Kazuyuki Yano

Elucidation of adsorption mechanism of bone-staining agent alizarin red S on hydroxyapatite by FT-IR microspectroscopy

To elucidate adsorption mechanism of alizarin red S (ARS), which is often used for staining bones in histology, adsorption of ARS on hydroxyapatite, Ca10(PO4)6(OH)2 (HAP), was investigated by a batch method, compared with alizarin, phenols, and benzenesulfonates. We found that ionized 1-, 2-OH groups (1-, 2-O(-)) of ARS can be electrostatically bound to Ca2+ on HAP, but that the 3-SO3(-) group of ARS hardly participates in adsorption on HAP. ARS-adsorbed HAP (ARS-HAP) in dark reddish violet was also prepared and analyzed by FT-IR microspectroscopy to gain structural information on bonding between ARS and HAP. The obtained spectrum, which was converted to difference spectra, indicated a single band of nu(C=O) at 1627 cm(-1) and two types of symmetric C=O stretching bands of nu(s)(C=O) + nu(C=C) at 1345 cm(-1) and nu(s)(C=O) + delta(O-C=C) at 1272 cm(-1). These bands imply the existence of a salt form in ARS-HAP via 1-, 2-OH groups of ARS. As a result of the existence of a chelate form in ARS-HAP via 1-OH and 9-C=O groups of ARS, two bands of nu(C=C) + nu(C=O) at 1572 cm(-1) and nu(C=O) + nu(C=C) at 1537 cm(-1) were also observed. In addition, ARS was almost desorbed from colored ARS-HAP at 50 degrees C by using neutral phosphate buffer to recover slightly pale pinkish HAP, or De-ARS-HAP. The desorbed ARS belongs to ARS previously adsorbed on HAP by salt formation, while the remaining color on De-ARS-HAP indicates ARS still adsorbed on HAP by chelate formation. Consequently, we elucidated two adsorption mechanisms of ARS on HAP: The major adsorption is salt formation made up with 1-, 2-O(-) of ARS and Ca2+ on HAP, and the minor adsorption is chelate formation made up with 1-O(-) and 9-C=O of ARS and Ca2+ on HAP.

Evaluation of glycogen level in human lung carcinoma tissues by an infrared spectroscopic method

Glycogen levels in the tissue samples obtained from carcinomas and normal sections of human lungs (26 patients) were studied by measuring the infrared band intensity at 1045 cm(-1) due to glycogen. As an internal standard peak, the band at 1545 cm(-1) (amide II) was chosen, and the ratios of these band areas (A1045/A1545) were compared with histological classification and differentiation of tumors. The glycogen level in the carcinoma tissues was significantly higher than that in the normal tissues (P < 0.01, n = 26). Further, the ratio of amounts of glycogen in the carcinomas and in the normal tissues for adenocarcinoma was higher than that for squamous cell carcinoma (P < 0.01). The increased degree of differentiation of the squamous cell carcinomas appeared to be correlated with an increase in the glycogen level. These results suggest that comparison of glycogen levels in the tumor and normal section of human lung may be used as a differentiating parameter for abnormality and histological classification of tumors. The present Fourier transform-infrared spectroscopy (FT-IR) method may become of wide application for studying various tissue samples.

Estimation of glycogen levels in human colorectal cancer tissue: relationship with cell cycle and tumor outgrowth

Abstract: In this study, we quantitatively measured glycogen levels in tissue samples obtained from tumors, regions adjacent to tumor, and regions of normal colorectum to determine whether the levels were related to cell cycle and cancer growth. Glycogen levels were analyzed in relation to histopathological factors, (tumor size and stage of disease) and cell cycle progression. The glycogen level was found to be highest in the cancer tissue, lower in normal tissue, and lowest in the adjacent tissue. The difference in glycogen level between the cancer tissue and the other two regions was significant (P < 0.05). There was a negative correlation between glycogen level and tumor size, but it was not significant. The level of glycogen in cancer tissues decreased as the stage of the disease progressed, but a significant difference was not found between stages. There was a negative correlation between the glycogen level and the proliferation index. There was a positive correlation between the glycogen level and the proportion of cancer cells in G1 phase, while there was a negative correlation with S and G2M phases. Glycogen levels were highest in cancers with a high proportion of cells in G1, and decreased with progression to S phase. It may be that glycogen is utilized in the progression to S phase, and the cancer tissues are supplied with glycogen from the tumors themselves as well as their adjacent tissues. Cancer growth may be inhibited by artificial control of the glycogen level in the G1 phase of cancer cells.

Applications of Fourier transform infrared spectroscopy, Fourier transform infrared microscopy and near-infrared spectroscopy to cancer research

Glycogen levels in human lung and colorectal cancerous tissues were measured by the Fourier transform (FT-IR) spectroscopic method. Reliability of this method was confirmed by chemical analyses of the same tissues used for the FT-IR spectroscopic measurements, suggesting that this spectroscopic method has a high specificity and sensitivity in discriminating human cancerous tissues from noncancerous tissues. The glycogen levels in the tissues were compared with the clinical, histological and histopathological factors of the cancer, demonstrating that glycogen is a critical factor in understanding the biological nature of neoplastic diseases. Furthermore, direct measurement of a very small amount of tissue by a FT-IR microscope suggested that it could be used as a diagnostic instrument for various tissue samples obtained via a fine needle biopsy procedure. The progressive alterations in rat mammary gland tumors were investigated by a near-infrared (NIR) spectrometer with a fiber optic probe. A lipid band due to the first overtone of n-alkane was used to quantitatively evaluate malignant changes in the tumors. NIR spectroscopy may offer the potential for non‒invasive, in vivo diagnosis of human cancers.

Effect of Repeated Application of C60 Combined with UVA Radiation Onto Hairless Mouse Back Skin

Abstract Repeated application of buckminsterfullerene (C60) toluene solution combined with ultraviolet A (UVA) radiation onto hairless mouse back skin resulted in enhancing formation of erythema as an acute disease but induced no carcinoma on the skin. This result is probably due to insolubility of C60 in the living cells to generate little singlet oxygen 1O2 by UVA radiation.

N-nitroso compounds. Part 3. Hydrolysis of N,N′-dimethyl-N′-(p-nitrophenyl)-N-nitrosourea in aqueous basic solution. Effect of a cationic micelle on the hydrolysis of substituted N-nitrosourea

The hydrolysis of the title N-nitrosourea has been studied kinetically in various amine buffers (pH 8–11) at 36.8 °C. The reaction proceeds through hydroxide ion attack at the carbonyl carbon to form a tetrahedral intermediate, which collapses to an arylcarbamate ion, releasing an N-nitrosamino fragment. The effect of the buffer concentration on the hydrolysis rate is explained in terms of general base catalysis. Nucleophilic attack of unhindered amines at the carbonyl carbon also occurs under the hydrolytic conditions. With bulkier amines, however, the concurrent nucleophilic reaction is excluded because of steric hindrance around the carbonyl group. The micellar effect on the hydrolysis rate has also been investigated using cetyltrimethylammonium bromide (CTABr). The rate enhancement of the hydrolysis by the CTABr micelle is shown to be six-fold at pH 8.05. However, with an alternative N-nitrosourea bearing a labile hydrogen on N′, a much larger micellar catalysis was observed for the elimination of the hydrogen in the presence of OH–.

Determination of Glycogen Levels in Human Lung Cancer and Normal Tissues by Fourier Transform Infrared Spectrometry

Glycogen levels in the carcinoma and normal tissues of human lungs were studied by the FT-IR method using the integrated area ratios of glycogen to protein (A1045/A1545). Further amounts of glycogen and protein contained in the same tissues were measured by an anthrone method and by using a dotMETRIC™ kit, respectively. Ratios of glycogen to protein obtained here were compared with the FT-IR spectroscopic data in relation to abnormality and histological classification of the tumors indicating a good correlation between them. The present results show that the FT-IR method is very reliable and that may be of wide application for studying glycogen levels in various tissues as well as diagnosis purpose.