Biochemical Characterization, Phytotoxic Effect and Antimicrobial Activity against Some Phytopathogens of New Gemifloxacin Schiff Base Metal Complexes

doi.org/10.1002/cbdv.202100365

FULL PAPER

Biochemical Characterization, Phytotoxic Effect and Antimicrobial

Activity against Some Phytopathogens of New Gemifloxacin Schiff

Base Metal Complexes

Amira A. Mohamed,*^aHazem S. Elshafie,*^bSadeek A. Sadeek,*^cand Ippolito Camele*^b

a

Department of Basic Science, Zagazig Higher Institute of Engineering and Technology, 44519 Zagazig, Egypt,

e-mail: amira222283@yahoo.com; aa.adaim@science.zu.edu.eg

School of Agricultural, Forestry, Food and Environmental Sciences, University of Basilicata,

b

Viale dell’Ateneo Lucano 10, 85100 Potenza, Italy, e-mail: hazem.elshafie@unibas.it; ippolito.camele@unibas.it

c

Department of Chemistry, Faculty of Science, Zagazig University, 44519 Zagazig, Egypt,

e-mail: s_sadeek@zu.edu.eg

Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original

work is properly cited.

String of Fe(III), Cu(II), Zn(II) and Zr(IV) complexes were synthesized with tetradentateamino Schiff base ligand

derived by condensation of ethylene diamine with gemifloxacin. The novel Schiff base (4E,4’E)-4,4’-(ethane-1,2-

diyldiazanylylidene)bis{7-[(4Z)-3-(aminomethyl)-4-(methoxyimino)pyrrolidin-1-yl]-1-cyclopropyl-6-fluoro-1,4-dihy-

dro-1,8-naphthyridine-3-carboxylic acid} (GMFX-en) and its metal complexes were identified and confirmed by

1

elemental analyses, FT-IR, UV/VIS, H-NMR spectra, magnetic susceptibility, conductometric measurements and

thermal analyses. The FT-IR spectral data showed the chelation behavior of GMFX-en toward the metal ions

through oxygen of carboxylate group and nitrogen of azomethine group. In the light of all spectral data, these

complexes presumably have octahedral geometry configurations. Thermal analysis specified that the decaying of

the metal complexes exist in two or three steps with the final residue metal oxides. Antimicrobial activity of the

new prepared metal complexes was screened against some common phytopathogens and their mode of action

has been also discussed. The potential phytotoxic effectiveness of the new complexes was furthermore inspected

on two commonly experimental plants. The complexes showed significant antimicrobial and phytotoxic effects

against the majority of tested phytopathogens and the two tested plants, respectively. The potential

antimicrobial activity of the complexes proved their possibility to be used successfully in agropharmacutical

industry to control many serious phytopathogens. The phytotoxicity of the studied complexes also indicated

their possibility as potential bio-based herbicides alternatives to weed control in crop fields.

Keywords: gemifloxacin Schiff base, metal complexes, antimicrobial activity, phytotoxicity, chelation theory,

phytopathogens.

electronic features, and good solubility in many

Introduction

solvents.^[1,2]The presence of azomethine group in

Schiff bases can be effortlessly produced by the Schiff base ligands performs a paramount role in their

intensification reaction of primary amines together antimicrobial activities. Schiff base ligand is capable of

with carbonyl compounds. Schiff base ligands have coordinating with metal ions and stabilize them in

been substantially studied in coordination chemistry various oxidation state and increased their antimicro-

due to their ease of synthesis, effectively steric, bial activity depending on dipole moment, solubility,

their enzymatic action, and cell permeability.^[3–5]Schiff

base complexes have been utilized as model for

biological procedures and as a catalytic for oxidation

Supporting information for this article is available on the

WWW under https://doi.org/10.1002/cbdv.202100365

and polymerization of organic compounds.^[6]Schiff

Chem. Biodiversity 2021, 18, e2100365

base may be bi. di, tri or polydentate chelates together

Results and Discussion

with metal ions through nitrogen and oxygen donor

atoms formulation steady complexes.^[7–13]

The new prepared metal complexes of Fe(III), Cu(II),

Gemifloxacin (GMFX) is an artificial wide ranging Zn(II) and Zr(IV) with GMFX-en are stable, colored, and

antimicrobial compound versus both Gram-positive non-hygroscopic in nature. The complexes were solv-

and negative bacteria.^[14]GMFX implicates an effective able in dimethylformamide and dimethyl sulfoxide.

antibacterial activity against respiratory tract The exquisite physical features and distinctive data of

pathogens.^[14]GMFX can be interrogated with aliphatic the synthesized ligand and its metal complexes were

or aromatic amines supply a Schiff base.^[14,15]In the measured (Table 1). The results of the chemical analysis

literature, an attention has been paid to fluoroquino- showed that all the complexes were air stable at the

lones Schiff base complexes with transition metals.^[16]room temperature. At room temperature, the molar

On the other hand, the heavy metals and antibiotics conductance of GMFX-en in the free state is

could produce polluting molecules and induce phyto- 17.50 Ω^{À 1}mol^{À 1}cm², and the same data for metal

toxicity for crops. So, it is important to evaluate the complexes has been reported to range from 100.82 to

phytotoxic effect of these new synthesized complexes 270.69 Ω^{À 1}mol^{À 1}cm². Conductance statistics demon-

for determining the possibility for their using in strate that the metal complexes were electrolyte in

agropharmaceutical industry.^[17]

contrast with GMFX-en.^[16,18]

In view of the above consideration, the goal of this

study was to evaluate the impact of certain elements,

such as Fe(III), Cu(II), Zn(II) and Zr(IV) on the efficiency

FT-IR Spectral Studies

of gemifloxacin in new form as GMFX-en (Figure 1). Table 2 lists the positions of the significant FT-IR bands

The compounds were investigated by elemental of Schiff base and its metal complexes. FT-IR of the

analysis, molar conductance measurements, FT-IR, UV/ complexes (Figure S1) is compared with those of

VIS, ¹H-NMR and thermal analyses. Whereas the GMFX-en orderly to mark the spot of assortment that

antimicrobial activity was assessed towards phytopa- possibly implicated in chelation. The GMFX-en spec-

thogenic fungi: Monilinia fructicola (G. Winter) Honey, trum revealed that the bands were obscured speci-

Penicillium digitatum (Pers.) Sacc., and Colletotrichum alized for the bands of NH₂group for ethylene diamine

acutatum J.H. Simmonds and three bacterial species: and C=O of pyridone group of GMFX. Registered a

Clavibacter michiganensis corrig. (Smith) Davis et al., strong band at 1634 cm^{À 1}refers to the C=N and it is

Xanthomonas campestris Pammel (Dowson) and Bacil- determined to condensation of the amino group with

lus megaterium (Pammel) Dowson. Phytotoxicity assay a pyridone group, as a result of the formation of Schiff

was performed against Solanum lycopersicum L. (toma- base linkage.^[19]Complexes spectra involved bands

to) and Lepidium sativum L. (cress).

expand from 3428–3433 cm^{À 1}generated from the

vibration of OÀ H signalizing the subsistence of water

molecules in all complexes.^[13]The influence of oxygen

of carboxylate group with metal ions has been

registered by the obscurity of the band at 1725 cm^{À 1}

Figure 1. Structure of GMFX-en (4E,4’E)-4,4’-(ethane-1,2-diyldiazanylylidene)bis{7-[(4Z)-3-(aminomethyl)-4-(methoxyimino)pyrrolidin-

1-yl]-1-cyclopropyl-6-fluoro-1,4-dihydro-1,8-naphthyridine-3-carboxylic acid}.

Chem. Biodiversity 2021, 18, e2100365

and presence of new band around 1635 cm^{À 1}in the

entire complexes spectra.^[20]Subsistence of the asym-

metric stretching vibration band in 1634–1636 cm^{À 1}

region and ν_symin the region of 1356–1388 cm^{À 1}with

Δν>200 cm^{À 1}for the ligated carboxylate group

indicated the carboxylate group mono dentate inter-

acting through one of oxygen atoms.^[13,21]Further-

more, the shift of the characteristic band of azome-

thine group from 1522 cm^{À 1}to 1547 cm^{À 1}in all

certain complexes spectra specified participation of

C=N group in interaction with metal ions. New bands

in the regions 467–470 cm^{À 1}and 420–432 cm^{À 1},

matched to ν (MÀ O) and ν (MÀ N) vibration supporting

the pointed out the mode of coordination (Fig-

ure 2).^[22,23]

Electronic Spectra and Magnetic Studies

The assignments of the spotted electronic absorption

bands of the GMFX-en and its metal complexes,

magnetic data, and molar absorptivity (ϵ) of the

established complexes are indexed in Table 3. Actually,

the occurrences of diverse bands in the electronic

spectra of GMFX-en display bands at 237, 268 and

345 nm (Figure 3) that may be referred to π-π* and

n-π* transitions, respectively.^[24–26]The electronic

spectrum of the complex A exhibits an L!M (C.T) at

450 nm.^[27]This complex presumably has octahedral

configuration, and geometry is confirmed by meas-

ured magnetic moment (μ_eff=1.80 B.M.).^[28–30]For

complex B electronic spectrum exhibits an L!M (C.T)

charge transfer band at 480 nm.^[31]The band detected

at 580 nm for copper complex may be referred to

2

²B_1g! E_gtransition.^[32,33]The Cu(II) complex’s mag-

netic moment (1.70 B.M.) is very similar to the spin

value (1.73 B.M.) predicted for one unpaired electron,

indicating an octahedral geometry. The electronic

spectrum of complexes C and D exhibit an M!L (C.T)

charge transfer bands at 445 and 424 nm. In the light

of these results, these complexes presumably have

octahedral configurations.^[34,35]

¹H-NMR Spectra

1

The H-NMR spectra of the ligand and its complexes

were registered to emphasize the binding of the Schiff

base to the metal ions (Table S1, Figure 4).^[36,37]The

complexes did not detect the signal at 11 ppm

(COOH) in the GMFX-en spectrum, asserting that the

ligand was deprotonated and chelated with metal

ions.^[38]Furthermore, owing to the existence of water

1

molecules in the complexes, the H-NMR spectra for

Chem. Biodiversity 2021, 18, e2100365

complexes indicate a novel peak in the range 4.20–

4.55 ppm. Matching the GMFX-en Schiff base’s key

peaks to their complexes, The spectra of complexes

with the chemical change induced by Schiff base

binding to the metal ion contain all of the peaks of the

free ligand.^[39–41]

Thermal Analyses (TG and DTG) and Thermodynamic

Parameters

The stoichiometry of the resultant volatile decaying

components as well as the properties of the com-

plexes was studied employing thermal analyses. The

TG decay stages with the temperature maximum and

weight loss for the complexes are indexed in Table 4

and presented graphically in Figure 5. TG data for

GMFX-en manifested one stage of decaying with

°

temperature maxima 171, 338 and 532 C correspond-

ing to the loss of 18C₂H₂+2HF+2NH₃+6NO+2N₂

with total weight loss amounted to 96.90% (calc.

97.02%). The TG curve of complex A exhibited two

°

stages from 25 to 132 C and 132 to 680 C. The initial

°

stage of decay, with a maximum temperature of 69 C

with a mass loss of 11.20% (calc. 11.18%) may be

assigned to the removal of seven non- absorbed

coordinated water molecules. The last step showed

mass loss of 76.18% (calc. 76.41%), leaving the residue

°

observed over 670 C comparable to 0.5Fe₂O₃+5 C

related to the mass loss of 12.62% (calc. 12.41%). The

thermogram of complex B showed three stages in the

°

extent 25 to 137 C, 137 to 211 C and 211 to 650 C.

The first step due to the loss of 5H₂O lattice water with

mass loses 8.64% (calc. 8.26%). The second step from

°

137 up to 211 C display loss of 10 C₂H₂+2H₂O species

showed mass loss of 27.34% (calc. 27.19%). The last

step showed 6C₂H₂+CO+2HF+H₂S+4NO₂+2NH₃+

H₂species corresponding to 51.02% (calc. 51.73%)

°

leaving at the back the residue observed over 998 C

analogous to CuO+5 C related to mass loss of 13.00%

(calc. 12.82%). The complex C with in the temperature

°

range of 35 to 1000 C, the complex started decom-

°

position at 35 C in three distinct degradation phases.

°

Within the temperature range of 35 to 116 C, the first

step of decomposition corresponded to the loss of

3H₂O lattice water, resulting in a weight loss of 5.52%,

which agrees well with the calculated value of 5.25%.

The loss of Schiff base ligand molecules and coordi-

nated water molecules were assigned to the second

and third stages of decaying, which happened

°

between 116 and 650 C. It represented an 82.57%

weight loss, which is in good agreement with the

measured estimate of 83.35%, leaving at the back the

Chem. Biodiversity 2021, 18, e2100365

Figure 2. Structures of complexes A, B, C and D. Complex A: M=Fe(III), n=3 (FeC₃₈H₆₂F₂N₁₂O₁₅Cl₃). Complex B: M=Cu(II), n=2

(CuC₃₈H₅₄F₂N₁₂O₁₅S). Complex C: M=Zn(II), n=2 (ZnC₃₈H₅₄F₂N₁₂O₉Cl₂). Complex D: M=Zr(IV), n=2 (ZrC₃₈H₅₆F₂N₁₂O₁₃Cl₂).

°

residue observed over 999 C analogous to ZnO+3 C Antimicrobial Activity

related to mass loss of 12.00% (calc. 11.40%). Lastly,

Bactericidal Effect

the thermogram of complex D showed two steps in

°

the range 31–125 C and 125–660 C. The first step The results of antibacterial activity of the compounds

due to loss of 5H₂O uncoordinated water molecules were represented in (Figure 6) where all tested treat-

showed mass loss of 8.23% (calc. 8.26%). The second ments showed antibacterial effects against all tested

step showed loss of ligand molecules and coordinated bacteria especially at the higher tested concentrations.

water

molecules

corresponding

to

76.77% The highest antibacterial activity against C. michiga-

(calc.76.01%), leaving at the back the residue observed nensis was spotted in the case of GMFX-en and

°

over 660 C analogous to ZrO₂+4C related to mass complex B at 250 ppm (Figure 4). Regarding B. mega-

loss of 15.00% (calc. 15.72%).

terium and X. campestris the highest substantial effect

The kinetic factors such as (ΔH*), (E*), (G*), and (S*) was observed in the case of GMFX-en at 250 ppm

(Table 5) were graphically tested using the Coats- followed by the complex C at 250 ppm. On the other

Redfern and Horowitz-Metzger models equations (Fig- hand, complex D at 100 ppm showed the lowest effect

ure S2).^[42,43]Decomposition steps of activation ener- against B. megaterium and the same complex did not

gies were found to be in the range of 61.44– show any effect against X. campestris (Figure 6).

229.49 kJmol1.^[44,45]The positive sign of G* for com-

plexes demonstrated that the final residue’s free

energy was higher than that of the initial compounds,

Fungicidal Effect

implying that all decomposition steps were non- GMFX-en and its complexes were verified for fungici-

spontaneous. For the subsequent decomposition dal activity versus three serious phytopathogenic

stages of a given complex, the values of the activation, fungi. (P. digitatum, C. acutatum and M. fructicola)

ΔG*, increased significantly. This can be explained by compared to the positive control Azoxystrobin (Ta-

the fact that substantially rising the values of TΔS* ble 6). Results were demonstrated as fungal growth

from one stage to the next overrides the values of inhibition percentage. All examined substances clari-

ΔH*. Negative ΔS* values for the degradation process, fied antifungal effect in a dose dependent manner.

on the other hand, suggested that the activated The best antifungal effect against P. digitatum was

complex was more ordered than the reactants or that observed in the case of the complexes A and B at

the reaction was sluggish.^[46,47]

100 ppm. On the other hand, the lowest effect was

observed in the case of GMFX-en at 50 ppm compared

to Azoxystrobin. The highest activity against C. acuta-

tum was obtained in the case of Azoxystrobin followed

Chem. Biodiversity 2021, 18, e2100365

by complex C and GMFX-en at 100 ppm. The lowest

effect was obtained with GMFX-en 50 ppm. There is

no observed activity in the case of the complexes A

and B at both tested doses and the complex B 50 ppm

(Table 6). Furthermore, the complexes A, B, C and D at

100 ppm displayed the highest activity against M. fruc-

ticola. The lowest substantial effect has been observed

in the case of GMFX-en 50 ppm.

Mode of Action

The obtained microbicide effect of the studied com-

pounds might be due to the chemical structures of the

free ligand as well as the toxicity of the studied metal

ions.^[48,49]The principle of cell permeability of the

tested microorganisms clarified the enhanced antimi-

crobial activity of newly prepared metal chelates.^[50–52]

The polarity of metal ions can be reduced in particular

by partially sharing the positive charge with the parent

ligand’s donor groups and as a result of orbital overlap

with the ligand orbitals.^[53]On the other hand, the

chelation process enhancing the delocalization of

electrons above the chelate ring, which increase the

lipophilicity of the central ions. Because of the increase

in lipophilicity, the tested compounds were able to

penetrate deeper into the microorganism’s cells.^[54,55]

Phytotoxicity Effect

GMFX-en and its metal complexes were found to be

extremely phytotoxic to all of the plants examined,

S. lycopersicum and L. sativum, at all tested concen-

trations. In contrast to the control treatment, the

studied compounds have decreased the seeds germi-

nation and radical elongation for both tested plants.

Conclusions

This study investigated the new trace metal complexes

resulted from the interaction of some biological ions

such as Fe(III), Cu(II) Zn(II) and Zr(IV) with GFFX-en

which formed from the condensation reaction of

GMFX with ethylene diamine. Results showed that the

studied compounds have notable antimicrobial effect,

in a dose-dependent manner, against all tested

phytopathogens. The chemical structure of the free

ligand GMFX-en and its toxic nature are both

important factors in its promising antimicrobial activity

of some trace elements. In addition, the examined

compounds showed a significant phytotoxic effect on

the two tested plants. The potential antimicrobial and

Chem. Biodiversity 2021, 18, e2100365

Figure 3. Electronic absorption spectra for GMFX-en and its metal complexes.

Chem. Biodiversity 2021, 18, e2100365

Figure 4. ¹H-NMR spectra for GMFX-en and its metal complexes.

Chem. Biodiversity 2021, 18, e2100365

°

Table 4. The maximum temperature T_max( C) and weight loss values of the decomposition stages for GMFX-en and its metal complexes.

°

Compounds

Decomposition

T

_max( C)

Weight loss (%)

Lost species

Calc.

Found

GMFX-en

First step

Total loss

Residue

172, 338, 532

96.90

3.10

97.02

2.98

18C₂H₂+2HF+2NH₃+6NO+2N₂

2C

Complex A

First step

Second step

Total loss

Residue

First step

Second step

Third step

Total loss

Residue

First step

Second step

Third step

Total loss

Residue

First step

Second step

Total loss

Residue

69

337

11.18

76.41

87.59

12.41

8.26

27.19

51.73

87.18

12.82

5.25

38.88

44.47

88.60

11.40

8.26

76.01

84.28

15.72

11.20

76.18

87.38

12.62

8.64

27.34

51.02

87.00

13.00

5.52

38.70

43.78

88.00

12.00

8.23

76.77

85.00

15.00

7H₂O

16C₂H₂+2HF+CO+3HCl+2NH₃+5N₂+2.5H₂O

0.5Fe₂O₃+5C

5H₂O

10C₂H₂+2H₂O

6C₂H₂+CO+2HF+H₂S+4NO₂+2NH₃+H₂

Complex B

Complex C

Complex D

78

183,269

489

CuO+5C

3H₂O

93

316

472, 888

14C₂H₂+2H₂O

2HF+2HCl+2NH₃+4 N₂+2NO₂+CO

ZnO+3C

5H₂O

78

426

17C₂H₂+2HF+2NH₃+2HCl+H₂O+2NO₂+4N₂

ZrO₂+4C

phytotoxic effects of the new metal complexes combination was settling on a water bath, and then,

°

indicated that they could be used effectively in the temperature was lowered to 0 C. Yellowish-white

controlling both serious phytopathogens and harmful precipitate was collected out, washed multiple spells

weeds. In addition, the new innovative complexes by way of ethyl alcohol and desiccated below vacuum

could minimize the risk of appearance new resistant over CaCl₂.

strains of fungal, bacterial, or harmful weeds to the

traditional chemical compounds.

Preparation of Metal Complexes

The dark red [Fe(GMFX-en)(H₂O)₂]Cl₃·7H₂O complex A

Experimental Section

was prepared by blending 0.5 mmol (0.401 g) of

GMFX-en in 30 mL absolute ethyl alcohol with

0.5 mmol (0.081 g) of FeCl₃in 20 mL ethyl alcohol. The

Chemicals

Whole chemicals utilized were of analytical reagent blend was refluxed for 3 h and the subsequent was

grade and applied with no additional purifying. GMFX, collected out and desiccated under below vacuum

ethylene diamine (en), glacial acetic acid, absolute over CaCl₂. The brown, light brown, dark brown solid

ethanol, FeCl₃, CuSO₄·5H₂O, ZnCl₂·H₂O, ZrOCl₂·8H₂O complexes [Cu(GMFX-en)(H₂O)₂]SO₄·5H₂O complex B,

K₂Cr₂O₇, H₂SO₄and AgNO₃were purchased from [Zn(GMFX-en)(H₂O)₂]Cl₂·3H₂O complex C and [ZrO

Obour Pharmaceutical Industrial Company and Aldrich (GMFX-en)(H₂O)]Cl₂·5H₂O complex D were set in

Chemical Co. All glass were steeped overnight in identical technique qualified above by utilizing

chromic blend (Potassium dichromate+concentrated CuSO₄·5H₂O, ZnCl₂·H₂O and ZrOCl₂·8H₂O, respec-

sulfuric acid) swill completely with bidistilled water tively.

°

and desiccated in an oven at 100 C

Instruments

Preparation of GMFX-en Schiff Base (C₃₈H₄₄F₂N₁₂O₆)

The Carbon, Hydrogen and Nitrogen were done

An ethanolic solution was formed by combining 30 mL elemental experiments done via a Perkin Elmer 2400

gemifloxacin (2 mmol, 0.77 g) with 40 mL ethylenedi- CHN elemental analyzer. The proportion of metal ions

amine (1 mmol, 0.07 mL) and refluxing it for 4 h in the was gravimetrically measured by transforming solid

existence of 1 mL glacial acetic acid. The subsequent compounds into metal oxide and using the atomic

Chem. Biodiversity 2021, 18, e2100365

Figure 5. TGA and DTG diagram for GMFX-en and its metal complexes.

absorption technique utilizing a spectrometer model (D₆)DMSO as solvent. TG-DTG processes were con-

PYE-UNICAM SP 1900 fitted with the corresponding ducted out under N₂atmospheric conditions within

lamp. FT-IR spectra in KBr discs were enrolled in the the temperature range from ambient temperature to

area from 4000–400 cm^{À 1}with FT-IR 460 PLUS Spec- 1000 C using TGA-50H Shimadzu, the sample weight

°

trophotometer.¹H-NMR spectra have been registered was precisely weighted in an aluminum crucible. The

on Varian Mercury VX-300 NMR Spectrometer utilizing absorption spectra were registered as solutions in (D₆)

Chem. Biodiversity 2021, 18, e2100365

temperature employing Hg[Co(CSN)₄] as a calibrant.

Melting points have been transcribed on the Büchi

apparatus. All tests were done with freshly formulated

solutions at room temperature. The molar conductivity

of the ligand solutions and their complexes at 1×10^{À 3}

M in dimethylformamide was registered at room

temperature using CONSORT K410.

Antimicrobial Investigation

Antibacterial Activity Assay

Tested bacteria. The tested bacterial strains were

C. michiganensis, X. campestris and B. megaterium and

have been conserved as pure cultures in the collection

of the School of Agricultural, Forestry, Food and

Environmental Sciences (SAFE), University of Basilicata,

Potenza, Italy.

Bactericidal assay. The method of disc diffusion was

executed.^[56]Every strain’s bacterial suspension was

made in sterile Millipore H₂O, then inserted into soft

agar (0.7%) and regulated by UV-spectrophotometer

(DAS s.r.l., Rome, Italy) to 10⁸colony form units per

milliliter (CFU/mL). In a Petri dish (Φ 9 cm) filled with

10 mL of king B nutrient media (KB), 4 mL of each

tested bacterial suspension (10%) was poured. Blank

Discs (6 mm) (OXOID, Milan, Italy) were mounted on

KB Petri and 20 μL of each suspension was transferred

at concentrations: 100 and 250 ppm. Tetracycline

(160 μg/mL) was utilized as control. The antimicrobial

°

effect was determined after 24 h at 37 C, by measur-

ing the diameter of inhibition zones (mm). The experi-

ment was carried out in triplicate with�SDs.

Antifungal Activity Assay

Tested fungi. The tested phytopathogenic fungi were

M. fructicola, P. digitatum and C. acutatum. All tested

fungi were cultured on potato dextrose agar (PDA)

and were previously identified using morphological

and molecular methods. The amplicons were se-

quenced right away and matched to those in GenBank

using Simple Local Alignment Search Tool program

Figure 6. Antibacterial activities of GMFX-en and its metal

complexes. Values are recorded as the mean of diameter of

inhibition zones (mm) from three replicates�SDs. (*): statisti-

cally significant according to Tukey B test at P<0.05: (**): stat-

istically significant at P<0.01. Where A=Complex A, B=

complex B, C=complex C, D=complex D and Tetracy.: Tetracy-

cline at 1.6 mg/mL.

°

from 1990 (BLAST, USA). They were stored at 4 C as

pure cultures in the mycotheca of SAFE, University of

Basilicata, Potenza, Italy.

Fungicidal assay. The fungicidal effectiveness of the

DMSO. Room temperature magnetic susceptibility of studied compounds was determined by using incorpo-

the powdered samples was applied to the scientific ration assay,^[57]into PDA medium at two concentra-

magnetic balance of Sherwood utilizing Gouy at room tions (50 and 100 ppm). Every Petri dish was inocu-

Chem. Biodiversity 2021, 18, e2100365

Table 6. Antifungal activity of GMFX-en and its metal complexes.

Compounds

PPM

Fungal growth inhibition (%)

P. digitatum

C. acutatum

M. fructicola

GMFX-en

100

50

100

50

100

50

100

50

100

50

PDA

72.5�4.1^b

28.0�3.5^c

100.0�0.0^a

62.5�8.2^b

100.0�0.0^a

62.5�6.5^b

82.5�7.5^ab

61.0�2.5^b

86.0�6.1^ab

52.0�4.3^bc

0.0�0.0^d

62.5�2.8^b

22.5�3.7^d

0.0�0.0^e

45.0�2.1^c

0.0�0.0^e

77.5�6.5^b

32.5�4.5^c

0.0�0.0^e

92.5�1.3a

65.0�6.2^b

36.0�3.1^c

87.5�5.7^a

62.5�6.2^b

85.0�1.8^a

75.0�5.2^ab

82.5�5.4^a

62.5�4.2^b

85.0�5.8^a

67.5�2.2^b

0.0�0.0^d

Complex A

Complex B

Complex C

Complex D

Cont.

Azoxy.

+ve cont.

58.1�1.2b

45.3�2.1^c

Values were recorded as the mean inhibition percentage of fungal growth (three replicates)�SDs. Values followed by different

letters in each vertical column were significantly different according to Tukey B test at P<0.05. Cont (PDA): potato dextrose agar

(negative control). Azoxy.: Azoxystrobin (Positive control at 0.8 μL/mL).

lated with a fungal disk (0.5 cm), from 96 h fresh nated seeds were enumerated. The experiment was

°

culture. All plates were incubated at 22 C for 96 h and repeated three times, and the germination index (GI)

the antifungal effect was estimated by measuring the was measured as following:

diameter of the fungal mycelium (mm) compared to

control (only PDA+Fungi). The percentage of myce-

SG:t � RE:t

G:I: ð%Þ ¼

� 100

(2)

lium growth inhibition (MGI) was calculated according

to Zygadlo et al.,^[58](Formula 1) compared to Azox-

ystrobin (0.8 μL/mL):

SG:c � RE:c

All values are expressed as the mean�SDs.

GI: germination index; SG.t: average germination

(Treated seeds); RE.t: average radical elongation

(treated seeds); SG.c: average germination (control);

RE.c: average radical elongation (control). The statisti-

ðGC À GTÞ

MGI ð%Þ ¼

� 100

(1)

GC

Where MGI is the percentage of mycelium growth cal analysis was performed using SPSS software with

inhibition, GC is the average diameter of fungal post hoc Tukey B (P<0.05).

mycelium in PDA (control), and GT is the average

diameter of fungal mycelium on the treated PDA dish.

Data Availability Statement

Phytotoxicity Assay

The data that support the findings of this study are

The potential phytotoxic efficacy of the tested com- available from the corresponding author upon reason-

pounds was investigated on tomato and garden cress able request.

plants.^[59,60]Seeds were sterilized for 1 min in 3% H₂O₂

then were immerged in Millipore H₂O. Seeds were

shaken gently for 2 h in dH₂O (control) or the above-

mentioned treatments. The tested concentrations

Acknowledgments

were 2000, 1000, 500, 250 and 100 ppm. All seeds The authors gratefully acknowledge Zagazig Univer-

were placed in Petri dishes with filter paper (9 cm, sity, Egypt for the support of this research work.

Whatman No. 1). Two mL of dH₂O or various com-

pounds was added to each plate then sealed with

Parafilm. All Petri dishes were incubated for 4 days in

°

growth chamber at 24 C and 60% humidity. The

radical length was calculated in cm and the germi-

Chem. Biodiversity 2021, 18, e2100365

acid (L) metal complexes’, Appl. Organomet. Chem. 2020,

Author Contribution Statement

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Received May 12, 2021

Accepted June 29, 2021

Article Doi

Biochemical Characterization, Phytotoxic Effect and Antimicrobial Activity against Some Phytopathogens of New Gemifloxacin Schiff Base Metal Complexes

DOI: 10.1002/cbdv.202100365

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