Key findings:

Key findings include: successful synthesis of two novel Schiff base compounds (A and B) from ceftriaxone and substituted benzaldehydes; in vitro evaluation of antimicrobial activities against gram-positive bacteria, Candida, and Aspergillus niger; improved antibacterial potency of some derivatives compared to ceftriaxone; and amoxicillin showing significant biological activity.

What is known and what is new?

The known aspect is the use of Schiff base compounds derived from cephalosporin antibiotics like ceftriaxone to enhance antimicrobial activity. The new contribution is the synthesis and characterization of two novel Schiff base compounds (A and B) from ceftriaxone and substituted benzaldehydes, and the evaluation of their in vitro antibacterial and antifungal activities against specific microorganisms, demonstrating improved potency compared to the parent drug.

What is the implication, and what should change now?

The implication of this study is that the synthesized Schiff base compounds derived from ceftriaxone have the potential to be developed as more potent antimicrobial agents. Changes needed include further optimization of the Schiff base structures, evaluation of their mechanism of action, and in-depth pharmacological studies to establish their clinical utility as novel antibacterial and antifungal therapeutics.

Ceftriaxone is  one of the types of antibiotic drug that returned to the cephalosporin/cephamycin family of beta lactam antibiotics [1]. Ceftriaxone is used for treatment for bacterial infections. Ceftriaxone, in vitro, has biological activity against different types of bacteria such as gram-positive and gram-negative aerobic bacteria [2], in addition to anaerobic bacteria. Ceftriaxone has the capacity to inhibit and kill different bacteria due to its ability to inter and block the form of the cell wall of bacteria [3]. When broken down by many types of beta-lactamases, ceftriaxone is stable, the beta-lactamases containing penicillinases, cephalosporinases, and extended-spectrum [4-5]. Ceftriaxone is the third generation of cephalosporins, and ceftriaxone has a wide range of effectiveness against many types of bacteria [6]. Compared to other drugs, it has a longer half-life and a big ability to penetrate the eyes, and ear [7-8]. Ceftriaxone has a wider and more potent spectrum of action against gram-negative bacteria compared to second generation drugs [9]. Ceftriaxone is often used as an antibacterial agent because of its notable efficacy against multi-drug-resistant Enterobacteriaceae, its relatively benign side-effect profile, and its extended half-life that permits the ease of daily or twice-day administration.

An azomethine or imine group is a characteristic of a class of chemicals known as Schiff bases, which Hugo Schiff, an Italian-German scientist, successfully synthesized in 1864 [10-11].

As part of Schiff's standard method, a carbonyl molecule and a primary amine were joined together using azeotropic distillation. To get an excellent yield, the reversible condensation reaction requires the elimination of the produced water. This is done by using a carbinolamine intermediate to promote the forward reaction. Schiff bases are special because they can be made in a lot of different ways [12]. This means that you can precisely control (i) the types of atoms that act as donors, (ii) the number of binding sites and their ability to form chelates, and (iii) the electronic and steric properties. Schiff bases are extensively used in organic compounds with diverse uses, such as serving as intermediates in organic synthesis, functioning as chemosensors and polymer stabilizers, acting as dyes and pigments in the food industry, playing a role in catalysis, and more [13].

In addition to exhibit a wide play activity of biological applications. The azomethine or imine group plays an essential role. Schiff base compounds have good medicine applications because Schiff bases have multi-active groups. These applications in many fields, such as biochemical processes and biological control. Schiff bases can chelate abilities due to the presence of azomethine and other coordinating groups [14].

The aim of this study was to synthesize Schiff base derivatives (C and D) and characterize these derivatives by spectroscopic methods. These derivatives of Schiff base were tested as antibacterials and antifungals and compared with ceftriaxone and amoxicillin drugs.

2.1 Synthesis of Schiff base derivatives (A and B)

Dissolve (0.001 mol, 0.14 g) of 4-dimethylaminobenzaldehyde and (0.001 mol, 0.14 g) of 4-chlorobenzaldehyde separately in 15 ml of absolute ethanol and (2–3) drops of glacial acetic acid. Subsequently, ceftriaxone (0.554 g, 0.001 mol) was added to each of these solutions for 4 hours. Following formation and filtering of the yellow precipitates, ethanol washing and vacuum drying take place [15-16]. ( Figure 1)

Figure 1: Chemical structures of derivatives A and B.

2.2 A study of the antibacterial activity of ceftriaxone and its synthetic modifications

The several bacterial strains, including Bacillus subtilis, and E. coli, were grown on Muller-Hinton agar plates using a sterile loop and streaking techniques, starting with the broth culture. Subsequently, a solitary well was generated in the agar. Each well received 50 and 100 μl of the corresponding dilution of ceftriaxone and produced derivatives, which were well absorbed. The dish was hermetically sealed and cultured at a temperature of 37 ºC overnight for additional testing on the next day [17].

2.3 Docking Experiment

The protein data bank (PDB code: 1MOQ) was used to get the crystallographic 3D structure of glucose-6-phosphate synthase (GP6 synthase). The compounds produced were displayed using ChemDraw Ultra (version 12.0) and stored in a sdf format file. The energy reduction was performed using Chem3D (version 12.0) and then converted to a pdb file. The docking procedure was executed using Autodock Tools (version 1.5.7) [18].The outcomes were saved in pdbqt file format subsequent to reducing the energies of both enzymes and ligands. The grid box was defined with dimensions of 60 × 60 × 60, and its center was positioned at coordinates 21.864 in the x dimension, 24.853 in the y dimension, and 9.8123 in the z dimension [19]. The program Discovery Studio was used to create visual depictions.

3.1 FTIR Characterization

The FTIR (cm^-1) of derivative (A): The C-H bond of aromatic rings appeared at 3043, while the C-H aliphatic at 2909 and 2842, and the carbonyl group of the azomethine appeared at 1644. The C-C of aromatic rings that appeared as sharp peaks 1597 [15, 20]. (Figure 2)

Figure 2: FTIR of derivative A1.

The FTIR (cm^-1) of derivative (B): The C-H bond of aromatic rings appeared at 3040, while the C-H aliphatic at 2922, and the carbonyl group of the azomethine appeared at 1652. The C-C of aromatic rings that appeared as sharp peaks 1599 [17, 21]. ( Figure 3)

Figure 3: FTIR of derivative A2.

3.2 Study of Docking

In the first stage of hexosamine biosynthesis, enzymes assist in the conversion of fructose-6-phosphate into glucosamine-6-phosphate, thus increasing the process. The GlcN-6-P is an important part of making uridine diphosphate N-acetyl glucosamine (UDP-NAG), which is a part of the peptidoglycan layer in microorganisms' cell walls [22]. The docking characteristics of the medication are shown in Figures 4 and 5. The ligand's 3D structures display a multitude of interactions, notably hydrogen bonding, as well as other interactions, including van der Waals and pi-alkyl. The computed values of -6.9 kcal/mol for ceftriaxone demonstrated that the affinity between the two compounds was comparable. The experimental findings further confirm the data that both compounds have antibacterial activity. Molecular docking analysis, the 3D structures of the ligand exhibit several interactions, namely hydrogen bonding, along with additional interactions such as van der Waals and pi-alkyl. The calculated values of -6.9 kcal/mol for ceftriaxone indicate that the affinity between the two molecules is similar. 

Figure 4: Docking of ceftriaxone drug against GP6 synthase.

Figure 5: Docking 3D of ceftriaxone drug against GP6 synthase.

3.4 Biological Activity 

In table 1, the derivative B has an increased effect on preventing bacterial growth in petri dishes. Derivative B had the most important effect on Bacillus subtilis microorganisms by (21 mm) zone inhibition. The parent drug has little impact on Enterococcus faecalis bacteria by (9 mm). The derivative B exhibits better biological activity compared to the standard drug and Azithromycin drug [23].

Table 1: Anti-bacterial activities for amoxicillin and parent drug (ceftriaxone) and derivatives synthesized (A and B).

In table 2, The most significant impact on Fungal was seen in Aspergillus niger due to derivative B. The derivative B has a higher level of biological activity compared to the standard drug.

Table 2: Anti-fungal activities for ceftriaxone and derivatives synthesized (A1 and A2).

Schiff-base compounds were synthesized from the ceftriaxone drug, and spectroscopic methods were used to determine their structures. The process of making it began with a reaction between ceftriaxone and the appropriate substituted benzaldehydes. The Schiff base compounds containing the ceftriaxone moiety have been evaluated in vitro for their antimicrobial activities against two types of bacteria: Gram-positive (Bacillus subtilis and Enterococcus faecalis) microorganisms and Candida and Aspergillus niger in different concentrations (50 and 100 μl). The results showed that some of these derivatives have good antibacterial activity compared to the parent drug, but amoxicillin has high biological activity.

1. Funding: No funding sources.

    

    

2. Conflict of interest: None declared.

    

    

3. Ethical approval: The study was approved by the Institutional Ethics Committee of University of Al-Qadisiyah.

    

1. Zeng, Linan, et al. "Safety of ceftriaxone in paediatrics: a systematic review." Archives of disease in childhood (2020). https://adc.bmj.com/content/archdischild/early/2020/03/06/archdischild-2019-317950.full.pdf. 

2. 2. Nakayama, Shu-ichi, et al. "New ceftriaxone-and multidrug-resistant Neisseria gonorrhoeae strain with a novel mosaic penA gene isolated in Japan." Antimicrobial agents and chemotherapy 60.7 (2016): 4339-4341..https://doi.org/10.1128/aac.00504-16. 

3. 3. Shimels, Tariku, Arebu I. Bilal, and Anwar Mulugeta. "Evaluation of ceftriaxone utilization in internal medicine wards of general hospitals in Addis Ababa, Ethiopia: a comparative retrospective study." Journal of pharmaceutical policy and practice 8 (2015): 1-8.https://link.springer.com/article/10.1186/s40545-015-0047-1. 

4. 4. Nasir, Nosheen, et al. "Risk factors for mortality of patients with ceftriaxone resistant E. coli bacteremia receiving carbapenem versus beta lactam/beta lactamase inhibitor therapy." BMC Research Notes 12.1 (2019): 1-5.https://bmcresnotes.biomedcentral.com/articles/10.1186/s13104-019-4648-7. 

5. 5. Kuang, Dai, et al. "Increase in ceftriaxone resistance and widespread extended-spectrum β-lactamases genes among Salmonella enterica from human and nonhuman sources." Foodborne Pathogens and Disease 15.12 (2018): 770-775.https://doi.org/10.1089/fpd.2018.2468. 

6. 6. Tamma, Pranita D., et al. "Molecular epidemiology of ceftriaxone-nonsusceptible Enterobacterales isolates in an academic medical center in the United States." Open forum infectious diseases. Vol. 6. No. 8. US: Oxford University Press, (2019).https://doi.org/10.1093/ofid/ofz353. 

7. 7. Pacifici, Gian Maria, and Giovanna Marchini. "Clinical pharmacology of ceftriaxone in neonates and infants: effects and pharmacokinetics." International Journal of Pediatrics 5.9 (2017): 5751-5778. https://scholar.archive.org/work/6lbgz3w4uzhqtcd74li5bdlf3a/access/wayback/http://ijp.mums.ac.ir/article_9101_1fa278382740927d39497e7d8203e0fc.pdf. 

8. 8. Jordan, Jennifer, R. Brigg Turner, and Teri Moser Woo. "DRUGS USED IN TREATING INFECTIOUS DISEASES." PHARMACOTHERAPEUTICS FOR ADVANCED PRACTICE NURSE PRESCRIBERS: 691.https://www.academia.edu/download/60383442/Pharmacotherapeutics_for_Advanced_Practice_Nurse_Prescribers__4t20190824-18614-1cockv1.pdf#page=710. 

9. 9. Vaidya, Ankur, and Shweta Jain. "Chemistry and Pharmacology of β-Lactam Analogs." Handbook of Research on Medicinal Chemistry: Innovations and Methodologies (2017): 339.https://books.google.com/books?hl=en&lr=&id=2GdQDwAAQBAJ&oi=fnd&pg=PA339&dq=Vaidya,+A.+and+S.+Jain,+Chemistry+and+Pharmacology+of+%CE%B2-Lactam+Analogs.+Handbook+of+Research+on+Medicinal+Chemistry:+Innovations+and+Methodologies,+2017:+p.+339.&ots=UcRVwa2fQP&sig=hud84kb48fi1_5HysIP056P5zOw. 

10. 10. Wei, Liang, and Chun‐Jiang Wang. "Synergistic catalysis with azomethine ylides." Chinese Journal of Chemistry 39.1 (2021): 15-24. https://doi.org/10.1002/cjoc.202000380. 

11. 11. Santos, Hugo, et al. "1, 3-Dipolar cycloaddition reactions of phthalic anhydrides with an azomethine ylide." Organic Chemistry Frontiers 2.6 (2015): 705-712.https://doi.org/10.1039/C5QO00062A. 

12. 12. Kumawat, Gopal Lal, et al. "Cyclic Voltammetric Studies of Biologically Active Azomethine 2’-Hydroxyacetophenone Sulfamethoxazole." Oriental Journal of Chemistry 35.3 (2019): 1117.https://pdfs.semanticscholar.org/6ae5/8010b8b00ef9212192306440f6e9252e9bd9.pdf. 

13. 13. Hossain, Md Saddam, et al. "Selected Schiff base coordination complexes and their microbial application: A review." Int. J. Chem. Stud 6.1 (2018): 19-31.https://www.researchgate.net/profile/Mdkudrat-E-Zahan/publication/322202143_Selected_Schiff_base_coordination_complexes_and_their_microbial_application_A_review/links/5a4b1f2da6fdcce197214841/Selected-Schiff-base-coordination-complexes-and-their-microbial-application-A-review.pdf. 

14. 14. Chaturvedi, D., and M. Kamboj. "Role of Schiff base in drug discovery research." Chem Sci J 7.2 (2016): e114.https://www.academia.edu/download/86704275/role-of-schiff-base-in-drug-discovery-research-2150-3494-1000e114.pdf. 

15. 15. Odabasoglu, Hakki Yasin, Taner Erdogan, and Fikret Karci. "Synthesis & characterization of heterocyclic disazo-azomethine dyes and investigating their molecular docking & dynamics properties on acetylcholine esterase (AChE), heat shock protein (HSP90α), nicotinamide N-methyl transferase (NNMT) and SARS-CoV-2 (2019-nCoV, COVID-19) main protease (Mpro)." Journal of Molecular Structure 1252 (2022): 131974.https://doi.org/10.1016/j.molstruc.2021.131974. 

16. 16. El-Atawy, Mohamed A., et al. "New nitro-laterally substituted azomethine derivatives; Synthesis, mesomorphic and computational characterizations." Molecules 26.7 (2021): 1927.https://doi.org/10.3390/molecules26071927. 

17. 17. Burlov, A. S., et al. "Synthesis, characterization, luminescent properties and biological activities of zinc complexes with bidentate azomethine Schiff-base ligands." Polyhedron 154 (2018): 65-76.https://doi.org/10.1016/j.poly.2018.07.034. 

18. 18. Erdoğan, Musa, et al. "Synthesis and characterization of some benzidine-based azomethine derivatives with molecular docking studies and anticancer activities." Chemical Papers 77.11 (2023): 6829-6847.https://link.springer.com/article/10.1007/s11696-023-02981-3. 

19. 19. Korkmaz, Adem, and Ercan Bursal. "Benzothiazole sulfonate derivatives bearing azomethine: Synthesis, characterization, enzyme inhibition, and molecular docking study." Journal of Molecular Structure 1257 (2022): 132641.https://doi.org/10.1016/j.molstruc.2022.132641. 

20. 20. Sahan, Fatma, et al. "New azo‐azomethine‐based transition metal complexes: Synthesis, spectroscopy, solid‐state structure, density functional theory calculations and anticancer studies." Applied Organometallic Chemistry 33.7 (2019): e4954.https://doi.org/10.1002/aoc.4954. 

21. 21. Alamro, Fowzia S., et al. "Experimental and theoretical investigations of three-ring ester/azomethine materials." Materials 15.6 (2022): 2312.https://doi.org/10.3390/ma15062312. 

22. 22. Mezoughi, Aysha. “The synthesis and evaluation of novel soluble lytic transglycosylase inhibitors”. Diss. Cardiff University, (2019).https://orca.cardiff.ac.uk/id/eprint/125559/. 

23. 23. Peiffer-Smadja, Nathan, et al. "In vitro bactericidal activity of amoxicillin combined with different cephalosporins against endocarditis-associated Enterococcus faecalis clinical isolates." Journal of Antimicrobial Chemotherapy 74.12 (2019): 3511-3514.https://doi.org/10.1093/jac/dkz388.