Effect of acute acidosis and alkalosis on leucine kinetics in man

Summary. The effects of acute pH changes on whole body leucine kinetics (1-¹³C-leucine infusion technique) were determined in normal subjects. Plasma insulin, glucagon, and growth hormone concentrations were kept constant by somatostatin and replacement infusions of the three hormones. When acidosis was produced by ingestion of NH4CI (4 mmol kg^-1 p. os; n = 8) arterialized pH decreased within 3 h from 7.39±0.01 to 7.31 ±0.01 (P<0.001) and leucine plasma appearance increased by 0.13 ±0.04 μmol kg^-1 min^-1 (P<0.02); in contrast, when alkalosis was produced by intravenous infusion of 4 mmol kg^-1 NaHCO₃ (n= 1, pH 7.47 ±0.01), leucine plasma appearance decreased by -0.09 ± 0.04 (xmol kg^-1 min^-1 (P<0.01 vs. acidosis). Whole body leucine flux also increased during acidosis compared to alkalosis (P<0.05), suggesting an increase in whole body protein breakdown during acidosis. Apparent leucine oxidation increased during acidosis compared to alkalosis (P=0.05). Net forearm leucine exchange remained unaffected by acute pH changes. Plasma FFA concentrations decreased during acidosis by -107 ±67 μmol l^-1 (P<0.05) and plasma glucose increased by 1.90±0.25 mmol l^-1 (P<0.02); in contrast, alkalosis resulted in an increase in plasma FFA by 83 ± 40 (μmol 1^-1 (P<0.02; P<0.01 vs. acidosis), suggesting an increase in lipolysis; plasma glucose decreased compared to acidosis (P<0.01). The data demonstrate that acute metabolic acidosis and alkalosis, as they occur in clinical conditions, influence protein breakdown, and in the opposite direction, lipolysis.     

Protein sparing and protein replacement in acutely injured patients during TPN with and without amino acid supply

The metabolic effects of TPN were studied in a selected group of trauma patients. Nineteen patients were randomly divided into two groups: the first was treated with glucose and insulin, the second with glucose, insulin and amino acids. Each patient in both groups received TPN isocaloric with respect to daily energy output and the treatment lasted five days. Each group was further divided into two subsets (severe or moderate catabolism) according to fasting energy output with respect to the expected energy expenditure. During the acute flow phase, both in moderate as well as in severe catabolism, glucose and insulin were effective for protein sparing; the maximum protein sparing effect was reached when giving a caloric intake equal to 130% of daily energy output. Glucose, insulin and amino acids were effective in replacement of nitrogen losses. In moderately catabolic patients nitrogen balance was significantly better than in severely catabolic patients. This study shows that early and short-term TPN is effective in controlling the flow phase of trauma. Glucose and insulin appear to be the determinants of the protein sparing effect when given in amounts equal to those needed; amino acids provided protein replacement when given in amounts equal to about 20% of energy output. Energy supply higher than 120–130% of daily energy output does not increase protein sparing and protein replacement, the only effect being a further increase in metabolism, which is possibly dangerous in critically ill patients.     

核酸提取方法在聚合酶链反应测定乙肝病毒核酸中的评价

目的 对不同核酸提取方法在荧光定量聚合酶链反应（FQ－PCR）检测血清乙型肝炎病毒核酸（HBV DNA）含量中应用进行评价，为临床方法学选择和实验结果的评价提供理论依据。方法 选用直接煮沸裂解法、NP－40煮沸裂解法、沉淀煮沸裂解法、磁珠核酸提取法等4种核酸提取方法提取乙型肝炎血清标本和含不同含量肝素的血清标本中HBV DNA模板，用荧光实时定量PCR法对其进行准确定量，从提取效率、抗干扰能力方面进行评价。结果 4种核酸提取方法中磁珠核酸提取法提取效率为最高，对数换算后，以此相对提取效率为100％，则直接煮沸裂解法为最低81％（P＜0．05）、NP－40煮沸裂解法为95％（P＞0．05）、沉淀煮沸裂解法为88．7％（P＜0．05）；从抗干扰能力方面，当肝素含量为500U／ml时，磁珠核酸提取法抗干扰能力最强，沉淀煮沸裂解法次之、直接煮沸裂解法和NP－40煮沸裂解法为最差，其HBV DNA含量抑制率分别为9．4％、13．5％、35．4％、52．7％。结论 本实验结果认为磁珠核酸提取法为实验室首选的核酸提取方法。     

Characterization of prolidase activity in erythrocytes from a patient with prolidase deficiency: comparison with prolidase I and II purified from normal human erythrocytes

OBJECTIVES: The purpose of this study was to investigate the effect of various amino acids and their metabolites on the activities of prolidase I and II from human erythrocytes compared to those in a patient with prolidase deficiency. DESIGN AND METHODS: Prolidase I and II from human erythrocytes were purified by using column chromatography. Prolidase activity against various iminodipeptides was determined by spectrophotometry using Chinard's method. RESULTS: The activities of prolidase I and II against glycylproline and methionylproline were enhanced by glycine, L- and D-isoforms of alanine and serine and D-isoforms of valine, leucine and isoleucine. L-isoforms of branched amino acids inhibited the activity of prolidase I. On the other hand, the activity of prolidase II was enhanced by all of these L-branched amino acids. The patient's prolidase activity was also enhanced by all the L- and D-branched amino acids. CONCLUSION: The activities of prolidase I and II against various iminodipeptides were prominently enhanced by glycine, but the effect of L-valine differed between the two enzymes. Enzymatic properties of the patient's prolidase were essentially the same as those of prolidase II.     

The effects of albumin bound fatty acids on the platelet inhibitory function of human endothelial cells

Abstract. This study was carried out to evaluate the effects of albumin-bound fatty acids on the anti-platelet effects of endothelial cells. Primary cultures of human endothelial cells (ECM), grown in confluent monolayers, were incubated with plasma or growth medium enriched with albumin-bound fatty acids (FA) for 2–20 h. The effects of ECM on ADP-induced platelet aggregation (PA) and collagen-induced PA and prostaglandin synthesis in platelet-rich plasma were tested.
ECM released an inhibitor of platelet reactions which resembled the activity of PGI₂ (prostacyclin). The inhibitory activity was increased by preincubation of ECM with arachidonic acid (AA). A moderate decrease of the activity was obtained by incubation with longchain saturated, monoenoic and dienoic unsaturated fatty acids. A pronounced decrease of the inhibitor was obtained by incubation with di-homo-γ-linolenic acid (DHLA).
Paired combinations of AA with the other fatty acids in the incubation medium partially restored the inhibitor activity obtained by the separate FA.
The stimulation of the inhibitor by AA was dose dependent and high concentrations of AA reduced this activity.
The present study indicates that the quantity and quality of the plasma free fatty acids can affect the endothelial cells' ability to act as a non-thrombogenic surface.     

Unconjugated serum bile acids as a marker of small intestinal bacterial overgrowth

Non-invasive methods to detect small intestinal bacterial overgrowth often lack specificity in patients who have undergone an ileal resection or have an accelerated intestinal transit. Since elevated serum unconjugated bile acid levels have been found in patients with clinical signs of bacterial overgrowth, we studied the clinical value of unconjugated serum bile acids as a marker of small intestinal bacterial overgrowth. Patients with culture-proven bacterial overgrowth had significantly elevated fasting unconjugated serum bile acid levels (median and range: 4.5; 1.4-21.5 mumol l-1) as compared to healthy subjects (0.9; 0.3-1.7 mumol l-1, P less than 0.005), to persons with an accelerated intestinal transit (1.0; 0.3-1.9 mumol l-1, P less than 0.005) and to persons who have undergone an ileal resection (2.1; 0.7-3.6 mumol l-1, P less than 0.005). The same was true 30 and 60 min after ingestion of a Lundh meal. Serum unconjugated bile acid levels above 4 mumol l-1 were found in eight of 10 patients with culture-proven small intestinal bacterial overgrowth whereas serum levels above 4 mumol l-1 were found in none of the patients from the three control groups. These results suggest that determination of unconjugated serum bile acids is of clinical value in the evaluation of patients suspected of small intestine bacterial overgrowth.     

Characterization of plasma acylcarnitines in patients under valproate monotherapy using ESI-MS/MS

Objectives: The effect of administration of the antiepileptic drug valproate (VPA), on the composition of the plasma acylcarnitine profile (including free carnitine) was investigated.

Design and methods: Plasma samples were obtained from 18 individuals (13♂:5♀; 15–65y) on long-term treatment with VPA (resulting in plasma levels of 14.6–135.0 mg/L; therapeutic conc.: 40–100 mg/L). Acylcarnitines (AC) in plasma were quantified by electrospray tandem mass spectrometry (ESI-MS/MS).

Results: VPA was found to increase the levels (mean ± SD, μM) of 3-hydroxy-isovalerylcarnitine (0.10 ± 0.04; controls: 0.02–0.06), C14:2 acylcarnitine (0.11 ± 0.05; controls: 0.02–0.08), propylglutarylcarnitine (0.06 ± 0.05; controls: 0.00–0.04), and C18-0H-acylcarnitine (0.09 ± 0.05; controls: 0.00−0.04). The free carnitine (C) (42.2 ± 9.0; controls: 22.3–54.9) and the total carnitine (52.3 ± 10.1; controls: 26.5–73.6) were not significantly altered by VPA. Other AC (C2-C18, monounsaturated and hydroxylated) were all within the control range and especially no increase of C8 (valproyl) carnitine was observed. A positive correlation was found between the ratios [AC] / [C] (p < 0.05) or [long-chain AC (C10-C18)] / [C] (p < 0.09) with the plasma VPA concentration.

Conclusions: The unequivocal increase in 3-hydroxy-isovalerylcarnitine is consistent with the increase of 3-hydroxy-isovaleric acid observed in urine of VPA treated patients. This finding suggests an interaction mechanism of VPA with specific enzymes, namely involved in leucine metabolism. Adult patients under VPA monotherapy do not suffer from carnitine deficiency; the effect of the accumulating acylcarnitines is ill-defined.     

Incorporation of 5-fluorouracil into rat liver tumor and normal tissues and the liver nucleotide profile after administration by the hepatic artery during portal venous branch ligation

A therapeutic dose of labelled 5-fluorouracil (5-FU) was infused via the hepatic artery during 30 min with or without ligation of the left portal venous branch in Wistar rats with a secondary liver cancer in the left lateral lobe. After another 60 min, the incorporation of 5-FU into the acid soluble fraction (ASF), ribonucleic acid (RNA) and deoxyribonucleic acid (DNA), was determined in tumor, ligated and unligated liver lobes, small intestine, kidney, and bone marrow. The liver nucleotide profile was examined with isotachophoresis. Portal venous branch ligation (PVBL) caused the following changes, compared with the unligated control group: in the tumor, the incorporation of 5-FU into RNA and DNA decreased and the ratio RNA/acid-soluble fraction labelling decreased. The incorporation increased in intestinal and bone marrow RNA. It was unchanged in liver and kidney. The ratio of tumor to peripheral normal-tissue (small intestine, bone marrow, and kidney) labelling of RNA and DNA decreased. Liver nucleotides (F) UTP, (F)UDP-glucuronic acid, (F)UDP-N-acetylhexosamine, and NAD were lower in the ligated than in the unligated liver lobe. ATP and energy charge did not decrease significantly. In conclusion, PVBL in conjunction with hepatic arterial administration of 5-FU increased systemic drug exposure and possibly decreased hepatic tumor anabolism. It has not been examined how this interferes with the therapeutic effect.     

平衡型氨基酸透析液血液透析治疗临床应用观察

目的设计平衡型氨基酸透析液，应用于维持性血液透析患者常规血液透析治疗，观察临床应用效果。方法选用解放军总医院第一附属医院长期维持性血液透析患者29例，透析2～3次／w，实验采用自身交叉对照设计，使用碳酸氢盐透析液1月后，再应用平衡型氨基酸透析液1月，分别观察血液生化指标、血浆蛋白、肾功能以及炎症指标的变化。结果血浆氨基酸谱显示绝大部分游离氨基酸及总游离氨基酸含量均恢复到与健康人相仿的水平；平衡型氨基酸透析液治疗后尿素氮、肌酐、尿酸等。肾功能指标有不同程度的升高，但电解质及钙、磷没有明显的变化；血浆总蛋白（TP）、白蛋白（ALB）、前白蛋白（PreA）、转铁蛋白（Trf）等较前无明显变化，但β2微球蛋白（β2-MG）和超敏C反应蛋白（SCRP）则较应用前有明显的降低（P〈0．01和P〈0．05）。结论平衡型氨基酸透析液能减少维持性血液透析患者的氨基酸丢失，部分改善其体内的氨基酸代谢紊乱，同时有可能减轻体内的微炎症反应。     

A biochemical perspective on the use of tandem mass spectrometry for newborn screening and clinical testing

The first newborn screen was a clinical test to detect a disorder of the biochemistry of the amino acid, phenylalanine. This disorder, known as phenylketonuria, produces profound mental retardation if not detected and treated early in life. Early screening programs relied on inexpensive population screening techniques that have all but been replaced by more accurate analytical methods such as tandem mass spectrometry (MS/MS). MS/MS enables a multianalyte approach for detecting biochemical disorders such that a metabolic profile is obtained rather than a single analyte measurement. The metabolic profile has clearly shown improvements in the detection of diseases such as phenylketonuria and several new disorders arising from errors in fatty acid oxidation and organic acid metabolism. MS/MS is a powerful tool for accessing the metabolic status of a newborn and can detect both inborn metabolic errors as well as examine the effect of acquired diseases or pharmacologic intervention on intermediary metabolism.