Naruji Sugiyama

Smoking-induced olfactory dysfunction in chronic sinusitis and assessment of brief University of Pennsylvania Smell Identification Test and T&T methods.

Background To evaluate the outcome of smell activity after endoscopic sinus surgery in chronic sinusitis, it is important to verify evidence of smoking-induced olfactory dysfunction. Methods In both the preoperative and the postoperative stages, the 5-odorant T&T olfactometer (T&T5) and the 40-item University of Pennsylvania Smell Identification Test (UPSIT40) were administered to 100 patients (84 men and 16 women; mean age, 49.5 years; range, 21–75 years) who underwent surgery for chronic sinusitis. Additionally, as more simplified measures, we adopted a 3-odorant T&T (T&T3) and a 12-item UPSIT (UPSIT12), and then compared findings with those of each original method. Results (1) Obvious correlations of scores were noticed both between T&T5 and T&T3 and between UPSIT40 and UPSIT12 (r = 0.964 and 0.893, respectively), (2) significant changes in postoperative scores were observed on all four measurements (p < 0.0001), (3) smoking-induced hyposmia was noticeable in older subjects but not in younger subjects, and (4) the correlations of postoperative scores and the age of smoker or the smoking dose was significant (UPSIT40, r = 0.825 or 0.642; UPSIT12, r = 0.666 or 0.428; T&T5, r = 0.447 or 0.476; T&T3, r = 0.457 or 0.500, respectively). Conclusion The abbreviated measures of T&T3 and UPSIT12 could be available enough to assess the effect of surgical intervention on olfaction, while the original UPSIT40 was considered to be the most sensitive among the four methods tested here for examining the impact of smoking on olfaction.

Evaluation of valproate effects on acylcarnitine in epileptic children by LC-MS/MS

BACKGROUNDnValproate (VPA) is a simple fatty acid and a substrate for the fatty acid β-oxidation pathway. Previous data suggested that the toxicity of VPA may be provoked by carnitine deficiency and the inhibition of mitochondrial β-oxidation.nnnOBJECTIVEnThe aim of the present study was to elucidate the effect of VPA treatment on carnitine and isomer-differentiated acylcarnitine disposition, and determined the relationships between acylcarnitines and blood VPA levels in long-term treated patients with VPA and/or other antiepileptic drugs.nnnMETHODSnSerum samples were obtained from children aged 1-15 years old treated for at least 6 months with VPA alone (n=28) or VPA combined with other anticonvulsants (n=23) and untreated controls (n=23). Serum acylcarnitines were separated from their isomers and quantified using high-performance liquid chromatography-tandem mass spectrometry.nnnRESULTSnWe found higher 3-hydroxyisovalerylcarnitine levels and trace amounts of valproylcarnitine in both VPA monotherapy and polytherapy patients. Other acylcarnitines, hexanoylcarnitine, C12, C14:1-carnitines and the ratio of long-chain acylcarnitine to free carnitine were also higher in VPA polytherapy individuals than in controls. VPA monotherapy does not result in decreases in free carnitine or in the accumulation of long-chain acylcarnitines. Blood VPA concentrations correlated positively with hexanoylcarnitine, C12, C14:1, C16:1, C18:1-carnitines in all VPA-treated children (n=51).nnnCONCLUSIONnLong-term VPA treatment in pediatric patients could affect some specific acylcarnitines, which is enhanced by the concomitant use of other anticonvulsants, and the formation of valproylcarnitine alone seems insufficient to develop severe carnitine deficiency at therapeutic doses of VPA.

Kinetic and molecular orbital analyses of dicarboxylic acylcarnitine methylesterification show that derivatization may affect the screening of newborns by tandem mass spectrometry

Newborns are routinely screened for organic acidemias by acylcarnitine analysis. We previously reported the partial catalytic methylesterification of dicarboxylic acylcarnitines by benzenesulfonic acid moiety in the solid extraction cartridge during extraction from serum. Since the diagnosis of organic acidemias by tandem mass spectrometry is affected by the higher molecular weight of these derivatized acylcarnitines, we investigated the methylesterification conditions. The kinetic constants for the methylesterification of carboxyl groups on the acyl and carnitine sides of carnitine were 2.5 and 0.24h(-1), respectively. The physical basis underlying this difference in methylesterification rates was clarified theoretically, illustrating that methylesterification during extraction proceeds easily and must be prevented.

A case of holocarboxylase synthetase deficiency with insufficient response to prenatal biotin therapy

Holocarboxylase synthetase (HCS) deficiency is an inborn error of biotin metabolism, leading to a multiple carboxylases deficiency. As the affected fetus sometimes presents with enlargement of the cerebral ventricles and intrauterine growth retardation (IUGR), prenatal administration of biotin has been attempted in some pregnancies. We present herein the case of a Japanese neonate with HCS deficiency who received maternal administration of biotin (10mg/day) from 33 weeks gestation. After biotin administration, the fetal body weight increased and gestation was continued to full term. However, lactic acidemia and metabolic acidosis were observed after birth. To evaluate the effects of prenatal therapy, we collected serum samples and measured the acylcarnitine profiles using high-performance liquid chromatography electrospray ionization tandem mass spectrometry. At birth, levels of propionylcarnitine and 3-hydroxyisovalerylcarnitine had already increased. At 2h after birth, these levels of acylcarnitines were further increased. At 3.5h after the start of biotin, these chemical findings were slightly improved. In conclusion, we considered that prenatal biotin therapy at 10mg/day may have been inadequate to avoid neonatal acidotic crisis in this case.

Assay for methylmalonyl coenzyme A mutase activity based on determination of succinyl coenzyme A by ultrahigh-performance liquid chromatography tandem mass spectrometry

Methylmalonic acidemia (MMA) is an inherited metabolic disease. In this condition, metabolism from methylmalonyl coenzyme A (CoA) to succinyl-CoA is inhibited because of either low methylmalonyl-CoA mutase (MCM) activity or adenosylcobalamin deficiency owing to altered vitamin B12 metabolism. A high-precision assay for detecting MCM activity would facilitate not only MMA diagnosis but also the ability to determine the severity of MMA. We developed an MCM assay method based on ultrahigh-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) that involves the determination of succinyl-CoA, which is formed in an enzyme reaction, using peripheral lymphocytes. Using 0.05, 0.5, and 5xa0μmol/L succinyl-CoA, the intra-assay coefficient of variation (CV) was less than 5.2xa0% and the inter-assay CV was less than 8.7xa0%. The MCM activities of five healthy individuals and four patients were investigated with this assay. The MCM activities of the patients were very low in relation to those of healthy individuals. Together, these results show that the UPLC–MS/MS method is useful for a detailed MCM activity assay.

Fatal lipid storage disorder

A 12-week-old female infant died from acute encephalopathy mimicking Reye syndrome. Because of positive serum hepatitis B surface antigen (HBs Ag) and perivascular inflammatory cell infiltration in the liver, she was diagnosed as having acute hepatitis. The most striking finding in the present case was extremely excessive lipid accumulation in the striated muscles including biceps brachii, tongue and cardiac muscles. The levels of serum, liver and muscle carnitine were within normal limits, though liver carnitine palmityl transferase (CPT) was markedly decreased in activity. Although the primary metabolic defect has yet to be elucidated, it is assumed that the fulminant hepatic failure induced lipid accumulation in the skeletal muscle by a certain abnormal lipid metabolism.

Determination of methylmalonyl coenzyme A by ultra high-performance liquid chromatography tandem mass spectrometry for measuring propionyl coenzyme A carboxylase activity in patients with propionic acidemia

Propionic acidemia (PA) is an inherited metabolic disease caused by low activity of propionyl coenzyme A (CoA) carboxylase (PCC), which metabolizes propionyl-CoA into methylmalonyl-CoA. Although many patients with PA have been identified by tandem mass spectrometry since the test was first included in neonatal mass screening in the 1990s, the disease severity varies. Thus, determining the specific level of PCC activity is considered to be helpful to grasp the severity of PA. We developed a new PCC assay method by the determination of methylmalonyl-CoA, which is formed by an enzyme reaction using peripheral lymphocytes, based on ultra high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). With methylmalonyl-CoA concentrations of 0.05, 0.5, and 5μmol/L, the intra-assay coefficients of variation (CVs) were 8.2%, 8.7%, and 5.1%, respectively, and the inter-assay CVs were 13.6%, 10.5%, and 5.9%, respectively. The PCC activities of 20 healthy individuals and 6 PA patients were investigated with this assay. Methylmalonyl-CoA was not detected in one PA patient with a severe form of the disease, but the remaining PA patients with mild disease showed residual activities (3.3-7.8%). These results demonstrate that determination of PCC activity with this assay would be useful to distinguish between mild and severe cases of PA to help choose an appropriate treatment plan.

Neonatal alkalemia associated with potential hypovolemia in an infant born to a severely dehydrated mother

A newborn male infant with a birthweight of 2626 g was delivered at 38 weeks gestation by cesarean section; the indication was prolonged bradycardia (80 beats/min). At birth, the infant exhibited respiratory distress due to meconium aspiration (Apgar score, 1/6 at 1/5 min); therefore, he was immediately intubated. His 32-year-old mother experienced severe hyperemesis at 36 weeks’ gestation secondary to cancer-induced pyloric stenosis; her oral intake was markedly limited. The physician in charge of her thought that her symptoms were transient and left her untreated. Over the next 2 weeks, because of inadequate fl uid intake, she lost 5 kg of bodyweight and developed severe metabolic alkalosis with hypercapnia, hypokalemia, hypochloremia, and acute renal failure ( Table 1). On admission to the neonatal intensive care unit, the following were noted: body temperature 36.2°C; pulse 126 beats/ min; respiration 42/min; and blood pressure 35/14 mmHg. Physical examination revealed: bilateral clear breath sounds, normal heart impulse and rhythm, and no murmur. The abdominal exam was normal, and the liver was not palpable. The infant also had marked metabolic alkalosis with hypercapnia, hypokalemia, and hypochloremia ( Table 1 ). Chest radiography showed bilateral granular opacity of the lungs without cardiomegaly, compatible with the meconium aspiration syndrome. Echocardiography revealed unimpaired cardiac function (ejection fraction = 70.9%) and a patent ductus arteriosus with left-to-right shunt. Renal, bladder and cranial ultrasound examinations were normal. The systolic blood pressure was maintained at approximately 30 mmHg during the initial therapy, which included intermittent mandatory ventilation (IMV), and 7% dextrose-in-water infusion at 60 mL/kg per day. During the initial therapy, we carefully monitored the infusion to avoid increasing pulmonary blood fl ow through the ductus arterisosus, which could result in pulmonary hypertension. Because no improvement was noted, the infusion of dopamine (5 x01 g/kg per min) and dobutamine (5 x01 g/kg per min) were begun at 2 h of age; in addition, a milrinone infusion (0.5 x01 g/kg per min) was begun at 4 h of age. Although adequate ventilation and oxygenation had been established, neither an increase in blood pressure nor diuresis was observed. At 4 h of age, the infant suffered a grand mal seizure, which was controlled with a Phenobarbital suppository (10 mL/kg). At 8 h of age, a fl uid challenge of 20 mL/kg containing albumin was given for a duration of 2 h. Subsequently, the blood pressure rapidly increased and urinary output rose to an acceptable level ( Fig. 1). After diuresis, supplementation with chloride (1.9 mmol/kg per day) and potassium (1.0 mmol/kg per day) was begun. At 10 h of age, the fractional excretion of sodium (FENa) was 1.7%, and the renal failure index (RFI) was 2.45, indicating probable prerenal failure. 4,5 At 24 h of age, metabolic alkalosis and hypokalemia transiently exacerbated ( Table 1 ; Fig. 2). We continued hydration and Cl − /K + supplementation, which resulted in a weaning from IMV and extubation at 4 days of age; correction of all abnormal fi ndings, including alkalemia, occurred at 5 days of age ( Table 1 ; Fig. 2 ). Subsequently, the neonate did well. Patient Report