Direct Evaluation of Molecular States of Piroxicam/Poloxamer Nanosuspension by Suspended-State NMR and Raman Spectroscopies

A nanosuspension of piroxicam (PXC) and poloxamer 407 (poloxamer) prepared by the wet milling method was directly evaluated at the molecular level from the viewpoint of both solution and solid phases. ¹³C solution-state NMR measurements revealed a reduction in the concentration of dissolved poloxamer in the nanosuspension. Furthermore, the fraction of dissolved poly­(ethylene oxide) (PEO) chain, which is the hydrophilic part of poloxamer, was higher than that of dissolved poly­(propylene oxide) (PPO) chain, the hydrophobic part. ¹³C suspended-state NMR and Raman spectroscopies detected both solid-state PXC and poloxamer involved in the nanoparticles. Interestingly, the coexistence of crystalline and amorphous PXC in the nanoparticle was demonstrated. The yellow color of the nanosuspension strongly supported the existence of amorphous PXC. Changes in the peak intensity depending on the contact time in the suspended-state NMR spectrum revealed that the PEO chain of poloxamer in the nanoparticle had higher mobility compared with the PPO chain. The PEO chain should project into the water phase and form the outer layer of the nanoparticles, whereas the PPO chain should face the inner side of the nanoparticles. Amorphous PXC could be stabilized by intermolecular interaction with the PPO chain near the surface of the nanoparticles, whereas crystalline PXC could form the inner core

Data_Sheet_1_Intraoperative intact parathyroid hormone monitoring and frozen section diagnosis are essential for successful parathyroidectomy in secondary hyperparathyroidism.docx

BackgroundTotal parathyroidectomy (PTx) is often performed to treat secondary hyperparathyroidism (SHPT). Successful PTx is essential to prevent recurrent and persistent SHPT because remnant parathyroid glands (PTGs) in the neck can be stimulated and may secrete excessive parathyroid hormone (PTH) in end-stage renal disease. However, to date, few studies have investigated factors contributing to successful PTx before the completion of surgery.Materials and methodsBetween August 2010 and February 2020, 344 patients underwent total PTx, transcervical thymectomy, and forearm autograft for SHPT at our institute. Factors contributing to successful PTx before the completion of surgery were investigated. Preoperative imaging diagnoses, including computed tomography, ultrasonography, technetium-99m methoxyisobutylisonitrile (99mTc-MIBI) scintigraphy, intraoperative intact PTH (IOIPTH) monitoring, and frozen section histologic diagnosis, were performed. Successful PTx was defined as intact PTH level 70% decrease in intact PTH levels measured 10 min after total PTx and transcervical thymectomy compared to intact PTH levels measured before skin incision. Logistic regression analysis was conducted to investigate factors contributing to PTx success.ResultsUnivariate analysis showed that the number of all PTGs identified preoperatively by imaging modalities and the specimens submitted for frozen section diagnosis, which surgeon presumed to be PTGs, were not significant factors contributing to successful PTx. However, multivariate analysis revealed that the number of PTGs identified by frozen section diagnosis (P ConclusionSufficient intact PTH level decrease observed on IOIPTH monitoring and the number of PTGs identified by frozen section diagnosis contributed to successful PTx for SHPT. IOIPTH monitoring and frozen section diagnosis are essential for achieving successful PTx for SHPT.</p

Comparison of the EAT between patients with and without receiving warfarin.

<p>The EAT was significantly prolonged in the group receiving warfarin.</p

Dose-dependent change in the dielectric permittivity in response to TF reagent and heparin.

<p>(A and B) Representative traces of the temporal change in the normalized dielectric permittivity (upper panel) and its derivative (lower panel) at 10 MHz in a healthy subject. (A) The TF reagent was serially diluted and added to CaCl₂. A lesser dilution of the TF reagent shifted the permittivity curve to the left in a dose-dependent manner, which means a larger amount of TF enhances the coagulation. The lower panel shows that the EAT, and the peak of the derivative also shifted to the left in a dose-dependent fashion. (B) A serial concentration of heparin is added to the citrated blood samples. A larger amount of heparin shifted the permittivity curve to the right in a dose-dependent fashion. (C) The EAT is normalized to the control conditions, and plotted against the serial dilution of the TF reagent. The normalized EAT shows a gradual shortening accompanied with a smaller dilution of the TF. (D) The EAT is normalized to the control sample, and plotted against the serial concentration of heparin, which shows dose-dependent prolongation. *, p <0.05 vs. control by paired <i>t</i> test.</p

Correlation between the EAT and PT, aPTT, and D-dimer.

<p>Scatter-plots are shown between the EAT and PT (A), aPTT (B), and D-dimer (C). All of them exhibited no statistically significant correlations.</p

Coagulation parameters in the DBCM and conventional assays in groups with different CHA₂DS₂-Vasc scores.

<p>The patients were classified into 3 groups according to their CHA₂DS₂-Vasc score (0–1, 2–3, and ≥4) for a comparison with the EAT (A), PT (B), aPTT (C) and D-dimer (D). The EAT and D-dimer showed a significant difference by ANOVA, and multiple comparisons revealed that the CHA₂DS₂-Vasc score ≥4 group had a significantly shorter EAT and higher D-dimer than the other 2 groups. Neither the PT nor aPTT exhibited any difference among the 3 groups. *, p <0.05.</p

Coagulation parameters in the DBCM and conventional assays in groups with different CHADS₂ scores.

<p>The patients were classified into 3 groups according to their CHADS₂ score (0, 1, and ≥2) for a comparison with the EAT, (A), PT (B), aPTT (C) and D-dimer (D). The EAT and D-dimer showed a statistically significant difference in the 3 groups by ANOVA. Multiple comparisons revealed that the CHADS₂ ≥2 group had a significantly shorter EAT than the CHADS₂ = 0 groups and the CHADS₂ ≥2 group had a significantly higher D-dimer level than the CHADS₂ = 0 or 1 groups. The PT and aPTT exhibited no significant difference. *, p <0.05.</p

Temporal and spectral changes in the dielectric permittivity.

<p>(A) The normalized permittivity after recalcification is plotted from samples with a blood clot formation (left) or without coagulation in patients who underwent an intravenous heparin administration (heparinized samples) (right). The dielectric permittivity gradually increases at a range between 2.5 MHz and 16 MHz in samples with clot formation, whereas heparinized samples show no change in the permittivity in the same frequency range. (B) Representative traces of a normalized permittivity (top) and its derivative (bottom) at 10 MHz plotted against the time from recalcification. The solid line represents the trace from a normal sample with blood clot formation, and the dotted line is that from a heparinized sample without clot formation. The temporal changes in the dielectric permittivity demonstrate a sigmoidal increase, and its derivative shows a single peak in the normal sample. The heparinized sample exhibits no increase in the permittivity, as well as the derivative. The end of acceleration time (EAT) was defined as the time at which the derivative of the permittivity reached the 10% value in the descending phase. (C) The maximum value of the derivative of the dielectric permittivity was plotted in normal samples with a blood clot formation and heparinized sample without clot formation (n = 50 for normals, and 10 for heparinized samples).</p

Characteristics of the patients classified by the CHADS₂ score.

<p>Characteristics of the patients classified by the CHADS₂ score.</p

Reproducibility and interchangeability of the DBCM analysis.

<p>(A) The within-run reproducibility of the EAT is assessed by the correlation between the first measurement of the EAT (EAT1) and second one (EAT2) in healthy subjects (n = 40). The time interval between the first and second measurements was 3 to 5 minutes. The EAT at the time of the two measurements had a high reproducibility (y = 0.58 + 0.96x, r = 0.95, p <0.0001). (B) The interchangeability between the EAT and activated coagulation time (ACT) was evaluated by a simple regression analysis (n = 25). The EAT shows no significant correlation to the ACT.</p