Plasma elemental responses to red meat ingestion in healthy young males and the effect of cooking method.

PURPOSE: Elemental deficiencies are highly prevalent and have a significant impact on health. However, clinical monitoring of plasma elemental responses to foods remains largely unexplored. Data from in vitro studies show that red meat (beef) is a highly bioavailable source of several key elements, but cooking method may influence this bioavailability. We therefore studied the postprandial responses to beef steak, and the effects of two different cooking methods, in healthy young males. METHODS: In a randomized cross-over controlled trial, healthy males (n = 12, 18-25 years) were fed a breakfast of beef steak (270 ± 20 g) in which the meat was either pan-fried (PF) or sous-vide (SV) cooked. Baseline and postprandial blood samples were collected and the plasma concentrations of 15 elements measured by inductively coupled plasma-mass spectrometry (ICP-MS). RESULTS: Concentrations of Fe and Zn changed after meal ingestion, with plasma Fe increasing (p < 0.001) and plasma Zn decreasing (p < 0.05) in response to both cooking methods. The only potential treatment effect was seen for Zn, where the postprandial area under the curve was lower in response to the SV meal (2965 ± 357) compared to the PF meal (3190 ± 310; p < 0.05). CONCLUSIONS: This multi-element approach demonstrated postprandial responsiveness to a steak meal, and an effect of the cooking method used. This suggests the method would provide insight in future elemental metabolic studies to evaluate responses to meat-based meals, including longer-term interventions in more specifically defined cohorts to clearly establish the role of red meat as an important source of elements.

Relative quantification of albumin and fibrinogen modifications by liquid chromatography tandem mass spectrometry in the diagnosis and monitoring of acute pancreatitis.

The increasing availability of liquid chromatography tandem mass spectrometry (LC-MS/MS) in clinical laboratories provides the opportunity to replace or complement present underperforming immuno- and chemometric assays. Amylase and lipase show limited specificity and sensitivity for pancreatic inflammation and lack the capacity of monitoring the disease due to their short half-lives. Previous findings suggested that cleavage products of the pancreatic enzyme carboxypeptidase A could be a more suitable indicator for defining and classifying pancreatic inflammation. The plasma proteins albumin and ß-fibrinogen were digested with trypsin and truncated forms (des-Leu-albumin, and des-Gln-ß-fibrinogen) quantified against their non-truncated forms by LC-MS/MS. Four hundred fifty eight samples from 83 patients were used to evaluate the novel method and affirm its suitability for detecting acute pancreatitis. A robust, selective, precise and accurate LC-MS/MS method was set up to measure the proportion of truncated proteins. Reference ranges for the proportion of the truncated albumin and ß-fibrinogen were from 2% to 9% and 3% to 25%, respectively. Acute pancreatitis patients had values above these ranges and were distinctly separated from reference control individuals. The longer circulating half-lives of albumin and fibrinogen compared to pancreatic enzymes themselves provide the potential to diagnose pancreatitis more specifically over a longer time period, to monitor the course of the disease, and to track recurrent complications. The wide range of the proportion and the differential half-life of both truncated proteins could also be used for assessing the severity of pancreatitis.

Laboratory medicine best practice guideline: vitamins a, e and the carotenoids in blood.

Despite apparent method similarities between laboratories there appear to be confounding factors inhibiting uniform reporting and standardisation of vitamin assays. The Australasian Association of Clinical Biochemists (AACB) Vitamins Working Party, in conjunction with The Royal College of Pathologists of Australasia Quality Assurance Programs, has formulated a guideline to improve performance, reproducibility and accuracy of fat-soluble vitamin results. The aim of the guideline is to identify critical pre-analytical, analytical and post-analytical components of the analysis of vitamins A, E and carotenoids in blood to promote best practice and harmonisation. This best practice guideline has been developed with reference to the Centers for Disease Control and Prevention (CDC) "Laboratory Medicine Best Practices: Developing an Evidence-Based Review and Evaluation Process". The CDC document cites an evaluation framework for generating best practice recommendations that are specific to laboratory medicine. These 50 recommendations proposed herein, were generated from a comprehensive literature search and the extensive combined experience of the AACB Vitamins Working Party members. They were formulated based on comparison between an impact assessment rating and strength of evidence and were classified as either: (1) strongly recommend, (2) recommend, (3) no recommendation for or against, or (4) recommend against. These best practice recommendations represent the consensus views, in association with peer reviewed evidence of the AACB Vitamins Working Party, towards best practice for the collection, analysis and interpretation of vitamins A, E and carotenoids in blood.

Vitamin B1 and B6 method harmonization: comparison of performance between laboratories enrolled in the RCPA Quality Assurance Program.

OBJECTIVES: The RCPA Quality Assurance Program (RCPA QAP) offers monthly proficiency testing for vitamins A, B1, B6, ß-carotene, C and E to laboratories worldwide. A review of the results submitted for the whole blood vitamin B1/B6 sub-program revealed a wide dispersion. Here we describe the results of a methodology survey for vitamins B1 and B6. DESIGN AND METHODS: A questionnaire was sent to thirteen laboratories. Eleven laboratories were returning QAP results for vitamin B1 (thiamine diphosphate) and five were returning results for vitamin B6 (pyridoxal-5-phosphate). RESULTS: All nine respondents provided a clinical service for vitamins B1 and B6. HPLC with fluorescence detection was the most common method principle. For vitamin B1, six respondents used a commercial assay whilst three used in-house methods; whole blood was the matrix for all. For vitamin B6, five respondents used commercial assays and four used in-house assays. The choice of matrix for vitamin B6 varied with three respondents using whole blood and five using plasma for analysis. Sample preparation incorporated protein precipitation and derivatization steps. An internal standard was employed in sample preparation by only one survey respondent. CONCLUSIONS: The immediate result of this survey was the incorporation of plasma vitamin B6 into the RCPA QAP vitamin program. The absence of an internal standard in current vitamin B1 and B6 assays is a likely contributor to the wide dispersion of results seen in this program. We recommend kit manufacturers and laboratories investigate the inclusion of internal standards to correct the variability that may occur during processing.

A mass-spectroscopic method for measuring des-Leu albumin--a novel marker for chronic pancreatitis.

OBJECTIVES: Chronic pancreatitis is a progressive inflammatory disease leading to pancreatic insufficiency. The diagnosis of chronic pancreatitis is challenging, especially in early disease and the current tests have low sensitivity, may be invasive or have limited availability. We previously identified a truncated form of albumin lacking the C-terminal leucine, des-Leu albumin, which was present at high concentration in pancreatitis. We have developed a liquid-chromatography tandem-mass spectrometry (LC-MS/MS) method for measuring this peptide and make some preliminary observations on patient samples. METHODS: Serum samples from patients with established pancreatitis and controls were obtained. Diluted serum samples or prepared standards were digested with trypsin. Aliquots of the digest were separated on a reversed-phase column using water:acetonitrile:formic acid mobile-phase with tandem-mass spectrometry detection. Percentage composition of des-Leu albumin was determined from a response curve. RESULTS: The C-terminal peptide, LVAASQAALG- of des-Leu albumin was identified by m/z 901→725, wild type albumin by m/z 1014→825. Additional fragments were monitored as internal reference for digestion and sample integrity. Inter-assay imprecision was estimated at 10%. The percentage composition of des-Leu albumin segregated with the diagnosis of established pancreatitis with median levels of des-Leu albumin of 68% in patients compared to 5% in controls. CONCLUSIONS: Des-Leu albumin is a promising novel biomarker for chronic pancreatitis. It allowed clear discrimination of patients with pancreatitis from controls and its long half-life may facilitate monitoring of disease activity. The method described could readily be undertaken in modern clinical chemistry laboratories and will form the basis for further study.