Quantitative analysis of ethanol–methanol–water ternary solutions using Raman spectroscopy

Raman spectroscopy was used for rapid in-situ measurement of alcohols in ethanol-methanol-water ternary systems. Mass fractions of the individual components were determined using calibration curves for binary systems of ethanol-water, methanol-water, and ethanol-methanol. Calibration curves were constructed by calculating the ratio of the Raman peak intensity of a component and that of an external standard (acetonitrile). Assuming additivity of the spectra, simultaneous equations were written, and mass fractions of ethanol, methanol, and water in the ternary solutions were determined by solving the system of equations through calculating an inverse matrix. The relative errors between the mass fractions obtained from the Raman spectra and those obtained from mass measurements were <0.6%.</p

Production of Ros during glucose metabolism.

The amount of ROS produced by (A) normoxically and (B) hypoxically cultured cells was estimated by ROS assay kit. Cancer cells were harvested after 3-day culture and normal cells were harvested after 5-day culture. The error bars represent standard deviation. Significant difference (*, p <0.05; **, p <0.001) in the amount of other acids produced between normoxic and hypoxic conditions, according to the paired t-test.</p

The effects of rotenone on the production of lactic acid and other acids during glucose metabolism by hypoxically cultured HSC-3.

The amount of other acids produced by hypoxically cultured HSC-3 cells was calculated by subtracting the amount of lactic acid produced from total acid production, which was obtained with pH-stat system. The number is the ratio of lactic acid [%]. The error bars represent standard deviation.</p

Schematic diagram showing the metabolic effects of the environmental oxygen concentration revealed in the present study.

Schematic diagram showing the metabolic effects of the environmental oxygen concentration revealed in the present study.</p

S1 Data -

(XLSX)</p

Production of lactic acid and other acids produced during glucose metabolism.

The amount of other acids produced by (A) normoxically and (B) hypoxically cultured cells was calculated by subtracting the amount of lactic acid produced from total acid production, which was obtained from Fig 1. The error bars represent standard deviation. Significant difference (*, p <0.05) in the amount of acid other than lactic acid between normoxic and hypoxic conditions, according to the paired t-test.</p

Glucose-derived acid production.

Glucose-derived acid production by (A) normoxically and (B) hypoxically cultured cells was measured as the amount of NaOH added by the pH-stat system under normoxic or hypoxic conditions. Cancer cells were harvested after 3-day culture and normal cells were harvested after 5-day culture. The error bars represent standard deviation. Significant difference (*, p <0.05) between the normoxic and hypoxic conditions, according to the paired t-test.</p

Cell growth in normoxic and hypoxic conditions.

Cells that had been pre-cultured in normoxic (A) or hypoxic (B) conditions were cultured under normoxic or hypoxic conditions. The error bars represent standard deviation. *Significant difference in the number of cells (p <0.05) between the normoxic and hypoxic culture conditions, according to the paired t-test.</p

Non-Enzymatic DNA Cleavage Reaction Induced by 5-Ethynyluracil in Methylamine Aqueous Solution and Application to DNA Concatenation

<div><p>DNA can be concatenated by hybridization of DNA fragments with protruding single-stranded termini. DNA cleavage occurring at a nucleotide containing a DNA base analogue is a useful method to obtain DNA with designed protruding termini. Here, we report a novel non-enzymatic DNA cleavage reaction for DNA concatenation. We found that DNA is cleaved at a nucleotide containing 5-ethynyluracil in a methylamine aqueous solution to generate 5′-phosphorylated DNA fragment as a cleavage product. We demonstrated that the reaction can be applied to DNA concatenation of PCR-amplified DNA fragments. This novel non-enzymatic DNA cleavage reaction is a simple practical approach for DNA concatenation.</p></div

Image2_Anti-cancer activity of Chaga mushroom (Inonotus obliquus) against dog bladder cancer organoids.jpeg

Despite its disadvantages, chemotherapy is still commonly used for the treatment of bladder cancer (BC). Developing natural supplements that can target cancer stem cells (CSCs) which cause drug resistance and distant metastasis is necessary. Chaga mushrooms are popular to have several health-promoting and anti-cancer potentials. Organoid culture can recapitulate tumor heterogeneity, epithelial environment, and genetic and molecular imprints of the original tissues. In the previous study, we generated dog bladder cancer organoids (DBCO) as a novel experimental model of muscle-invasive BCO. Therefore, the present study aimed to examine the anti-tumor potentials of Chaga mushroom extract (Chaga) against DBCO. Four strains of DBCO were used in the present study. Treatment with Chaga inhibited the cell viability of DBCO in a concentration-dependent way. Treatment of DBCO with Chaga has significantly arrested its cell cycle and induced apoptosis. Expression of bladder CSC markers, CD44, C-MYC, SOX2, and YAP1, declined in the Chaga-treated DBCO. Also, Chaga inhibited the phosphorylation of ERK in DBCO. Expression of downstream signals of ERK, C-MYC, and Cyclins (Cyclin-A2, Cyclin-D1, Cyclin-E1, and CDK4) was also inhibited by Chaga in DBCO. Interestingly, the combinational treatment of DBCO with Chaga and anti-cancer drugs, vinblastine, mitoxantrone, or carboplatin, showed a potentiating activity. In vivo, Chaga administration decreased tumor growth and weight of DBCO-derived xenograft in mice with the induction of necrotic lesions. In conclusion, Chaga diminished the cell viability of DBCO by inhibiting proliferation-related signals and stemness conditions as well as by arresting the cell cycle. Collectively, these data suggest the value of Chaga as a promising natural supplement that could potentiate the effect of adjuvant chemotherapy, lower its adverse effects, and thus, limit the recurrence and metastasis of BC.</p