Outcomes of Joint-Preserving Surgery for Forefoot Deformities in Patients With Rheumatoid Arthritis.

BACKGROUND: With the advent of effective disease-modifying medications, the surgical treatment of forefoot deformities in patients with rheumatoid arthritis (RA) has evolved from joint-sacrificing to joint-preserving surgery. However, it is unclear whether joint-preserving surgery is effective for the full range of metatarsophalangeal joint involvement. Hence, this study investigated the postoperative outcomes of joint-preserving surgery for rheumatoid forefoot deformities with a wide range of joint destruction. METHODS: This retrospective observational study included 68 feet from 68 patients with RA who underwent joint-preserving surgery for forefoot deformities between 2014 and 2020. The Larsen grade classification was used to assess the first metatarsophalangeal joint destruction and classify patients into 4 groups as follows: 0 and 1 (n = 14), 2 (n = 21), 3 (n = 19), and 4 and 5 (n = 14). The Self-Administered Foot and Ankle Evaluation Questionnaire (SAFE-Q) score, hallux valgus angle (HVA), and intermetatarsal angle (IMA) were determined before surgery and at the last follow-up visit. RESULTS: The median observation duration was 40 (range, 24-78) months. SAFE-Q scores of all groups significantly improved in all subscales at the last observation, with no significant differences among the study groups. Radiographic evaluations of all groups revealed significant improvements in HVA and IMA after surgery, with no significant differences among the groups. CONCLUSION: In patients using the surgical approaches described in this study, joint-preserving surgery for rheumatoid forefoot deformities led to satisfactory clinical and radiographic improvements, regardless of the severity of joint destruction. LEVEL OF EVIDENCE: Level III, case-control study.

Mechanism for inhibition of cytotoxicity of Shiga toxin by luteolin.

Enterohemorrhagic or Shiga toxin-producing Escherichia coli is a food-poisoning bacterium that grows in the intestine to produce Shiga toxin (Stx). In this study, the effects of 20 polyphenols on the cytotoxicity of Stx1 and Stx2 in Vero cells were investigated. Among these, epigallocatechin gallate, butein, isorhapontigenin, hesperetin, morin, luteolin, resveratrol, and rhapontigenin showed inhibitory effects on the cytotoxicity of Stxs at 0.4 mmol/L. Furthermore, Vero cells pre-treated with these polyphenols were resistant to Stx at 0.4 mmol/L. However, luteolin showed the most potent inhibitory and cytoprotective effect against Stxs at 0.08 mmol/L or more. This inhibitory mechanism of luteolin was determined using a cell-free protein synthesis system and quantitative reverse transcription PCR assay to detect depurination of 28S rRNA in Vero cells. Luteolin did not inhibit the cell-free protein synthesis by Stxs, suggesting that the enzymatic activity of the Stx A subunit was not inhibited by luteolin. The depurination of 28S rRNA by Stxs was also investigated in Vero cells. The 28S rRNA depurination by Stxs was suppressed in Vero cells treated with Stxs which had been pretreated with luteolin. These results suggest that luteolin inhibits the incorporation of Stxs into Vero cells. This is the first report to show that luteolin inhibits the cytotoxicity of both Stx1 and Stx2 by inhibiting the incorporation of Stxs into Vero cells.

Crystal Structures of Lacticaseibacillus 4-Deoxy-L-threo-5-hexosulose-uronate Ketol-isomerase KduI in Complex with Substrate Analogs.

Some probiotics including lactobacilli, colonize host animal cells by targeting glycosaminoglycans (GAGs), such as heparin, located in the extracellular matrix. Recent studies have shown that several lactic acid bacteria degrade GAGs. Here we show the structure/function relationship of Lacticaseibacillus rhamnosus 4-deoxy-L-threo-5-hexosulose-uronate ketol-isomerase (KduI) crucial for metabolism of unsaturated glucuronic acid produced through degradation of GAGs. Crystal structures of ligand-free and bound KduIs were determined by X-ray crystallography and the enzyme was found to consist of six identical subunits and adopt a ß-helix as a basic scaffold. Ligands structurally similar to the substrate were bound to the cleft of each enzyme subunit. Several residues located in the cleft interacted with ligands through hydrogen bonds and/or C-C contacts. In addition to substrate analogs, a metal ion coordinated to four residues, His198, His200, Glu205, and His248, in the cleft, and the enzyme activity was significantly inhibited by a chelator, ethylenediaminetetraacetic acid. Site-directed mutants in Arg163, Ile165, Thr184, Thr194, His200, Arg203, Tyr207, Met262, and Tyr269 in the cleft exhibited little enzyme activity, indicating that these residues and the metal ion constituted an active site in the cleft. This is the first report on the active site structure of KduI based on the ligand-bound complex.

Baicalein Inhibits Stx1 and 2 of EHE: Effects of Baicalein on the Cytotoxicity, Production, and Secretion of Shiga Toxins of Enterohaemorrhagic Escherichia coli.

Shiga toxin-producing enterohaemorrhagic Escherichia coli (EHEC) O157:H7 is an important foodborne pathogen. Baicalein (5,6,7-trihydroxylflavone), a flavone isolated from the roots of Scutellaria baicalensis, is considered as a potential antibacterial agent to control foodborne pathogens. Among seven compounds selected by in silico screening of the natural compound database, baicalein inhibited the cytotoxicity of both Shiga toxins 1 and 2 (Stx1 and Stx2) against Vero cells after pretreatment at 0.13 mmol/L. In addition, baicalein reduced the susceptibility of Vero cells to both Stx1 and Stx2. Real-time qPCR showed that baicalein increased transcription of stx1 but not of stx2. However, baicalein had no effects on production or secretion of Stx1 or Stx2. Docking models suggested that baicalein formed a stable structure with StxB pentamer with low intramolecular energy. The results demonstrate that inhibitory activity of baicalein against the cytotoxicity of both Stx1 and Stx2 might be due to of the formation of a binding structure inside the pocket of the Stx1B and Stx2B pentamers.

Glyceraldehyde-3-phosphate dehydrogenase of Mycoplasma pneumoniae induces infection-related glomerulonephritis
.

Infection-related glomerulonephritis (IRGN) was previously thought to be due mostly to Streptococcus species, but is now known to be caused by a variety of other pathogens. Nephritis-associated plasmin receptor (NAPlr) was originally isolated from group A streptococci as the protein responsible for acute poststreptococcal glomerulonephritis, and was shown to be identical to streptococcal glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Here, we describe a 7-year-old boy diagnosed with Mycoplasma pneumoniae IRGN presenting with acute nephritic syndrome. Laboratory data revealed a significant increase in serum anti-M. pneumoniae antibody titer. Renal biopsy revealed diffuse global endocapillary proliferation and cellular crescents in 5/43 glomeruli examined. Although antistreptolysin O antibody titer and serum complement C3 level were within the respective normal ranges, glomeruli showed positive staining for NAPlr and upregulation of plasmin activity. In addition, positive staining for NAPlr in the glomeruli was abolished by preabsorption of anti-NAPlr antibody with recombinant M. pneumoniae GAPDH. Western blotting analysis revealed anti-NAPlr antibody reactivity with a band at around the predicted size of GAPDH in the protein isolate of M. pneumoniae (37 kDa). Furthermore, immobilized M. pneumoniae GAPDH bound to anti-NAPlr antibody as well as plasmin in vitro. These data suggest that M. pneumoniae GAPDH has a function similar to streptococcal GAPDH (NAPlr) and may induce plasmin-related glomerular damage in M. pneumoniae IRGN. NAPlr could be a marker of glomerulonephritis related to infection not only by streptococci but also by &M. pneumoniae.

Compensatory Function of the Subtalar Joint for Lower Extremity Malalignment.

It is important to evaluate the subtalar joint and hip-knee-ankle alignment to understand lower extremity alignment. In this review, we focused on the compensatory changes in the subtalar joint alignment for the deformity of the knee and ankle joint, reviewing previous research. The subtalar joint alignment was compensatory valgus in patients with varus knee and ankle deformity, whereas it was uncertain whether the subtalar joint alignment was compensatory varus in patients with valgus knee and ankle deformity. The subtalar joint valgus alignment improved after total knee arthroplasty or high tibial osteotomy for varus knee deformity, even if the deformity was severe. In contrast, whether the subtalar joint alignment changed after the surgery for ankle or valgus knee deformity has not been considered. Further research on the compensatory function of the subtalar joint is needed.

Initial treatment with pulse methylprednisolone followed by short-term prednisolone and tonsillectomy for childhood IgA nephropathy.

BACKGROUND: Although a combination therapy, comprising 2-year high-dose oral prednisolone (PSL), is effective for treating childhood immunoglobulin A nephropathy (IgAN), severe adverse effects and residual proteinuria occur in some patients after the therapy. METHODS: To clarify the efficacy of intravenous pulse methylprednisolone (IVMP; 15-20 mg/kg; maximum 600 mg/day; for 3 consecutive days/week for 3 weeks) followed by short-term reduced-dose PSL (initially 1 mg/kg; maximum 30 mg on alternate days; tapered off within approximately 12 months) and tonsillectomy as an initial treatment, we retrospectively reviewed the clinical courses of 54 consecutive children with IgAN (32 boys; mean age at onset, 12.2 years; follow-up period of > 2 years) after initiating the treatment. According to the Japanese pediatric IgAN guidelines, we divided the 54 patients into the following two groups: group 1, comprising 24 patients with severe IgAN, and group 2, comprising 30 patients with mild IgAN. RESULTS: After the treatment, proteinuria disappeared in all patients at a median of 1.6 months (group 1, 2.8 months; group 2, 0.4 months) and hematuria disappeared in 47 patients (87%) at a median of 13.2 months (group 1, 15.9 months; group 2, 13.2 months). During the follow-up period (median 5 years), no severe adverse effects were observed in any patient. At the last visit, although two patients (4%) had mild proteinuria, none developed hypertension or renal insufficiency. CONCLUSIONS: As an initial treatment, IVMP followed by short-term PSL and tonsillectomy appears to be effective for treating childhood IgAN.

Effect of adipose-derived mesenchymal stem cell transplantation on vascular calcification in rats with adenine-induced kidney disease.

Previous studies have investigated the use of mesenchymal stem cells (MSCs) to treat damaged kidneys. However, the effect of adipose-derived MSCs (ASCs) on vascular calcification in chronic kidney disease (CKD) is still poorly understood. In the present study, we explored the potential of ASCs for the treatment of CKD and vascular calcification. CKD was induced in male Sprague-Dawley rats by feeding them a diet containing 0.75% adenine for 4 weeks. ASCs transplantation significantly reduced serum inorganic phosphorus (Pi) as compared to that in the control. The histopathology of the kidneys showed a greater dilation of tubular lumens and interstitial fibrosis in the control group. Calcium and Pi contents of the aorta in the ASCs transplantation group were lower than those in the control group. Von Kossa staining of the thoracic aorta media revealed that ASCs transplantation suppressed vascular calcification. Thus, this study revealed that autogenic ASCs transplantation inhibits kidney damage and suppresses the progression of vascular calcification in the CKD rat model, suggesting that autogenic ASCs transplantation is a novel approach for preventing the progression of CKD and vascular calcification.

Independent Risk Factors and 2-Year Outcomes of Acute Kidney Injury after Surgery for Congenital Heart Disease.

BACKGROUND: Data are limited regarding risk factors for acute kidney injury (AKI) following cardiac surgery in children with congenital heart disease (CHD). This observational study was performed to examine temporal trends in AKI incidence according to the Pediatric Risk, Injury, Failure, Loss, End-Stage Renal Disease (pRIFLE) criteria, identify independent risk factors for AKI after cardiac surgery, and examine associations between AKI and long-term mortality. METHODS: We retrospectively evaluated 418 patients (259 males, 159 females; median age, 5 months) who underwent cardiac surgery for CHD between April 2007 and August 2013. Patients were followed up for 2 years. AKI was defined according to the pRIFLE criteria as ≥25% decrease in estimated creatinine clearance. RESULTS: AKI developed postoperatively in 104 cases (24.9%). Approximately 80% belonged to the "Risk" category according to the pRIFLE criteria, and only 21 cases (5%) required renal replacement therapy (peritoneal dialysis in all cases). Multivariate analysis revealed 3 independent risk factors for onset of AKI: young age (<1 year), surgery in Risk Adjustment in Congenital Heart Surgery (RACHS-1) category ≥4, and long cardiopulmonary bypass (CPB) time (≥90 min). Twenty-three patients (22%) with AKI died during the 2-year follow-up. In multivariate cox hazard regression analysis, the most significant contributor to risk of mortality was AKI. CONCLUSIONS: Postoperative AKI was strongly associated with young age, high RACHS-1 category, and prolonged CPB time. In addition, mortality rate was higher in patients who survived after recovery from AKI than in those without AKI, even among the lower pRIFLE categories.

Predictors of relapse and long-term outcome in children with steroid-dependent nephrotic syndrome after rituximab treatment.

BACKGROUND: In patients with complicated steroid-dependent nephrotic syndrome (SDNS), rituximab (RTX) followed by immunosuppressive agent (IS) can maintain remission without the use of prednisolone (PSL). However, available data on the predictive factors for relapse and the long-term outcome after this protocol are few. METHODS: We retrospectively analyzed 43 SDNS patients who were followed-up for a long time (>2 years, mean 5.4 years) after a single dose of RTX (375 mg/m2) from September 2007. After RTX, PSL was tapered off within 6 months; monotherapy with IS, such as cyclosporine or mycophenolate mofetil, was continued to prevent post-RTX relapse. For patients who achieved >12 months of PSL-free remission, IS was also tapered off. RESULTS: Thirty-nine patients (91 %) could discontinue PSL without relapses at a median of 154 days after the initial RTX. The first relapse of NS occurred in 39 patients (91 %) at a median of 586 days; additional RTX doses were administered in 28 patients (65 %). Kaplan-Meier analysis showed that shorter CD19 cell depletion (<150 days) and younger age at RTX initiation (<12.5 years) were significantly associated with high risk for first relapse after RTX (log rank p < 0.05). In multivariate analysis, mycophenolate mofetil therapy as maintenance IS after RTX was the only predicted risk factor for first relapse (hazard ratio 2.75; p = 0.027). At the last follow-up, IS was still used in 33 patients (77 %); treatment-free remission (>12 months) was achieved in only five patients (12 %). CONCLUSIONS: The introduction of RTX may not be necessarily associated with improved long-term outcome.

Correction: Adipose Tissue-Derived Mesenchymal Stem Cells in Long-Term Dialysis Patients Display Downregulation of PCAF Expression and Poor Angiogenesis Activation.

[This corrects the article DOI: 10.1371/journal.pone.0102311.].

Late-onset adverse events after a single dose of rituximab in children with complicated steroid-dependent nephrotic syndrome.

BACKGROUND: Rituximab (RTX) is regarded as a relatively safe and effective treatment for children with steroid-dependent nephrotic syndrome (SDNS). However, late-onset adverse events after RTX, including neutropenia, hypogammaglobulinemia, and increased risk of infections, have been rarely reported in this cohort. MATERIALS AND METHODS: This was a single-center retrospective analysis of adverse events during B-cell depletion periods after a single dose of RTX (375 mg/m2) in 60 patients with complicated SDNS (total 126 doses). After RTX, maintenance therapy with cyclosporine (CsA) or mycophenolate mofetil (MMF) was continued, and prednisolone was discontinued within 6 months. To detect potential drug toxicity, clinical and laboratory parameters were measured before and 1 week after RTX infusion and every month thereafter during B-cell depletion periods (at least 6 months). A single dose of RTX was added if NS relapsed despite maintenance therapy with MMF or CsA after the re-emergence of CD19+ B cells (> 1% of total lymphocytes) in the peripheral blood. RESULTS: Severe neutropenia (neutrophil count < 500/mm3) was identified in 3 patients and hypogammaglobulinemia (IgG levels < 500 mg/dL) in 9 patients. During B-cell depletion periods (median 5 months; range 1 - 20 months), 2 patients required hospitalization because of bacterial infections. However, no lifethreatening infections were identified in our cohort. CONCLUSION: Although neutropenia and hypogammaglobulinemia should be kept in mind as late-onset adverse events of RTX therapy in patients with complicated SDNS, severe infections during B-cell depletion periods are infrequent when our treatment strategies are implemented.

Tolvaptan in a pediatric patient with diuretic-resistant heart and kidney failure.

Despite conventional diuretic therapy, volume overload persists in many patients with decompensated heart failure. Adverse effects of diuretics are common, including worsening kidney function and electrolyte disturbance. Furthermore, decreased kidney function also affects the response to diuretics and is associated with an increased risk of mortality. A 10-year-old boy with congestive heart failure (CHF) complicated by advanced chronic kidney disease (CKD) presented with oliguria and generalized edema. He was being treated with furosemide and spironolactone, and these doses were increased to 3 mg/kg/day after admission. Although edema decreased temporarily, the symptoms worsened and furosemide resistance developed 2 months later. Tolvaptan (0.1 mg/kg/day) was started, resulting in a gradual increase in the plasma sodium level and adequate decongestion of the volume overload state. Cardiac function also improved. The use of tolvaptan should be considered in pediatric cases of conventional diuretic-resistant CHF, even when complicated by advanced CKD.

Positive role of rituximab in switching from cyclosporine to mycophenolate mofetil for children with high-dose steroid-dependent nephrotic syndrome.

BACKGROUND: Recent randomized studies indicate that mycophenolate mofetil (MMF) is inferior to cyclosporine (CsA) in preventing relapses of nephrotic syndrome (NS). During the last decade, rituximab (RTX) has emerged as a rescue therapy in patients with complicated, frequently relapsing, or steroid-dependent NS. CASE-DIAGNOSIS/TREATMENT: After introducing RTX in our single center, we analyzed 26 patients with steroid-dependent NS who had relapses while receiving long-term CsA and who were subsequently switched to MMF. MMF was adjusted to maintain a targeted predose mycophenolic acid (MPA) level of 2-5 µg/ml. Moreover, for patients who required MMF and high-dose prednisolone (PSL) to maintain remission, a single infusion of RTX (375 mg/m(2)) was added. The primary endpoint was the probability of achieving PSL-free remission for >1 year. At a mean follow-up of 28.8 ± 9.9 months, 11 of 26 patients (42 %) required RTX treatment, and 22 of those patients (85 %) achieved PSL-free sustained remission. The mean predose MPA levels for patients who achieved PSL-free sustained remission were significantly higher compared with those for patients who did not (3.1 µg/ml vs. 1.7 µg/ml, p < 0.05). CONCLUSIONS: After RTX introduction, most patients were able to switch from CsA to MMF and achieve sustained PSL-free remission.

Adipose tissue-derived mesenchymal stem cells in long-term dialysis patients display downregulation of PCAF expression and poor angiogenesis activation.

We previously demonstrated that mesenchymal stem cells (MSCs) differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD) patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months) and from healthy controls (HC-MSCs) to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF) was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α), we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF) expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients.

Comparison of multipotency and molecular profile of MSCs between CKD and healthy rats.

We previously showed that mesenchymal stem cells (MSCs) can differentiate into a functional miniature kidney, suggesting that MSCs may be a cell source for kidney regeneration. However, MSCs from long-term dialysis patients, which have been exposed to uremic toxin, can exhibit reduced viability. Therefore, the aim of this study was to examine the gene expression profiles and differentiation capabilities of bone marrow- and adipose-derived MSCs from chronic kidney disease (CKD) model rats. CKD was induced in rats by adenine feeding, and then MSCs were isolated from bone marrow (BMSCs) and adipose tissue (ASCs). After confirming MSC surface marker expression, comprehensive gene expression profiles were obtained by RT-PCR array. MSCs were differentiated into adipocytes, osteoblasts, and chondrocytes, and histological and/or functional assays were performed. Tgfb3 expression was up-regulated, while Bmp6, Gdf15, Mmp2, and Vegfa were down-regulated in CKD-ASCs compared with Control-ASCs. There were no significant differences in the gene expression of stemness markers, and the morphology of cells that underwent adipogenesis, osteogenesis, and chondrogenesis, or GPDH activity between CKD and control groups. Comparing BMSCs with ASCs, gene expression of Bglap, Bmp4, Igf1, Itgax, Pparg, Ptprc, and Tnf were up-regulated, while Col1a1, Mmp2, Sox9, and Vegfa were down-regulated in both CKD and control groups. Uremic toxin in CKD rats had a small effect on the gene expression and differentiation of MSCs. However, long-term exposure to uremic toxin and the differences in gene expression of MSCs derived from bone marrow or adipose tissue may affect renal regeneration.

Glucose utilization in the brain during acute seizure is a useful biomarker for the evaluation of anticonvulsants: effect of methyl ethyl ketone in lithium-pilocarpine status epilepticus rats.

Enhancement of glucose utilization in the brain has been well known during acute seizure in various kinds of animal model of epilepsy. This enhancement of glucose utilization might be related to neural damage in these animal models. Recently, we found that methyl ethyl ketone (MEK) had both anticonvulsive and neuroprotective effects in lithium-pilocapine (Li-pilo) status epilepticus (SE) rat. In this article, we measured the uptake of [(14)C]2-deoxyglucose ([(14)C]DG) in the Li-pilo SE and Li-pilo SE with MEK rat brain in order to assess whether the glucose utilization was a useful biomarker for the detection of efficacy of anticonvulsive compounds. Significant increase of [(14)C]DG uptake (45 min after the injection) in the cerebral cortex, hippocampus, amygdala and thalamus during acute seizure induced by Li-pilo were observed. On the other hand, the initial uptake of [(14)C]DG (1 min after the injection) in the Li-pilo SE rats was not different from the control rats. Therefore, the enhancement of glucose metabolism during acute seizure was due to the facilitation of the rate of phosphorylation process of [(14)C]DG in the brain. Pretreatment with MEK (8 mmol/kg) completely abolished the enhancement of glucose utilization in the Li-pilo SE rats. The present results indicated that glucose utilization in the brain during acute seizure might be a useful biomarker for the evaluation of efficacy of anticonvulsive compounds.

Lactate is an alternative energy fuel to glucose in neurons under anesthesia.

The uptake of [14C]lactate was measured in the brains of mice anesthetized with pentobarbital or chloral hydrate. The results showed significant increase of the [14C]lactate uptake in the brain under both anesthesia. Despite energy metabolism in the brain being suppressed by both pentobarbital and chloral hydrate, the [14C]lactate uptake was unexpectedly increased under anesthesia. [14C]Lactate uptake in rat brain injured by infusion of quinolic acid was significantly decreased, and the reduction of [14C]lactate uptake was parallel to neural cell death, suggesting that exogenous lactate might be selectively taken up by neuron. These results indicated that lactate rather than glucose might serve as an energy substrate for neuron in intact brain under anesthesia.

Liquid chromatographic determination of acetylcholine based on pre-column alkaline cleavage reaction and post-column tris(2,2'-bipyridyl)ruthenium(III) chemiluminescence detection.

A method for the determination of acetylcholine (ACh) has been developed using liquid chromatography with chemiluminescence detection. This method is based on the pre-column alkaline cleavage of ACh to form trimethylamine (TMA) and the post-column tris(2,2'-bipyridyl)ruthenium(III) chemiluminescence detection of TMA. ACh was converted to TMA with high yield at 180 degrees C in the presence of lithium hydroxide, and the produced TMA was separated on a cation-exchange/reversed-phase dual-functional column using a mixture of 0.2 M potassium phosphate buffer (pH 5.9) and acetonitrile (20:1, v/v) as the mobile phase. The eluate was online mixed with acidic tris(2,2'-bipyridyl)ruthenium(III) solution, and the generated chemiluminescence was detected. The detection limit (signal-to-noise ratio = 3) for ACh was 0.80 nmol/mL, which corresponded to 1.1 pmol TMA per injection volume of 5 microL. This is simple and robust method that does not need an expensive device and unstable enzymes, and was applied to the determination of ACh in pharmaceutical formulations.

Electrooxidation of vanillyl alcohol in acidic aqueous solution using rotating ring-disk electrode voltammetry.

The electrooxidation of benzylic alcohol derivative in acidic aqueous solution shows an oxidation pre-peak in the cyclic voltammogram, which means that the reaction is proceeding via an ECE mechanism where the second electron transfer occurs at a less positive potential. From the result of the rotating ring-disk electrode voltammetry, the initial oxidation response of the electrode reaction can be extracted.