Synthesis, Raman, FT-IR, NMR spectroscopic characterization, antimicrobial activity, cytotoxicity and DNA binding of new mixed aza-oxo-thia macrocyclic compounds

Series of new mixed aza-oxo-thia macrocyclic ligands 1,9(2,6)-ditriazina-2,8,10,16-tetraaza-3,7,11,15-tetraoxo-5,13-dithia-cyclohexadecaphan-1⁴,9⁴-diphenyl (L₁); 1,10(2,6)-ditri azina-2,9,11,18-tetraaza-3,8,12,17-tetraoxo-5,6,14,15-tetrathia-cyclooctadecaphan-1⁴,10⁴-diphenyl (L₂); 1,11(2,6)-ditriazina-2,10,12,20-tetraaza-3,9,13,19-tetraoxo-6,16-dithia-cyclocosa-phan-1⁴,11⁴-diphenyl (L₃); 1,12(2,6)-ditriazina-2,11,13,22-tetraaza-3,10,14,21-tetraoxo-6,7,17,18-tetrathia-cyclodocosaphan-1⁴,12⁴-diphenyl (L₄) were synthesised. The structural features of the compounds have been studied by elemental analyses, Mass, FT-Raman, FT-IR, ¹H and ¹³C NMR spectroscopy. The antimicrobial activities of the ligands were evaluated using disk diffusion method in dimethyl sulfoxide (DMSO) as well as the minimal inhibitory concentration (MIC) dilution method, against several bacteria and yeast cultures. The obtained results from both methods were assessed in side-by-side comparison with commercial antibacterial and antifungal agents. In most cases, the compounds show strong antifungal activity in the comparison tests. Cytotoxic activities of the ligands against two different human cancer cell lines, stomach (23132/87) and lung (A549) were determined by MTT assay. DNA fragmentation assay tested cell lines were used to analyze the DNA ladder formation which is a characteristic of apoptotic cell death. The binding of the ligands with calf thymus DNA (CT-DNA) has also been investigated by absorption spectroscopy.     

Stable carbamazepine colloidal systems using the cosolvent technique

The aim of this paper was to prepare stable carbamazepine nanosuspensions containing 10 mg/ml drug concentration by screening different polymers. Stable formulations were created by the cosolvent technique with polyethylene glycol (PEG-300) and water as the cosolvents. Rapid growth of long needle shaped CBZ crystals was observed in the absence of polymer. The presence of hydroxypropyl methylcellulose (HPMC) or methylcellulose (MC) inhibited crystal growth and the mean particle sizes were in the range 10–20 nm. Simultaneous presence of HPMC and polyvinylpyrrolidon (PVP PF17) polymers in CBZ suspensions enhanced the overall stability of the formulations. The additional stability improvement was attributed to the interaction between the polymers by the formation of hydrogen bonds. Suspension stability was evaluated over 5 months where the particle size remained constant. FT-Raman studies showed the existence of form I within the stable CBZ suspensions.     

Quantitative analysis of mannitol polymorphs. FT-Raman spectroscopy

Mannitol is a polymorphic excipient which is usually used in pharmaceutical products as the beta form, although other polymorphs (alpha and delta) are common contaminants. Binary mixtures containing beta and delta mannitol were prepared to quantify the concentration of the beta form using FT-Raman spectroscopy. Spectral regions characteristic of each form were selected and peak intensity ratios of beta peaks to delta peaks were calculated. Using these ratios, a correlation curve was established which was then validated by analysing further samples of known composition. The results indicate that levels down to 2% beta could be quantified using this novel, non-destructive approach. Potential errors associated with quantitative studies using FT-Raman spectroscopy were also researched. The principal source of variability arose from inhomogeneities on mixing of the samples; a significant reduction of these errors was observed by reducing and controlling the particle size range. The results show that FT-Raman spectroscopy can be used to rapidly and accurately quantitate polymorphic mixtures.     

胱氨酸、半胱氨酸及乙酰半胱氨酸的傅里叶变换拉曼光谱研究

目的为研究蛋白质等高分子化合物的分子结构奠定基础。方法拉曼光谱可作为研究分子晶体中分子振动和晶格振动的重要手段。本文采用傅里叶变换拉曼光谱仪对胱氨酸、半胱氨酸及乙酰半胱氨酸进行光谱检测,对这3种氨基酸的光谱差异进行分析,并对其分子振动光谱与结构特征的相关性进行了分析和探讨。结果与结论获得了3种氨基酸的傅立叶拉曼特征谱信息,为进一步研究它们的SERS提供了基础。     

Particle Size Dependent Molecular Rearrangements During the Dehydration of Trehalose Dihydrate-In Situ FT-Raman Spectroscopy

Purpose. (1) To characterise the different phases of trehalose using FT-Raman spectroscopy. (2) To monitor the changes in the structure of trehalose dihydrate on isothermal heating at 80°C. Methods. Different phases of trehalose were prepared and FT-Raman spectra obtained. Trehalose dihydrate was sieved to <45 m and >425 m particle size fractions and FT-Raman spectra were obtained at various time intervals during heating at 80°C. Results. During heating at this temperature, the spectra of a <45 m particle size fraction showed a loss of peak resolution with time and after 210 minutes resembled the spectrum of amorphous trehalose prepared by lyophilisation, indicating that the material was rendered amorphous by heating. In contrast, spectra obtained from a >425 m particle size fraction altered with time and became characteristic of the crystalline anhydrate. The approximate kinetics of this transformation to the anhydrate were monitored by analysis of peak intensity ratios with time. A two stage rearrangement was indicated; some functional groups appeared to manoeuvre into the spatial arrangement found in the anhydrate initially before the rest of the ring structure relaxed into this conformation. This may be due to some parts of the molecule being immediately affected by the loss of the water molecules on dehydration prior to the subsequent reorientation of the entire molecule into the anhydrate crystal lattice. Conclusions. The <45 m particle size fraction becomes disordered on dehydration induced by heating at 80°C whilst the >425 m particle size fraction crystallises to the anhydrate under the same conditions.     

FT-Raman spectroscopy of calcium hydroxide medicament in root canals

AIM: To investigate chemical changes in calcium hydroxide introduced into human root canals as a medicament using Fourier transform-(FT) Raman spectroscopy. METHODOLOGY: Ten necrotic maxillary anterior teeth were selected in 10 patients. The teeth were divided into five treatment groups, according to the survey time. Root canal instrumentation was performed with hand instruments until the master apical file was size 40. Calcium hydroxide paste, in a 1 : 1.25 mixture by weight of powder and distilled water, was introduced directly into the root canal with a lentulo-spiral filler and then condensed with a finger plugger. The access cavity was sealed with a temporary dressing. After 2 and 4 days, then 2, 4 and 6 weeks, the calcium hydroxide paste was sampled with a K-file and then analysed using FT-Raman spectroscopy. The excitation source was an Nd : YAG laser with an excitation wavelength of 1064 nm. All spectra were taken with a laser power of 200 mW, 275-1185 scans, and 4 cm(-1) resolution. The conversion of calcium hydroxide to calcium carbonate was calculated on the basis of the spectral data obtained from the mixtures of both compounds. RESULTS: The calcium hydroxide paste in the apical region showed weak bands at 1088 and 284 cm(-1), in addition to bands associated with calcium hydroxide. The weak bands, assigned to calcium carbonate, became stronger with time. Calcium carbonate content increased rapidly in the first 2 days and then tended to increase slowly. Approximately 11% of the calcium hydroxide at the apical portion of the canal was converted to calcium carbonate after 6 weeks. However, little alteration of the paste was noticed in the samples from the middle portion of the canal. CONCLUSIONS: Calcium hydroxide medicament in root canals became transformed into calcium carbonate in the apical region within 2 days. Although the transformation continued with time, approximately 90% of the calcium hydroxide remained unchanged after 6 weeks.     

NaOCl effects on primary and permanent pulp chamber dentin

OBJECTIVES: The dentin quality of primary and permanent pulp chamber was inspected by Fourier-transformed Raman spectroscopy (FT-Raman) and scanning electron microscopy (SEM). Fragments of pulp chamber dentin were obtained from 20 human molar crowns (primary and permanent). METHODS: The fragments were assigned to 8 groups (n=5)-Primary teeth: G1, pulp chamber dentin; G2, pulp chamber dentin irrigated with NaOCl 1% (30min); G3, pulp chamber dentin irrigated with NaOCl 1% (30min) and etched by 35% phosphoric acid; G4, pulp chamber dentin etched by 35% phosphoric acid. Permanent teeth: G5, pulp chamber dentin; G6, pulp chamber dentin irrigated with NaOCl 1% (30min); G7, pulp chamber dentin irrigated with NaOCl 1% (30min) and etched by 35% phosphoric acid; G8, pulp chamber dentin etched by 35% phosphoric acid. The spectra were subjected to the Cluster analysis. The SEM images were scored. RESULTS: Inorganic content: There was a difference between primary and permanent dentin. The groups treated with NaOCl were statistically similar between them, but differed from the groups not treated. Organic content: There was no difference between primary and permanent dentin. The groups became similar after NaOCl and phosphoric acid treatments. The microscopic images showed the presence of calcospherites on permanent dentin and their absence on primary dentin. CONCLUSIONS: The NaOCl changed the inorganic content in both dentitions; regardless of the following phosphoric acid etching. However, the chemical changes caused by NaOCl were not detected by SEM when it was followed by etching.     

The effect of excitation laser power in Raman spectroscopic measurements of the degree of conversion of resin composites

ObjectivesTo evaluate the effect of excitation laser power in Raman spectrometry by comparing the spectra and the degree of conversion (DC) values obtained using excitation powers between 300 and 1000 mW.MethodsFive commercial and three experimental resin composites were light cured at 1200 mW/cm² for 10–20 s from a commercial blue-violet LED dental curing unit. Raman spectra were collected from composite specimens within 9 min after light-curing. The excitation laser (1064 nm) was focused on the spot of 0.4 mm in diameter. The following powers were used for specimen excitation (mW): 300, 400, 600, 800, and 1000. From Raman spectra, the DC values were calculated and compared among different laser powers. Also, vector-normalized Raman spectra collected using the lowest excitation power (300 mW) were compared to those collected using the maximum excitation power (1000 mW).ResultsVarying the excitation laser power between 300 and 1000 mW resulted in statistically significant differences in both the DC values and the intensity of particular spectral features. The effect of varying laser power on Raman spectra and obtained DC values was material-dependent. The DC values measured within an individual material using different laser powers varied between 3.2 and 7.2% (absolute DC difference). The spectral bands affected by variations in laser power were assigned to symmetric and asymmetric stretching of −CH₂ (2900-3100 cm⁻¹), symmetric stretching of aliphatic CC (1640 cm⁻¹) and scissoring of C–H (1458 cm⁻¹).SignificanceThe DC can be artificially elevated through increasing excitation laser power. This effect should be considered in Raman spectroscopic evaluations of DC in specimens during ongoing post-cure polymerization.     

Quantitative determination of diclofenac sodium and aminophylline in injection solutions by FT-Raman spectroscopy

The FT-Raman quantification of diclofenac sodium and aminophylline commercial injection solutions was performed. The efficiency of various spectra treatment procedures including classical univariate intensity ratio and multivariate partial least squares (PLS) and principal component regression (PCR) methods was compared. First, the calibration models were built using unnormalised spectra. Next, spectra normalised by the intensity of a selected band of CH₃CN added as an internal standard to the studied samples were utilised. To compare the predictive ability of the models constructed, the relative standard error of prediction (RSEP) was calculated. The errors found for multivariate calibrations were a few times smaller than those for the univariate ones. Usually, the most effective was the PLS method, for which RSEP values of the order of 1–2% for calibration and 2–3% for testing data sets were obtained.

Four commercial preparations of diclofenac sodium and one of aminophylline containing by weight, 2.4% of the active pharmaceutical ingredient (API) were quantified applying the developed models. Concentrations found from the Raman data analysis correlate with the declared values and the results of reference analyses. For the studied diclofenac sodium solutions they amount to 99.2–101.2% of the former and 101.2–102.4% of the latter quantities for the PLS models optimised for each medicine based on unnormalised spectra. These values for the aminophylline preparation were found to be 101.0 and 99.1%, respectively. It shows that the proposed procedure based on the chemometric treatment of FT-Raman spectra can be a fast and convenient alternative to the standard pharmacopoeial procedures of API quantification even in relatively diluted injection solutions.     

The chemical aspects of Raman spectroscopy: Statistical structure-spectrum relationship in the analyses of bioflavonoids

Raman spectroscopy has been accepted as a useful tool for the characterization of natural products. However, to identify a specific compound in a mixture sample of natural products using Raman spectra alone is highly challenging if not impossible. We demonstrated an effective solution to such issues using a method combining statistical Raman spectroscopy and Mass spectrometry. The method was validated with a successful application to the identification of the major anthocyanin components in a purple yam (Dioscorea purpurea) extract. Of particular interest is that statistical grouping of the bioflavonoid standards that formed the database of this study was found to correspond closely to the conventional chemical classification. An initial theory on the chemical aspects of Raman spectroscopy pertaining to the connectivity of Raman-active functional groups in bioflavonoids was developed based on the statistical correlation between chemical classification and Raman spectroscopy.