Fabrication and Characterization of S-Benzyldithiocarbazate Schiff Base Microcrystals by a Reprecipitation Method for Enhanced Antibacterial Activity

In this study, the microcrystallization of the 2-thiophenecarboxaldehyde Schiff base of S-benzyldithiocarbazate (i.e. benzyl (2E)-2-[(thiophen-2-yl)methylidene]hydrazine-1-carbodithioate [TASBnDTC]) was fabricated by a reprecipitation method in an organic solvent-water system using different crystallization parameters, including temperature and the concentration of the target compound. The size, anisotropy, crystalline phase, and surface morphology of the TASBnDTC microcrystals were characterized by dynamic light scattering and scanning electron microscopy. The stability of the Schiff base microcrystals was also evaluated. Different sizes of surfactant-dispersed TASBnDTC microcrystals (1505, 2194, and 2447 nm) were fabricated from three different concentrations of the Schiff base (0.001 M, 0.002 M, and 0.003 M, respectively) in an acetone-water system. The TASBnDTC microcrystals were also evaluated by X-ray powder diffraction and were found to differ slightly in molecular form but were otherwise similar, irrespective of the different TASBnDTC concentrations. The synthesized Schiff bases and their microcrystals were also screened for their antibacterial activities against four different gram-positive and gram-negative bacterial strains (Escherichia coli, Bacillus subtilis, Pseudomonas aeruginosa, and Staphylococcus aureus) using the agar well diffusion method. The growth inhibition was enhanced by 8.0 to 10.75 mm against the four bacteria by TASBnDTC microcrystals compared to the bulk molecular form, which showed no inhibitory activity at all. However, the inhibition was less that that achieved with the standard streptomycin antibiotic, which gave zones of inhibition of 18.0 to 23.0 mm against the four bacterial strains. Overall, the Schiff base microcrystals show potential for use in various biological applications. They also have potential physical and optical applications due to their high surface-to-volume ratio and the molecular alignment on the surface of the microcrystals.

Antibacterial activity; Nanoscience; Organic microcrystal; Reprecipitation method; Schiff base

In recent times, nanotechnology has been integrated in many parts of our everyday lives. The fabrication of organic and inorganic nano- and microcrystals is one area of growing interest from the perspectives of both fundamental science and applications (Poerwadi et al., 2020; Strokova et al., 2020). However, organic microcrystals have received little attention so far because of difficulties with their characterization and the thermal instability of most of the organic compounds.

Organic microcrystals with sizes of several tens of nanometers to the sub-micrometer crystal size have been fabricated by either “top-down” approaches, such as milling methods (Li et al., 2020), or by “bottom-up” approaches, such as reprecipitation (Kasai et al., 1992; 2012) and supercritical fluid crystallization (SCFC) (Komai et al., 1999). In the “top-down” approaches, the microcrystals are fabricated by size reduction techniques, whereas in the “bottom-up” approaches, the microcrystals are fabricated from atoms or molecules by synthetic chemistry or self-assembly processes. The most common “bottom-up” approach is the so-called reprecipitation method first reported by Kasai et al. (1992). The reprecipitation method has many advantages, as it is simple, quick, and inexpensive. In addition, it is applicable to many kinds of organic compounds. However, this method cannot be applied in the case of water-soluble organic dyes or barely water-soluble organic compounds, such as pigments. However, Tripathy et al. (1998) recently introduced a new approach for fabricating well-defined organic microcrystals through electrostatic self-assembly. Similarly, Baba et al. (2000) introduced a novel microwave-irradiation process to improve the conventional reprecipitation method.

Most of the current research focuses on the preparation of free-standing organic microcrystals, and far less work has been conducted on inorganic semiconductor microcrystals. These are organic microcrystals with sizes ranging from a few tens to a few hundred nanometers, and they have received substantial attention in the past few decades (Tripathy et al., 1998; Oikawa et al., 2000). This interest arises largely from the observation that their unique size-dependent optical, electronic, and chemical properties differ from those of either their single molecules or their bulk crystals (Jortner and Rao, 2002; Chiang et al., 2012; Indrasti et al., 2020). The excitement of nanoscience and the potential applications of these microcrystals are now driving intensive research activities, and organic nano-/microcrystals are now finding a broad range of applications in the areas of lithography (Magdassi and Moshe, 2003), organic photoconductors (Miyashita et al., 2008) and pharmaceutical formulations for drug delivery and biomedical applications (Kumar and Lal, 2014).

Typically, organic microcrystals have a high surface energy and large curvature, which changes the chemical nature of the organic molecules on the surface and renders the small organic microcrystals thermodynamically unstable. For this reason, obtaining organic nano-/microcrystals even with sizes below 100 nanometers is very difficult, and this size is still much larger than that of semiconductor or noble metal nano-/microcrystals (Li et al., 2017). Therefore, much research has focused on the use of additives, capping agents, or inorganic or polymer matrices to control the size and shape of organic microcrystals (Abyan et al., 2005). One approach has been the use of Schiff bases containing imine group (?CH=N?) as these bases have structural peculiarities and wide pharmacological and biological applications (Fasina et al., 2013; Mondal et al., 2017).

Schiff bases are usually formed by the condensation of a primary amine with a carbonyl compound (an aldehyde or ketone) in the presence of an acid catalyst. A careful survey of the literature shows only limited reports on the fabrication of Schiff-base micro-scale crystals by reprecipitation methods (Mahajan et al., 2017; Patil et al., 2017; Kumar et al., 2019). In recent years, interest has grown in the chemistry of S-alkyl/aryl dithiocarbazates owing to their potential biological activities and their great behavior as intermediates in organic reactions (Malik et al., 2020). To the best of our knowledge, the fabrication of this type of Schiff base by a reprecipitation method has not been reported previously. Therefore, this paper summarizes the details of a preliminary investigation on the fabrication, characterization and antibacterial studies of a S-benzyldithiocarbazate Schiff base and its microcrystals.

This study provides some new insight into the microcrystallization of Schiff base compounds by the reprecipitation method using an organic solvent-water system. Adjusting the concentration of the target compound in the organic solution and the temperatures of the microcrystallization process was confirmed as a feasible and practical way to control the size and quality of the organic microcrystals. The rate and efficiency of dispersion of the organic solvent in water govern the microcrystal size. These insights into microcrystallization by the reprecipitation method would allow further development of organic microcrystal fabrication in the Rayleigh regime with fully controlled properties. Varying the concentration of TASBnDTC with acetone at 0.001, 0.002, and 0.003 M yielded microcrystals of the compound with the same crystal structure. Interestingly, this preliminary study showed that these Schiff base microcrystals have slight antibacterial activities against four strains of bacteria (inhibition zone ranges of 8.0 to 10.75 mm), whereas the bulk molecular form has no antibacterial activity at all. Therefore, the Schiff base microcrystals warrant further studies by varying the moiety of the Schiff base as well as exploring its minimum inhibition concentration. These studies are currently underway in our laboratories.

        We are grateful to the Chemical Sciences [FIC Grant No. UBD/RSCH/1.4/FICBF(b)/2020/024], the Environmental and Life Sciences, Geosciences and Centre for Advanced Material and Energy Sciences, Universiti Brunei Darussalam for the necessary support in carrying out the research work.

Abyan, M., Bertorelle, F., Fery-Forgues, S., 2005. Use of Linear Polymers to Control the Preparation of Luminescent Organic Microcrystals. Langmuir, Volume 21(13), pp. 6030–6037

Ali, N.H.S.O., Hamid, M.H.S.A., Putra, N.A.A.M.A, Adol, H.A., Mirza, A.H., Usman, A., Siddiquee, T.A., Hoq, M.R., Karim, M.R., 2020. Efficient Eco-Friendly Syntheses of Dithiocarbazates and Thiosemicarbazones. Green Chemistry Letters and Reviews, Volume 13(2), pp. 129–140

Baba, K., Kasai, H., Okada, S., Oikawa, H., Nakanishi, H., 2000. Novel Fabrication Process of Organic Microcrystals using Microwave-Irradiation. Japan Journal of Applied Physics, Volume 39(12A), pp. 1256–1258

Becker, R., Liedberg, B., Käll, P.O., 2010. CTAB Promoted Synthesis of Au Nanorods–Temperature Effects and Stability Considerations. Journal of Colloid and Interface Science, Volume 343(1), pp. 25–30

Burda, C., Chen, X., Narayanan, R., El-Sayed, M.A., 2005. Chemistry and Properties of Nanocrystals of Different Shapes. Chemical Reviews, Volume 105(4), pp. 1025-1102

Chiang, H., Iimori, T., Onodera, T., Oikawa, H., Ohta, N., 2012. Gigantic Electric Dipole Moment of Organic Microcrystals Evaluated in Dispersion Liquid with Polarized Electroabsorption Spectra . The Journal of Physical Chemistry, Volume 116(14), pp. 8230–8235

Dalavi, D.K., Bhopate, D.P., Bagawan, A.S., Gore, A.H., Desai, N.K., Kamble, A.A., Mahajan, P.G., Kolekar, G.B., Patil, S.R., 2014. Fluorescence Quenching Studies of CTAB Stabilized Perylene Nanoparticles for the Determination of Cr(VI) from Environmental Samples: Spectroscopic Approach. Analytical Methods, Volume 6(17), pp. 6948–6955

Fasina, T.M., Ejiah, F.N., Dueke-Eza, C.U., Idika, N., 2013. Substituent Effect on the Antimicrobial Activity of Schiff Bases Derived from 2-Aminophenol and 2-Aminothiophenol. International Journal of Biological Chemistry, Volume 7, pp. 79–85

Indrasti, N.S., Ismayana, A., Maddu, A., Utomo, S.S., 2020. Synthesis of Nano-silica from Boiler Ash in the Sugar Cane Industry using the Precipitation Method. International Journal of Technology, Volume 11(2), pp. 422–435.

Jahangirian, H., Haron, M.J., Ismail, M.H.S., Rafiee-Moghaddam, R., Afsah-Hejri, L., Abdollahi, Y., Rezayi, M., Vafaei, N., 2013. Well Diffusion Method for Evaluation of Antibacterial Activity of Copper Phenyl Hydroxamate Synthesized from Canola and Palm Kernel Oils. Digest Journal of Nanomaterials and Biostructures, Volume 8(3), pp. 1263–1270

Jortner, J., Rao, C.N.R., 2002. Nanostructured Advanced Materials. Perspectives and Directions. Pure and Applied Chemistry, Volume 74(9), pp. 1491–1506

Kasai, H., Nakanishi, H., Oikawa, H., 2012. Application 42 - Fabrication Technique of Organic Nanocrystals and Their Optical Properties and Materialization. In: Nanoparticle Technology Handbook, Hosokawa M., Nogi, K., Naito, M., Yokoyama, T. (eds.), Elsevier, Amsterdam, pp. 596-601

Kasai, H., Nalwa, H.S., Oikawa, H., Okada, S., Matsuda, H., Minami, N., Kakuta, A., Ono, K., Mukoh, A., Nakanishi, H., 1992. A Novel Preparation Method of Organic Microcrystals. Japan Journal of Applied Physics, Volume 31(8A), pp. 1132–1134

Komai, Y., Kasai, H., Hirakoso, H., Hakuta, Y., Katagi, H., Okada, S., Oikawa, H., Adschiri, T., Inomata, H., Arai, K., Nakanishi, H., 1999. Preparation of Organic Microcrystals using Supercritical Fluid Crystallization Method Preparation of Organic. Japan Journal of Applied Physics, Volume 38(1A), pp. 81–83

Kumar, R., Lal, S., 2014. Synthesis of Organic Nanoparticles and their Applications in Drug Delivery and Food Nanotechnology: A Review. Journal of Nanomaterials & Molecular Nanotechnology, Volume 3(4), pp 1–11

Kumar, S., Mittal, S.K., Kaur, N., 2019. Enhanced Performance of Organic Nanoparticles of a Schiff Base for Voltammetric Sensor of Cu(II) Ions in Aqueous Samples. Analytical Methods, Volume 11(3), pp. 359–366

Li, W., Baren, J.V., Berges, A., Bekyarova, E., Lui, C.H., Bardeen, C.J., 2020. Shaping Organic Microcrystals using Focused Ion Beam Milling. Crystal Growth and Design, Volume 20(3), pp. 1583–1589

Li, Z.Z., Liang, F., Zhuo, M.P., Shi, Y.L., Wang, X.D., 2017. White-Emissive Self-Assembled Organic Microcrystals. Small, Volume 13(19), pp. 1–6

Magdassi, S., Moshe, M.B., 2003. Patterning of Organic Nanoparticles by Ink-Jet Printing of Microemulsions. Langmuir, Volume 19(3), pp. 939–942

Mahajan, P.G., Kolekar, G.B., Patil, S.R., 2017. Recognition of D-Penicillamine using Schiff Base Centered Fluorescent Organic Nanoparticles and Application to Medicine Analysis. Journal of Fluorescence, Volume 27(3), pp. 829–839

Malik, M.A., Lone, S.A., Wani, M.Y., Talukdar, M.I.A., Dar, O.A., Ahmad, A., Hashmi, A.A., 2020. S-Benzyldithiocarbazate Imine Coordinated Metal Complexes kill Candida albicans by causing Cellular Apoptosis and Necrosis. Bioorganic Chemistry, Volume 98, pp. 1–11

Miyashita, Y., Baba, K., Kasai, H., Nakanishi, H., Miyashhita, T., 2008. A New Production Process of Organic Pigment Nanocrystals. Molecular Crystals and Liguid Crystals, Volume 492(1), pp. 268/[632]-274/[638]

Mondal, S., Mandal, S.M., Mondal, T.K., Sinha, C., 2017. Spectroscopic Characterization, Antimicrobial Activity, DFT Computation and Docking Studies of Sulfonamide Schiff Bases. Journal of Molecular Structure, Volume 1127, pp. 557–567

Oikawa, H., Fujita, S., Kasai, H., Okada, S., 2000. Electric Field-Induced Orientation of Organic Microcrystals with Large Dipole Moment in Dispersion Liquid. Colloid and Surfaces A: Physicochemical and Engineering Aspects, Volume 169(1-3), pp. 251–258

Patil, K.S., Mahajan, P.G., Patil, S.R., 2017. Fluorimetric Detection of Sn²⁺ Ion in Aqueous Medium using Salicylaldehyde Based Nanoparticles and Application to Natural Samples Analysis. Spectrochimica Acta-Part A: Molecular and Biomolecular Spectroscopy, Volume 170, pp. 131–137

Poerwadi, B., Kartikowati, C.W., Oktavian, R., Novaresa, O., 2020. Manufacture of a Hydrophobic Silica Nanoparticle Composite Membrane for Oil-Water Emulsion Separation. International Journal of Technology, Volume 11(4), pp. 364–373

Rahman, S.N.S.A., 2018. Fabrications of Polycyclic Aromatic Hydrocarbon and Schiff Bases Nanocrystals by Reprecipitation Method. Master’s Thesis, MSc Program, Universiti Brunei Darussalam, Brunei Darussalam

Raj, S.S.S., Yamin, B.M., Yussof, Y.A., Tarafder, M.T.H., Fun, H.-K., Grouse, K.A., 2000. Trans-Cis S-Benzyl Dithiocarbazate. Acta Crystallographica Section E Crystal Structure Communications, Volume 56(10), pp. 1236–1237

Strokova, V.V, Baskakov, P.S., Ayzenshtadt, A.M., Nelyubova, V.V., 2020. Creation of Biocidal Coatings using the Stabilization of Silver Nanoparticles in Aqueous Acrylic Dispersions. International Journal of Technology, Volume 11(1), pp. 5–14

Tripathy, S.K., Katagi, H., Kasai, H., Balasubramanian, S., Oshikiri, H., Kumar, J., Oikawa, Hidetoshi, O., Okada, S., Nakanishi, H., 1998. Self Assembly of Organic Microcrystals 1: Electrostatic Attachment of Polydiacetylene Microcrystals on a Polyelectrolyte Surface. Japan Journal of Applied Physics, Volume 37(2-3B), pp. 343–345

Yariv, I., Lipovsky, A., Gedanken, A., Lubart, R., Fixler, D., 2015. Enhanced Pharmacological Activity of Vitamin B12 and Penicillin as Nanoparticles. International Journal of Nanomedicine, Volume 10(1), pp. 3593–3601