recorded in DMSO at room temperature. The two absorption maxima at 278 nm and 318 nm in the spectrum of HL were assigned toπ→π* transitions of carbonyl oxygen and azomethine (–C=N–) moiety. No significant changes were observed in these bands on complexation.


## 3.6. Emission Spectra

The emission spectrum of Eu(III) complex was recorded in the solid state in the range of 300–700 nm by selective excitation wavelength at 318 nm. The three emission peaks observed were at 578, 592, and 612 nm. The bands at 578, 592, and 612 nm were assigned toD05→F07, D05→F17, and D05→F27, respectively. The intensity of 612 nm band was found to be more intense than other two bands [36, 37].From the spectral and analytical data, the general molecular formula for complexes is given as [Ln(L)3·nH2O]. The tentative structure for the complexes is given in Figure 14.Figure 14
Tentative structure of [Ln (L)3·nH2O].


## 3.7. Plant Growth Activity

The effect of HL and its La(III) complex at different concentrations in the growth of root and shoot of the germinated wheat seeds was analyzed with statistical analysis and data are compiled in Table4. Among the three concentrations used, that is, 1 × 10−5, 1 × 10−6, and 1 ×10−7M, the growth of root and shoot is more at 1 × 10−6M concentration. The growth activity exhibited by HL (Group 5) and La(III) complex (Group 6) is compared with that of control (Group 1), solvent (Group 2), standard auxin (Group 3), and metal salt (Group 4). At 1 × 10−5M concentration, the percentage germination is less, while it is good and almost equal at 1 × 10−6 and 1 ×10−7M concentrations. This indicates that, at 10−6M concentration, both HL and La(III) complex have more growth promoting activity than the standard auxin used.Table 4
Plant growth activity on wheat seeds.
Sl. no.
Treatments
Germination percentage
Root length(cm)
Shoot length(cm)
1 × 10−5 M
1
Group 1**
100
18.60 ± 1.89fghi
16.14 ± 0.56bc*
2
Group 2
97.60
18.08 ± 1.95ghi
16.03 ± 0.53bc
3
Group 3
80.00
17.87 ± 2.88hi
15.55 ± 1.67c
4
Group 4
85.00
20.97 ± 2.37cd
16.31 ± 1.48bc
5
Group 5
76.00
16.82 ± 1.29i
13.09 ± 1.91e
6
Group 6
80.50
14.77 ± 0.92j
13.56 ± 2.47de
1 × 10−6 M
1
Group 1
98.50
18.04 ± 0.92ghi
16.55 ± 2.03bc
2
Group 2
98.00
17.91 ± 2.63hi
16.28 ± 1.63c
3
Group 3
98.60
21.35 ± 1.30c
17.06 ± 2.22bc
4
Group 4
98.00
20.05 ± 2.14cdefg
16.84 ± 2.15bc
5
Group 5
100
25.70 ± 1.25b
18.10 ± 2.23b
6
Group 6
98.50
28.28 ± 0.99a
20.06 ± 1.22a
1 × 10−7 M
1
Group 1
100
18.90 ± 2.51efgh
15.98 ± 2.04bc
2
Group 2
99.00
18.82 ± 3.08efghi
16.17 ± 2.94bc
3
Group 3
98.00
19.32 ± 0.73defgh
15.27 ± 2.52cd
4
Group 4
96.00
20.26 ± 2.37cdef
16.92 ± 1.77bc
5
Group 5
98.50
20.74 ± 1.83cde
16.48 ± 3.17bc
6
Group 6
98.00
21.78 ± 2.63c
16.24 ± 1.13bc
*Values are the mean of triplicates. Mean values followed by different letters are significantly different within a row or column at 5% probability level; that is, α = 0.05 (P<0.05) according to Duncan statistics.
*
*Group 1: water, Group 2: DMSO and water mixture, Group 3: 2,4-D, Group 4: lanthanum nitrate, Group 5: HL, Group 6: La (III) complex.


### 3.7.1. Shoot Length

At 1× 10−5M concentration, surprisingly Groups 5 and 6 have shown a decreased activity while at 1 × 10−7M, a significant change was observed. But at 1 × 10−6M, a significant enhancement in shoot length is observed on complexation (Group 6). The graph is given in Figure 15.Figure 15
Comparison of shoot length Group 1 = Water, Group 2 = DMSO and Water mixture, Group 3 = 2,4-D, Group 4 = Lanthanum nitrate, Group 5 = HL, Group 6 = La(L)3·2H2O.


### 3.7.2. Root Length

At 1×10−5M concentration, Group 4 has shown significant increase in root length compared to Groups 3, 5, and 6. At 1 × 10−6M, Groups 5 and 6 have shown significant enhancement in root length compared to even Group 3 (the commercial auxin). When compared among the Groups 5 and 6, the activity has enhanced on complexation (Group 6). A similar observation is made at 1 × 10−7M concentration, but the extent of enhancement is less compared to that at 1 × 10−6M. The comparative graph is given in Figure 16.Figure 16
Comparison of root length. Group 1 = Water, Group 2 = DMSO and Water mixture, Group 3 = 2,4-D, Group 4 = Lanthanum nitrate, Group 5 = HL, Group 6 = La(L)3·2H2O.


## 3.7.1. Shoot Length

At 1× 10−5M concentration, surprisingly Groups 5 and 6 have shown a decreased activity while at 1 × 10−7M, a significant change was observed. But at 1 × 10−6M, a significant enhancement in shoot length is observed on complexation (Group 6). The graph is given in Figure 15.Figure 15
Comparison of shoot length Group 1 = Water, Group 2 = DMSO and Water mixture, Group 3 = 2,4-D, Group 4 = Lanthanum nitrate, Group 5 = HL, Group 6 = La(L)3·2H2O.


## 3.7.2. Root Length

At 1×10−5M concentration, Group 4 has shown significant increase in root length compared to Groups 3, 5, and 6. At 1 × 10−6M, Groups 5 and 6 have shown significant enhancement in root length compared to even Group 3 (the commercial auxin). When compared among the Groups 5 and 6, the activity has enhanced on complexation (Group 6). A similar observation is made at 1 × 10−7M concentration, but the extent of enhancement is less compared to that at 1 × 10−6M. The comparative graph is given in Figure 16.Figure 16
Comparison of root length. Group 1 = Water, Group 2 = DMSO and Water mixture, Group 3 = 2,4-D, Group 4 = Lanthanum nitrate, Group 5 = HL, Group 6 = La(L)3·2H2O.


## 4. Conclusion

A series of lanthanide complexes were prepared by treating novel 2-{[2-(2, 4-dichloro-phenoxy)-acetyl]-hydrazonomethyl}-phenoxy)-acetic acid with lanthanide(III) nitrates. The structure of HL was confirmed by various spectroscopic techniques. NMR spectral data of HL reveal its existence in E and Z isomeric forms in solution at room temperature. The coordination mode of ligand is well established from elemental analysis, molar conductivity, IR, NMR, mass, electronic spectral, and thermal studies. The results confirm that ligand has coordinated in bidentate fashion through carboxylic acid group via deprotonation. Based on spectral and analytical results, tentative structure for complexes is given in Figure14. The results obtained on plant growth activity indicate that germination percentage is more at 1 × 10−6M and 1 × 10−7M concentrations. When compared between the standard, HL and lanthanum(III) complex, an enhancement in the plant growth activity was observed on complexation and the best activity was observed at 10−6M concentration.



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*Source: 810892-2012-11-25.xml*