Charge-transfer chemistry of two corticosteroids used adjunctively to treat COVID-19. Part II: The CT reaction of hydrocortisone and dexamethasone donors with TCNQ and fluoranil acceptors in five organic solvents

Graphical abstract


Introduction
On December 9, 2019, a novel coronavirus (severe acute respiratory syndrome coronavirus 2, SARS-CoV-2) originated from a seafood market in Wuhan, China, rapidly becoming a destabilizing global pandemic. By early March 2020, after the SARS-CoV-2 outbreak had affected millions worldwide, the World Health Organization (WHO) termed the acute respiratory syndrome resulting from SARS-CoV-2 infection the COronaVIrus Disease 2019  and declared it a pandemic [1][2][3], become the biggest outbreak that the world ever seen [4]. COVID-19 has killed more than 6.15 million and infected more than 490 million people worldwide (as of April 1, 2022) [5]. Several pharmacological approaches have been implemented based on previous experience and evidence from prior outbreaks of respiratory-targeting viruses, such as SARS-CoV in China in 2003 and the MERS-CoV in the Middle East in 2012 [6,7]. These therapeutic approaches include corticosteroids, tocilizumab, azithromycin, lopinavir/ritonavir, remdesivir, chloroquine, and hydroxychloroquine. For more than 70 years, corticosteroids have been used in the treatment of inflammatory diseases, and oral infections due to their strong immunomodulatory and anti-inflammatory properties; in general, corticosteroids modulate the immune response to treat a wide variety of diseases by preventing and attenuating inflammation [8,9]. Corticosteroids are widely used adjunctively to treat critically ill patients with COVID-19 because they reduce inflammationinduced lung injury by suppressing lung inflammation [10][11][12][13][14]. Corticosteroids decrease patients' dependency on mechanical ventilation, the duration of shock on circulatory support, the length of intensive care unit (ICU) stays, and potentially even mortality.
Charge transfer (CT) or donor-acceptor complexations are interactions that involve transferring an electronically charged particle between two molecules, the one that donates the charge is commonly known as the electron-donating (donor; D) molecule, whereas the one that accepts the charge is commonly known as electron-accepting (acceptor; A) molecule. The transferring could be symbolized as (D ? A), and the generated complex by this interaction could be formulated as [D +Á A ÀÁ ] [20][21][22][23][24][25][26][27]. After Robert Mulliken introduced the concept of CT interaction in 1969, Roy Foster widely disseminated it, and since then, it has become well established that CT interactions involve the formation of a weak bond combined with a great color change. The strong color changes used to identify the resultant CT complex reflect a change in the UV/Vis spectrum of D and/or A molecules appearing in the form of either a new absorption band that did not exist in the spectra of the D and A molecules or increasing of the intensity of the already existing UV/Vis bands.
CT interactions researchers noted that the generated products possess unique chemical, physical, and biological properties via this type of interaction. Beneficiations of CT interactions are not limited to basic sciences (biology, biochemistry, chemistry, physics) but extended to applied sciences (medicine, pharmacology, technology, industry, material science, engineering) [21,. In the past years, we have been interested in examining the CT behavior of numerous biologically active donor molecules by reacting them with different organic and inorganic accepting molecules in different media . Since the onset of the SARS-CoV-2 pandemic, we shifted our focus from biologically active donor molecules to the CT properties of the pharmacological agents used to combat COVID-19 and their modifiability. We think obtaining new insights into the CT behavior of active compounds used in the treatments for COVID-19 will provide insight into the mechanisms underlying their effective properties and can help scientists alike to optimize and improve these treatment protocols and potentially even the vaccines (Janssen, Moderna, AstraZeneca, Pfizer-BioNTech). In a four-part work, we pioneered this new research effort with a comprehensive investigation into the CT chemistry of azithromycin, an antibiotic widely used in the treatment of COVID-19 [34][35][36][37].
As a continuation of these works, our newest series aims to furnish a big-picture perspective on the CT property of hydrocortisone and dexamethasone, corticosteroids widely used in the treatment of COVID-19. In part I [122], we described the CT interactions of these two corticosteroids with the 2,3-dichloro-5,6-dicyano-pbenzoquinone acceptor. In the current part of the work (Part II), we described the CT interactions of hydrocortisone and dexamethasone with two new accepting-molecules, 7,7,8,8-tetracyano quinodimethane (termed as TCNQ) and fluoranil (termed as TFQ) ( Fig. 2) in five organic solvents.

Reagents and solvents
The used solvents in this study: MeOH, EtOH, MeCN, CH 2 Cl 2 , and CHCl 3 were spectroscopic-grade solvents and obtained from Fluka (Lausanne, Switzerland). The investigated organic acceptors supplied by the Merck KGaA (Darmstadt, Germany) were 7,7,8,8tetracyanoquinodimethane (TCNQ) and fluoranil (TFQ). The examined donors obtained from Sigma-Aldrich (Saint Louis, MO, USA) were hydrocortisone (abbreviated here as D1) and hydrocortisone (abbreviated here as D2). All the donors and acceptors were obtained from the commercial sources at the highest purity available, which were ! 98 % HPLC for D1, 99 % HPLC for D2, 98 % for TCNQ, and 97 % for TFQ.

UV/Visible spectra
The UV/Visible spectrophotometry was applied to collect the UV/Visible spectra of the D1 and D2 soluble complexes with TCNQ and TFQ acceptors in the investigated solvents. These electronic spectra were helpful for:      (iii) Verify the wavelength (in nm) corresponding to the CT electronic absorption band (k CT ) for all soluble CT complexes.
To obtain the UV/Visible spectra, first, standard solutions of D1, D2, TCNQ, and TFQ were prepared in a specific solvent (MeOH, EtOH, MeCN, CH 2 Cl 2 , and CHCl 3 ) (5 Â 10 À4 M). Second, the D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ systems in each solvent were prepared by mixing equal amounts (1 mL) of the donor and acceptor standard solutions in a 5 mL volumetric flask and completing the volume up to the mark by the solvent. Finally, all the prepared systems were scanned using a Cary 7000 UV-vis-NIR Spectrophotometer from Agilent Technologies, Australia, in the 200-800 nm region. The electronic features of each CT system were obtained by comparing the UV/Visible spectra of the systems with those of the corresponding free components (D1, D2, TCNQ, and TFQ).

Infrared (IR) spectra
The IR spectrophotometry was applied to obtain the IR spectra of the D1 and D2 solid CT complexes with the acceptors in the investigated solvents. Observing and analyzing the IR spectra of the formed CT complexes were helpful for: i) Observing the changes in the characteristic IR spectral bands of free reactants after the complexation process. ii) Confirming the CT interaction between D1 and D2 with TCNQ and TFQ acceptors in each solvent. iii) Proposing the suitable mechanisms of the investigated CT interactions.
To obtain the IR spectra, concentrated solutions of D1, D2, TCNQ, and TFQ (2 mmol in 20 mL solvent) were prepared in a specific solvent (MeOH, EtOH, MeCN, CH 2 Cl 2 , and CHCl 3 ). Second, the concentrated D1 and D2 solutions were mixed with the corresponding acceptor concentrated solution to generate the D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ systems. These systems were wellstirred for $3 min at room temperature and left overnight. Third, the generated colored precipitates were removed from the beakers using filter paper, washed three times with the solvent, and ovendried. Finally, the solid CT complexes were scanned from 400 to 4000 cm À1 using an ALPHA Bruker Fourier-Transform Infrared Spectrometer from Bruker Optik GmbH, Germany. The vibrational features of each complex were obtained by comparing the IR spectra of the solid products with those of the corresponding free components (D1, D2, TCNQ, and TFQ).

Elemental analysis
A PerkinElmer Microanalyzer (model 2400 Series II CHNS; USA) determined the carbon, hydrogen, and nitrogen contents (%) for the D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ solid CT complexes. The elemental composition results were used to verify the molar ratio of the CT reaction between D1 and D2 with the investigated acceptors (TCNQ and TFQ).

Stoichiometric of the CT reaction in solution state
The stoichiometric of the CT reaction between D1 and D2 with the investigated accepting-molecules (TCNQ and TFQ) in the solution state was obtained using two well-known methods: the spectrophotometric titration method and Job's continuous variation method.

Free reactants
We found that D1 and D2 donor molecules need slight heat to be dissolved in MeCN, CH 2 Cl 2 , and CHCl 3 solvents, but in MeOH and EtOH solvents, they completely dissolved without any heat. D1 and D2 0 s solutions in all solvents were colorless. In contrast, the TFQ acceptor is completely soluble in all solvents without any heat, whereas the TCNQ acceptor is easily dissolved in MeCN  The TCNQ acceptor creates a light green solution in MeOH solvent. TCNQ in MeOH solvent behaves differently from the other solvents (MeCN, CH 2 Cl 2 and CHCl 3 ). The shape and intensity of the UV/Visible spectrum of the TCNQ in MeOH are different from those in the other solvents. Methanolic solution of TCNQ exhibits two intense broad bands that look like a plateau. These two bands centered at 339 and 394 nm. The band's intensity at 339 nm is higher than that at 394 nm. There is also a small broad absorption band found around 740 nm exists in the spectrum of the TCNQ in MeOH.  TCNQ solution in the MeCN, CH 2 Cl 2 , and CHCl 3 solvents displayed a single, strong, and intense absorption band. This brand has a small shoulder band in all solvents. The width of the band is approximately the same in all solvents, around $60 nm in width. The maximum of the band (k max ) is 394 nm in MeCN solvent and 400 nm in CH 2 Cl 2 and CHCl 3 solvents.
Also, the TFQ electronic absorption properties can be classified into three groups: (a) TFQ in the MeOH and EtOH solvents: TFQ acceptor creates a beige solution in MeOH and EtOH solvents. TFQ behaved similarly in these two solvents. It created two absorption bands. A sharp and very strong band appeared at 287 nm in both solvents, and this sharp band was coupled with a less intense broadband ranging from 330 to 485 nm. The shape of the broadband is slightly different; in MeOH solvent, it looks like a dome (k max = 362 nm), but in EtOH solvent, it looks like a plateau (k max = 340 nm).
(b) TFQ in the MeCN solvent: The TFQ acceptor forms a remarkable solution in MeCN solvent compared with its solution in other solvents. The color of TFQ's solution in MeOH, EtOH, CH 2 Cl 2 , and CHCl 3 solvents ranged from beige to light-yellow, but in MeCN solvent, TFQ gives a distinguish light-pink color. TFQ's solution in MeOH, EtOH, CH 2 Cl 2 , and CHCl 3 solvents absorbs until 485 nm, but in its acetonitrile solution, its absorption extended to 600 nm. Two absorption bands appeared in the spectrum of TFQ's solution in MeCN solvent. These bands were accumulated at 332 and 487 nm. Both bands were broad, but the band's intensity at 332 nm is 5-times that at 487 nm. The band at 332 nm was coupled with a small shoulder band at 287 nm. This shoulder appears as a very strong and sharp band in the spectrum of TFQ's solution in MeOH and EtOH solvents.
(c) TFQ in the CH 2 Cl 2 and CHCl 3 solvents: A single board absorption band was found in the UV/Visible spectra of TFQ's solutions in CH 2 Cl 2 and CHCl 3 solvents. This broad-      (Table 1).

CT complexes
Figs. 7 and 8 contain the UV/Vis spectra of the D1-TCNQ and D2-TCNQ complexes in the MeOH, MeCN, CH 2 Cl 2 , and CHCl 3 solvents alongside that of the free reactants, respectively. The corresponding spectra of the D1-TFQ and D2-TFQ complexes are presented in Figs. 9 and 10, respectively. Generally, D1 and D2 molecules have similar chemical structures. The main difference between the two molecules is that the D2 molecule possesses a CAF bond, whereas the D1 molecule does not. This structure similarity reflects the similarity of the chemical behavior of D1 and D2 molecules when complexed with TCNQ and TFQ acceptors in all solvents. In general, the generation of a CT complex is characterized by a pronounced change in the solution color during the mixing of the donor and the acceptor solutions in an appropriate solvent. This change in solution color is coupled with the change in the electronic spectra of the free donor or/and free acceptor. The spectral changes could be either a new absorption band that did not exist in the electronic spectrum of free donor and free acceptor, an increase in the intensity of the characteristic band referred to the donor or the acceptor, or both. Analysis of the UV/ Vis spectra of the D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ complexes presented in Figs. 7-10 revealed that the CT complexation of D1 and D2 with TCNQ acceptor in MeOH, MeCN, CH 2 Cl 2 , and CHCl 3 , and the CT complexation of D1 and D2 with TFQ acceptor in MeOH, EtOH, MeCN, CH 2 Cl 2 , and CHCl 3 strongly enhanced the intensity of the characteristic bands of free TCNQ and TFQ acceptors in the specific solvent.

Bandgap energies
Tauc's plot [123][124][125] was used to determine the minimum energy for an electron to jump from a valence band to a conduction band; bandgap energy (termed as E g ). Figs. 11 and 12 contain the Tauc's plots for the TCNQ and TFQ acceptors dissolved in the inves-tigated solvents, respectively. The derived values of E g are tabulated in Table 2. The E g value of the TCNQ and TFQ acceptors depends on the solvent's type, as indicated in Figs. 13 and 14.
The highest E g value was displayed by TCNQ solution dissolved in CH 2 Cl 2 solvent, where the methanolic solution of TCNQ had the lowest value of E g . Interestingly, the TFQ acceptor shows the same outcome. Its solution in CH 2 Cl 2 solvent had the highest E g value, whereas its solution in MeOH solvent had the lowest E g value. Tauc's Plots for the D1-TCNQ and D2-TCNQ complexes prepared in the investigated solvents are presented in Fig. 15; those for the D1-TFQ and D2-TFQ complexes are shown in Fig. 16. The derived E g values for D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ complexes are tabulated in Table 2. The complexation of TCNQ acceptor with D1 slightly increased the value of E g in all solvents, but when TCNQ was complexed with D2, the value of E g was slightly decreased in all solvents. In contrast, the complexation of TFQ acceptor with both D1 and D2 slightly decreased the value of E g in all solvents.

Stoichiometric of the CT interactions
The stoichiometric CT reactions between D1 and D2 with the investigated accepting-molecules (TCNQ and TFQ) in the solidstate were verified by the elemental analyses and spectrophotometric titration and Job's continuous analysis variation methods in the solution state. The elemental results (C%, N%, and H%) for the D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ solid CT complexes collected by a PerkinElmer Microanalyzer are listed in Table 3. The collected elemental results proposed that the CT interactions between D1 with TCNQ and TFQ and D1 with TCNQ and TFQ proceed at a 1:1 M ratio in all investigated solvents. The spectrophotometric titration method and Job's continuous variation method are the most common techniques used to verify the stoichiometry of a chemical reaction in solution stat using UV/Vis absorption spectra. Figs. 17 and 18 represent the composition of D1 and D2 with TCNQ and TFQ acceptors in the investigated solvents obtained by the spectrophotometric titration method, respectively. Figs. 19 and 20 represent the composition of D1 and D2 with TCNQ and TFQ acceptors in the investigated solvents obtained by Job's continuous variation method, respectively. All plots in Figs. 17-20 suggest that the CT interactions between D1 with TCNQ and TFQ and between D1 with TCNQ and TFQ in solution-state proceeds with a

Determining spectroscopic parameters
Four common spectroscopic parameters were determined. These parameters were the molar absorptivity (e max ), formation constant (K CT ), transition dipole moment (l eg ), and oscillator strength (f) [126][127][128]. The units of e max , K CT , and l eg parameters are (L mol À1 cm À1 ), (L mol À1 ), and (Debye), respectively. The e max , K CT , l eg , and f parameters were calculated for the D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ complexes in the investigated solvents to understand the CT characteristics from D1 and D2 molecules towards TCNQ and TFQ molecules. Graphical representation of the 1:  This outcome suggests a strong interaction between D1 and D2 molecules with TCNQ acceptors with relatively high probabilities for CT transitions [38,69,76,88,102,111,122]. A strong, positive linear correlation between l eg and f values with Pearson's correlation coefficients (r) lies in the range + 0.9968 À +0.9992 as depicted in Fig. 23. This high mutual relationship between the two parameters (l eg and f) indicates that the values of l eg increased as the values of f increased.

Infrared analysis
Infrared (IR) spectra of pure reactants (D1, D2, TCNQ, and TFQ) are shown in Fig. 24. The intense bands of un-complexed D1 molecule were located at 502 cm À1 s(CACAC), 566 d twi (CH 2 ), 636 d twi (- The very strong band observed at 1054 cm À1 (D2) and 1041 cm À1 (D1) originated from the stretching vibration of the CAO bond. The very strong band observed at 1660 cm À1 (D2) and 1642 cm À1 (D1) was originated from the stretching vibration of the C@O bond of the carbonyl group, whereas those appeared at 1713 cm À1 (D2) and 1704 cm À1 (D1) conjugated to the C@O bond of the carboxylic group. The band resonated at 1280 cm À1 in the IR spectrum of the D2 could be assigned to the m(CAF) vibrations [129]. Methyl (CH 3 ) and methylene (CH 2 ) groups in D1 and D2 molecules displayed several absorption bands. D1 and D2 molecules possess multiple CH 2 and CH 3 groups [eight CH 2 groups and two CH 3 groups in D1; five CH 2 groups and three CH 3 groups in D2], and these groups make the IR spectrum of D1 and D2 noise complicated, especially in the area from 700 to 1500 cm À1 . The bending vibrations of-CH 2 groups: d twi (CH 2 ), d wag (CH 2 ), d rock (CH 2 ), and d sci (CH 2 ) are responsible for the bands located at 566, 779, 1277, and 1351 cm À1 in the IR spectrum of D1, respectively. The corresponding wavenumbers for the D2 were 536, 761, 1239, and 1358 cm À1 , respectively [130,131]. The bending vibrations of ACH 3 groups: d twi (CH 3 ), d wag (CH 3 ), d rock (CH 3 ), and d sci (CH 3 ) are responsible for the bands located at 636, 890, 1372, and 1434 cm À1 in the IR spectrum of D1, respectively. The corresponding wavenumbers for the D2 were 612, 897, 1403, and 1448 cm À1 , respectively. The m(CH 2 ) and m(CH 3 ) vibrations create a broad absorption band with a medium-intensity accumulation at 2946 cm À1 (D2) and 2920 cm À1 (D1). The in-plane bending, outof-plane bending, and stretching vibrations generated from the OAH bonds in the D1 molecule created the bands at 685, 936, and 3414 cm À1 , respectively. The corresponding wavenumbers for the D2 were 690, 984, and 3393 cm À1 , respectively.
In the IR spectrum of pure TCNQ, the band at 2216 cm À1 , combined with a small broad shoulder at 2282 cm À1 , resulted from the  TCNQ and TFQ molecules are typical electron-accepting units because they contain strong terminal electron-withdrawing groups. When a molecule possesses terminal electronwithdrawing groups, they take the electron density from the molecule's core, making the core deficient in electron density and thirsty to accept electronic charges from another molecule (electron-donating molecule). The terminal electron-withdrawing groups in TCNQ are the four C"N groups and are the two C@O and four CAF groups in TFQ. These withdrawing groups greatly decrease the electron density of the TCNQ and TFQ's aromatic rings and increase the aromatic ring's need for electrons. When mixing solutions of D1 or D2 with solutions of TCNQ or TFQ, electron charges were transferred from the donating sites in the electrondonating molecule (D1 or D2) to the aromatic ring of the electron-accepting molecule (TCNQ or TFQ) [D1 ? TCNQ; D2 ? TCNQ; D1 ? TFQ; D2 ? TFQ]. This donating-accepting interaction affects the solution's color, the distinguish electronic spec- Table 4 The molar absorptivity (e max ), formation constant (K CT ), transition dipole moment (l eg ), and oscillator strength (f) for the D1 and D2 complexes prepared in the investigated solvents.

Solvent
D1-TCNQ complex D2-TCNQ complex  tral bands of the free reactants, and the distinguish IR spectral bands of the free reactants. The IR spectra displayed in Fig. 25 are for the solid D1-TCNQ, D2-TCNQ, D1-TFQ, and D2-TFQ complexes generated in different solvents. These IR spectra revealed several spectral changes in the distinguish IR bands of free reactants (D1, D2, TCNQ, TFQ) after the CT complexation. Here we mentioned three examples: i) The bands resulting from the C@O stretching vibrations of the carbonyl and carboxylic groups of free D1 and D2 shifted to lower frequencies after CT complexation with TCNQ and TFQ. From 1704-1642 cm À1 in the free D1 to around $1686-1630 cm À1 in the D1-TCNQ and D1-TFQ complexes. The same was detected for the D2; the bands were moved from 1713-1660 cm À1 to around $1700-1642 cm À1 in the D2-TCNQ and D2-TFQ complexes. ii). The CT complexation between TCNQ with D1 and D2 affected the intensity and position of the characteristic bands belonging to the m(C"N) in the free TCNQ. The position of the bands was moved from 2216 and 2282 cm À1 in the free TCNQ to around $2190 and 2210 cm À1 in its complexes with D1 and D2 (D1-TCNQ, D2-TCNQ).
iii) In the free TFQ, the m(CAF) vibrations resonated at 988 cm À1 as a very strong, intense band, but in the D1-TFQ and D2-TFQ complexes, this was appeared at around $1020 cm À1 as a medium intensity band.

Conclusions
Hydrocortisone (D1) and dexamethasone (D2) have received considerable attention because they are widely used to treat COVID-19. We aim to furnish a big-picture perspective on the charge-transfer (CT) properties of corticosteroids D1 and D2. In part I, we examined the CT property of D1 and D2 towards the DDQ accepting-molecule in five different organic solvents. This part (part II) described the CT reaction between D1 and D2 with two new organic accepting-molecules (TCNQ and TFQ) in five different organic solvents. Calculated values of oscillator strength (f) and transition dipole moment (l eg ) suggested that the interaction between the investigated corticosteroids with TCNQ acceptor is much stronger than their interaction with TFQ acceptor. IR measurements suggested that electron charges transferred from the donating sites in D1 and D2 to the aromatic ring of TCNQ and TFQ (n ? p* transitions).

Data availability
Data will be made available on request.

Declaration of Competing Interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Wavenumber (cm -1 )