Proposal for a new N-stage classification system for intrahepatic cholangiocarcinoma.

Background: The number of metastatic lymph nodes (MLNs) is not considered in the nodal status (N classification) of intrahepatic cholangiocarcinoma (ICC) in the current 8thEdition of the American Joint Committee on Cancer (AJCC) staging system. The aim of this study was to find out the optimal cut-off point based on the number of MLNs and establish a modified AJCC staging system for ICC according to the new N category. Methods: A total of 675 ICC patients diagnosed between 2004 and 2015 were retrieved from the Surveillance, Epidemiology and End Results (SEER) database. The optimal cut-off value of MLNs affecting survival was determined by X-tile software. The relative discriminative power was assessed by Harrell's concordance index (C-index) and Akaike information criterion (AIC). Results: The proposed new nodal category subdivided patients into three groups (N0, no MLN; N1, 1-3 MLNs; and N2, ≥ 4 MLNs) with significantly different overall survival (P < 0.001). Multivariable analysis revealed that the new nodal category was an independent prognostic factor (P < 0.001). Both the C-index and AIC for our modified staging system were better than those for the 8th AJCC edition (0.574 [95% confidence interval 0.533-0.615] versus 0.570 [95% confidence interval 0.527-0.613], and 853.30 versus 854.21, respectively). Conclusion: The modified AJCC staging system based on the number of MLNs may prove to be a useful alternative for predicting survival of ICC patients in clinical practice.

Lenvatinib as First-Line Treatment for Unresectable Hepatocellular Carcinoma: A Systematic Review and Meta-Analysis.

Lenvatinib was approved in 2018 as a first-line treatment for patients with unresectable hepatocellular carcinoma (HCC). This systematic review and meta-analysis aimed to provide the most updated evidence about the efficacy and safety of lenvatinib as a first-line treatment for unresectable HCC. An electronic search of the PubMed database, Web of Science, Embase, and Cochrane Library was undertaken to identify all relevant studies up to May 2022. The pooled effect sizes were calculated based on the random-effects model. One phase III randomized controlled trial and 23 retrospective studies of 2438 patients were eligible for analysis. For patients treated with lenvatinib as first-line treatment, the pooled median overall survival (OS), median progression-free survival (PFS), 1-year OS rate, 1-year PFS rate, objective response rate (ORR), and disease control rate (DCR) were 11.36 months, 6.68 months, 56.0%, 27.0%, 36.0% and 75.0%, respectively. Lenvatinib showed a significantly superior efficacy compared with sorafenib (HR for OS, 0.85 and HR for PFS, 0.72; OR for ORR, 4.25 and OR for DCR, 2.23). The current study demonstrates that lenvatinib can provide better tumor responses and survival benefits than sorafenib as a first-line treatment for unresectable HCC, with a comparable incidence of adverse events.

[Retracted] MicroRNA­744 inhibits tumor cell proliferation and invasion of gastric cancer via targeting brain­derived neurotrophic factor.

Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the cell migration assay data shown in Fig. 2D were strikingly similar to data that had appeared in different form in other articles by different authors. Owing to the fact that the contentious data in the above article had already been published elsewhere, or were already under consideration for publication, prior to its submission to Molecular Medicine Reports, the Editor has decided that this paper should be retracted from the Journal. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a reply. The Editor apologizes to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 16: 5055­5061, 2017; DOI: 10.3892/mmr.2017.7167].

Differentiating ultra-high temperature milk and reconstituted milk using an untargeted peptidomic approach with chemometrics.

An untargeted peptide profiling based on ultra-performance liquid chromatography quadrupole time-of flight mass spectrometry with chemometrics was performed to differentiate ultra-high temperature processed milk and reconstituted milk. Thirty-three marker peptides were identified, primarily released from the C- or N-terminal of ß-casein and αs1-casein. These peptides were produced by heating and protease hydrolysis. Additional heating and storage time experiments showed that the level of 18 marker peptides increased with heat load and storage time, whereas 15 peptides were solely influenced by heat load. The peptides from ß-casein showed higher sensitivity to thermal stress compared to those from αs1-casein. Additionally, eight modified peptides of casein were identified as indicators of milk thermal processing. The identified marker peptides can distinguish ultra-high temperature processed milk and reconstituted milk, and are suitable for monitoring heating processes and storage of milk.

Milk lipids characterization in relation to different heat treatments using lipidomics.

Heat treatment is an important processing technique related to milk quality and nutritional value in the dairy industry. In this study, changes in milk lipids in response to different heat treatments were comprehensively characterized using a lipidomic approach. Ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) were used to identify and quantify 29 classes and 788 different lipids. In general, heat treatment promoted milk lipid hydrolysis and oxidation; in particular, ultra-high temperature (UHT) treatment resulted in more phospholipid hydrolysis than did pasteurization and extended shelf-life (ESL) treatment. Heat treatment resulted in further lipid oxidation reactions and a reduction in the amount of mild oxidation products. Moreover, the levels of lysophospholipids and free fatty acids (including oxidized free fatty acids) can be used to distinguish UHT-treated milk. In turn, oxidized phosphatidylcholine, oxidized phosphatidylethanolamine, ether-linked phosphatidylethanolamine, diacylglycerol, triacylglycerol, and oxidized triacylglycerol can be used to differentiate raw, pasteurized, and ESL milk. These biomarkers can potentially be used in the dairy industry to monitor the degree and method of heat treatment of milk.

Conversion surgery for initially unresectable pancreatic ductal adenocarcinoma following induction therapy: a systematic review of the published literature.

Patients with unresectable pancreatic ductal adenocarcinoma (UR-PDAC) are traditionally treated with palliative chemotherapy. The aim of this study was to evaluate the safety and efficacy of conversion surgery for initially UR-PDAC following induction therapy. The PubMed and Embase databases were systematically searched for eligible studies published between January 2000 and October 2020. Thirty-two series involving 1270 patients with 1056 locally advanced (LA) disease and 214 distant metastases were reviewed. The median mortality and morbidity was 0% (range 0-10%) and 47.1% (range 8.6-93.3%), respectively. Lymph-node negativity, negative resection margin and pathological complete response were observed in a median of 62.9% (38.5-90.9%), 84.4% (32.8-100%) and 6.7% (0-45.8%) of the specimens. The median survival was 32 (16.4-63.9) months with a 3-year survival rate of 47% (22-80%). Meta-analysis demonstrated that conversion surgery of initially UR-PDAC was associated with a significantly improved survival (hazard ratio [HR] = 0.55; 95% confidence intervals (CI) 0.45-0.66, P < 0.001). There was no significant difference in survival between the group with LA disease and that with distant metastases after conversion surgery (HR = 0.96; 95% CI 0.72-1.28, P = 0.790). Conversion surgery improved long-term survival of patients with initially UR-PDAC who had favorable response to induction therapy.

Dietary Conjugated Linoleic Acid Modulates the Hepatic Circadian Clock Program via PPARα/REV-ERBα-Mediated Chromatin Modification in Mice.

Scope: Disruptions of circadian rhythm cause metabolic disorders and are closely related to dietary factors. In this study, we investigated the interplays between the dietary conjugated linoleic acid (CLA)-induced hepatic steatosis and the circadian clock regulation, in association with lipid homeostasis. Methods and Results: Exposure of mice to 1.5% dietary CLA for 28 days caused insulin resistance, enlarged livers, caused hepatic steatosis, and increased triglyceride levels. Transcriptional profiling showed that hepatic circadian clock genes were significantly downregulated with increased expression of the negative transcription factor, REV-ERBα. We uncovered that the nuclear receptor (NR) PPARα, as a major target of dietary CLA, drives REV-ERBα expression via its binding to key genes of the circadian clock, including Cry1 and Clock, and the recruitment of histone marks and cofactors. The PPARα or REV-ERBα inhibition blocked the physical connection of this NR pair, reduced the cobinding of PPARα and REV-ERBα to the genomic DNA response element, and abolished histone modifications in the CLA-hepatocytes. In addition, we demonstrated that CLA promotes PPARα driving REV-ERBα transcriptional activity by directly binding to the PPAR response element (PPRE) at the Nr1d1 gene. Conclusions: Our results add a layer to the understanding of the peripheral clock feedback loop, which involves the PPARα-REV-ERBα, and provide guidance for nutrients optimization in circadian physiology.

Effects of the Oxygen Content and Light Intensity on Milk Photooxidation Using Untargeted Metabolomic Analysis.

Photooxidation is the main cause of milk quality deterioration during processing and transportation. Oxygen and light are responsible for dairy milk photooxidation. The objective of this study was to determine the characteristic metabolites after photooxidation and how they are affected by oxygen exposure and light intensity. Ultra-performance liquid chromatography coupled with triple time-of-flight mass spectrometry and multivariate data analysis were used for the high-throughput evaluation of milk photooxidation. Four products were identified as biomarkers: uric acid, riboflavin, lumichrome, and indole-3-carboxaldehyde. Afterward, the effects of oxygen content and light intensity on the biomarkers were investigated, and a sensory evaluation was performed. Both oxygen exposure and light intensity affected the contents of photooxidation biomarkers in milk samples. The sensory score correlated well with the oxygen content but not with the light intensity. The untargeted metabolomic method was an effective tool to identify biomarkers for milk photooxidation evaluation.

Long-term whole-grain rye and wheat consumption and their associations with selected biomarkers of inflammation, endothelial function, and cardiovascular disease.

BACKGROUND/OBJECTIVES: Whole-grain (WG) intake has been associated with a lowered risk of developing type 2 diabetes, cardiovascular disease, and some cancers in epidemiological studies. Reduced subclinical inflammation could be one important mechanism behind such associations. This study investigated whether high long-term WG rye and wheat intakes were associated with lower concentrations of biomarkers of inflammation, endothelial function, and protein biomarkers associated with cardiovascular disease. SUBJECTS/METHODS: We assessed WG intake by food frequency questionnaire (FFQ) and by measuring alkylresorcinols (ARs) in plasma and adipose tissue, respectively. Selected biomarkers in free-living 109 women and 149 men were analyzed from two clinical subcohort studies (Swedish Mammography Cohort-Clinical (SMC-C) and Cohort of Swedish Men-Clinical (COSM-C), respectively. Total WG rye and wheat (WGRnW) and the ratio of WG rye to WG rye and wheat (WGR/WGRnW) were estimated from FFQs. ARs were measured in plasma and adipose tissue by gas chromatography-mass spectrometry (GC-MS) and the biomarkers by ELISA. RESULTS: We found no consistent associations between WG intake assessed by different methods and the selected biomarkers. However, WGRnW intake was inversely associated with cathepsin S (P-trend < 0.05) and total AR and C17:0/C21:0 in plasma were inversely associated with the endostatin concentration (P-trend < 0.05) adjusted for BMI, age, and sex. CONCLUSION: The results give limited support to the hypothesis that a high WG wheat and rye intake is associated with lower concentrations of common biomarkers of inflammation and CVD that have previously been reported inversely associated with WG intake or an overall healthy lifestyle.

Establishing an untargeted-to-MRM liquid chromatography-mass spectrometry method for discriminating reconstituted milk from ultra-high temperature milk.

The counterfeit use of reconstituted milk to simulate fresh milk in some countries is largely driven by profiteering; hence, establishing a robust market-surveillance method is an important objective. In this study, an untargeted metabolomics approach that uses ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was established to discover biomarkers that can be used to classify UHT and reconstituted milk. Furthermore, 36 positive- and 24 negative-ionization mode features were selected as candidate markers to establish a UPLC-tandem mass spectrometry (UPLC-MS/MS) multiple reaction monitoring (MRM) method. The support vector machine (SVM) method was used to process the responses of the selected potential markers, and 100% classifiability was observed. The marker metabolites could be divided into three categories by hierarchical clustering analysis: peptides, lipids, and nucleic acids. The results suggest that the untargeted-to-MRM metabolomics method is an effective tool for distinguishing between UHT and reconstituted milk.

Determination of the levels of tryptophan and 12 metabolites in milk by liquid chromatography-tandem mass spectrometry with the QuEChERS method.

Tryptophan and metabolites have important biological functions in humans. Milk is an important source of tryptophan intake. In this study, we developed a method to detect levels of tryptophan and 12 metabolites in milk. The analytes were extracted by using the QuEChERS (quick, easy, cheap, effective, rugged, and safe) procedure and analyzed by liquid chromatography-tandem mass spectrometry with electrospray ionization. The proposed method resulted in suitable accuracy (standard deviation ≤10.31%) and high sensitivity (the limits of quantification were between 0.05 and 5 ng/mL). Recoveries were in the range of 44 to 126%. Finally, the developed method was successfully applied to compare the content of tryptophan and metabolites in 4 milk products produced by different processes: pasteurized milk, UHT milk, milk powder, and yogurt. The results of partial least squares-discriminant analysis (PLS-DA) showed that different types of processed milk could be distinguished clearly according to the method used here. The determined tryptophan and metabolites levels in milk can provide a new reference for evaluation of milk.

Long-Term Whole Grain Wheat and Rye Intake Reflected by Adipose Tissue Alkylresorcinols and Breast Cancer: A Case-Cohort Study.

Whole grain rye (WGR) and whole grain wheat (WGW) have been suggested to protect against the development of breast cancer. In this study, we estimated long-term intake of WGR and WGW, using both a food frequency questionnaire (FFQ) and alkylresorcinol concentrations in adipose tissue biopsies, in relation to the risk of developing invasive breast cancer in a case-cohort study (n = 414 in the case group, n = 933 in the subcohort group) on the Danish "Diet, Cancer and Health" cohort. The median follow-up time of the subcohort was 5.3 years. Total WGR and WGW intake estimated with FFQ or reflected by total alkylresorcinol concentration in adipose tissue was not significantly associated with risk of breast cancer. However, after adjustment for total WGR and WGW intake, women in the highest quartile of relative WGR intake, reflected by the alkylresorcinol C17:0/C21:0 ratio, had a higher risk of overall breast cancer and estrogen-receptor-positive (ER+) breast cancer than women in the lowest quartile of relative WGR intake, while the risk of estrogen-receptor-negative (ER-) breast cancer incidence was unaffected. Similar results were obtained with the FFQ data. Based on these data, further investigation of the role of specific grain types in reducing or increasing breast cancer risk, and their overall impact on health, is warranted.

Freeze-dried bilberry (Vaccinium myrtillus) dietary supplement improves walking distance and lipids after myocardial infarction: an open-label randomized clinical trial.

Bilberries, Vaccinium myrtillus, have a high content of phenolic compounds including anthocyanins, which could provide cardiometabolic health benefits following acute myocardial infarction (AMI). We hypothesized that standard medical therapy supplemented with freeze-dried bilberry after AMI would have a more beneficial effect on cardiovascular risk markers and exercise capacity than medical therapy alone. Patients were allocated in a 1:1 ratio within 24 hours of percutaneous coronary intervention in an 8-week trial either to V myrtillus powder (40 g/d, equivalent to 480 g fresh bilberries) and standard medical therapy or to a control group receiving standard medical therapy alone. High-sensitivity C-reactive protein and exercise capacity measured with the 6-minute walk test were the primary biochemical and clinical end points, respectively. Fifty subjects completed the study. No statistically significant difference in high-sensitivity C-reactive protein was detected between groups. The mean 6-minute walk test distance increased significantly more in the bilberry group compared to the control group: mean difference 38 m at follow-up (95% confidence interval 14-62, P = .003). Ex vivo oxidized low-density lipoprotein was significantly lowered in the bilberry group compared to control, geometric mean ratio 0.80 (95% confidence interval 0.66-0.96, P = .017), whereas total cholesterol and low-density lipoprotein cholesterol did not differ significantly between groups. Anthocyanin-derived metabolites in blood increased significantly in the bilberry group during the intervention and were different after 8 weeks between the bilberry group and control. Findings in the present study suggest that bilberries may have clinically relevant beneficial effects following AMI; a larger, double-blind clinical trial is warranted to confirm this.

Evaluation of alkylresorcinols in adipose tissue biopsies as a long-term biomarker of whole-grain wheat and rye intake in free-living Swedish men and women.

OBJECTIVE: Wheat and rye, the most consumed whole grains (WG) in the Nordic countries, contain alkylresorcinols (AR) in their bran. AR concentrations in human adipose tissue might reflect long-term WG rye and wheat intake. We aimed to evaluate AR concentrations in adipose tissue biopsies as a long-term biomarker of WG wheat and rye intake in free-living Swedish men and women. DESIGN: Cross-sectional study. AR concentrations in adipose tissue biopsies were analysed and compared with long-term WG intake assessed by three FFQ (repeated over a period of 14 years in men, 17 years in women) and with plasma AR concentrations. SETTING: The Cohort of Swedish Men between 1997 and 2010 and the Swedish Mammography Cohort between 1987 and 2003, Sweden. SUBJECTS: Men (n 149) and women (n 109). RESULTS: Long-term WG rye intake estimated with repeated FFQ correlated (r=0·31-0·41, P<0·01) with adipose-tissue AR concentrations, while WG wheat intake correlated only weakly (r=0·17-0·33, P<0·05). Total AR concentration in adipose tissue was 61 % lower in women than in men at similar energy-adjusted WG wheat and rye intakes, but plasma concentrations were similar. AR concentrations in adipose tissue correlated well with plasma concentrations (r=0·49-0·81, P<0·001). CONCLUSIONS: AR in adipose tissue reflected long-term WG rye but not WG wheat intake, probably due to poor precision in estimating WG wheat intake by FFQ. AR in adipose tissue appears promising as a biomarker of long-term WG rye intake but should be adjusted for sex.

MicroRNA­744 inhibits tumor cell proliferation and invasion of gastric cancer via targeting brain­derived neurotrophic factor.

Gastric cancer is the fourth most common malignancy and the third leading cause of cancer­associated deaths worldwide. It has previously been demonstrated that microRNAs (miRNAs) are actively involved in the pathogenesis of gastric cancer. Therefore, miRNAs have been proposed as promising therapeutic targets in gastric cancer patients. MiR­744 is aberrantly expressed in different types of human cancer. However, the expression pattern and biological roles of miR­744 in gastric cancer remain unknown. The present study demonstrated that miR­744 expression was low in gastric cancer tissues and cell lines. Low expression levels of miR­744 was significantly correlated with lymph node metastasis, invasive depth and TNM staging in gastric cancer patients. The restoration of miR­744 expression inhibited cell proliferation and invasion in vitro. Bioinformatic prediction, luciferase reporter assay, reverse transcription­quantitative polymerase chain reaction and western blot analysis verified that brain­derived neurotrophic factor (BDNF) is a direct target of miR­744 in gastric cancer cells. Furthermore, BDNF was upregulated in gastric cancer tissues and inversely correlated with miR­744 expression. Furthermore, enforced BDNF expression reversed the tumor­suppressing effects of miR­744 on the proliferation and invasion of gastric cancer cells, indicating that BDNF is a functional mediator of miR­744 in gastric cancer. The present study suggests that miR­744 is a potential prognostic biomarker and treatment target in gastric cancer patients.

Pir-B inhibits the DC function and disturbs the Th17/Treg balance in lung cancer murine model.

Paired immunoglobulin-like receptor B (Pir-B) was an inhibitory receptor expressed on the surfaces of dendritic cells (DCs). Pir-B inhibit T helper (Th) 1 response and induce Th2 cell differentiation, leading to the imbalance of Th1/Th2 cells. However, the role and potential mechanism of Pir-B on the balance of Th17/regulatory T cells (Tregs) is still largely unknown in lung cancer murine model. In the present study, the DC function and Th17/Treg balance were destroyed during the progression of lung cancer and this was accompanied by an increased expression of Pir-B. After transfection with Pir-B siRNA or administration of IL-6 in vitro, the decreased response of Th17 cells were restored, whereas the augmented differentiation of Tregs was diminished. Further, the transfer of Pir-B silenced DCs or the injection of IL-6 in vivo increased Th17 response and decreased Treg differentiation. Our study has demonstrated that Pir-B inhibits the DC function and disturbs the Th17/Treg balance via IL-6 pathway during the progression of lung cancer, contributing to inhibited antitumor immunity.

Targeted metabolomics reveals differences in the extended postprandial plasma metabolome of healthy subjects after intake of whole-grain rye porridges versus refined wheat bread.

SCOPE: We previously found that whole-grain (WG) rye porridges suppressed appetite and improved glucose metabolism. This study aimed to investigate potential plasma metabolites that may be related to differences in those appetite and glucose responses. METHODS AND RESULTS: Twenty-one health subjects consumed six isocaloric breakfasts in a randomized cross-over study. Plain WG rye porridges (40 and 55 g), rye porridge enriched with different inulin: gluten proportions (9:3 g; 6:6 g; 3:9 g), and a 55 g refined wheat bread (control) were served as part of complete breakfast, followed by a standardized lunch. NMR metabolomics assessed 36 plasma metabolites and short chain fatty acids were measured by GC-MS from baseline up to 8 h. Pre-lunch plasma essential amino acids reflected protein composition and post-lunch plasma short chain fatty acids varied with fiber content in breakfasts. No correlations were observed between measured metabolites and glucose, insulin, or appetite responses. CONCLUSIONS: Differences in protein and fiber contents in breakfasts altered postprandial plasma amino acids and short chain fatty acids, respectively, but were unrelated to appetite and glucose responses. Further studies are warrant to identify the underlying mechanisms for the beneficial effects on appetite and second meal glucose responses after rye-based foods.

Effects of transcutaneous acupoint electrical stimulation on the imbalance of Th1, Th2, Th17 and Treg cells following thoracotomy of patients with lung cancer.

An imbalance in the various T lymphocytes, including T-helper (Th)1, Th2 and Th17 cells, and regulatory T (Treg) cells, has been associated with immune dysfunction, and may occur following thoracotomy of patients with lung cancer. The use of transcutaneous acupoint electrical stimulation (TAES) has previously been demonstrated to exert immunoregulatory effects; therefore, the present study aimed to evaluate whether TAES was able to attenuate postoperative immune suppression in patients with lung cancer. Thoracic surgical patients with lung cancer (n=27) underwent TAES (frequency, 2/100 Hz; intensity, 4-12 mA) at the bilateral large intestine 4, pericardium 6, small intestine 3 and San Jiao 6 acupuncture points for 30 min, prior to incision, and at 20, 44, 68, 92 and 116 h following thoracotomy. The number of Th1, Th2, Th17 and Treg cells, and the protein and mRNA expression levels of related cytokines were measured by flow cytometry, ELISA and polymerase chain reaction, respectively. The balance of Th1, Th2, Th17 and Treg cells in the peripheral blood of patients with lung cancer was disrupted following thoracotomy. TAES administration increased the percentage of Th1 and Th17 cells, the protein expression levels of interleukin (IL)-2 and interferon-γ, the mRNA expression levels of T-bet and RAR-related orphan receptor-γt, and decreased the percentage of Th2 cells, IL-10 protein expression levels, and GATA binding protein 3 mRNA expression levels. The results of the present study demonstrated that TAES was able to partially attenuate the postoperative immune depression of patients with lung cancer, by regulating the balance of Th1, Th2, Th17 and Treg cells, and the expression levels of related cytokines and transcription factors; therefore, TAES may be considered to be a promising strategy for treating postoperative immune dysfunction in patients with lung cancer.

Validation of Reported Whole-Grain Intake from a Web-Based Dietary Record against Plasma Alkylresorcinol Concentrations in 8- to 11-Year-Olds Participating in a Randomized Controlled Trial.

BACKGROUND: Whole-grain (WG) intake is important for human health, but accurate intake estimation is challenging. Use of a biomarker for WG intake provides a possible way to validate dietary assessment methods. OBJECTIVE: Our aim was to validate WG intake from 2 diets reported by children, using plasma alkylresorcinol (AR) concentrations, and to investigate the 3-mo reproducibility of AR concentrations and reported WG intake. METHODS: AR concentrations were analyzed in fasting blood plasma samples, and WG intake was estimated in a 7-d web-based diary by 750 participants aged 8-11 y in a 2 school meal × 3 mo crossover trial. Reported WG intake and plasma AR concentrations were compared when children ate their usual bread-based lunch (UBL) and when served a hot lunch meal (HLM). Correlations and cross-classification were used to rank subjects according to intake. The intraclass correlation coefficients (ICCs) between subjects' measurements at baseline and after the UBL were used to assess reproducibility. RESULTS: Correlations between reported WG wheat + rye intake and plasma AR were 0.40 and 0.37 (P < 0.001) for the UBL and the HLM diets, and 78% and 77% were classified in the same or adjacent quartiles for the UBL and HLM diets, respectively. The ICC over 3 mo was 0.47 (95% CI: 0.38, 0.55) for plasma total ARs and 0.64 (95% CI: 0.58, 0.70) for reported WG intake. Correlations were higher when using the AR C17:0 homolog as a biomarker, reflecting rye intake instead of plasma total ARs [UBL: r = 0.47; HLM: r = 0.43, P < 0.001; ICC = 0.51 (95% CI: 0.43, 0.59)]. CONCLUSIONS: Self-reported WG wheat + rye intake among children showed moderate correlations with plasma AR concentrations. Substantial intraindividual variation was found in WG intake and plasma AR concentrations. The AR homolog C17:0 may be used as a biomarker for WG intake when the WG intake primarily comes from rye as in the present study. This trial was registered at clinicaltrials.gov as NCT01457794.

Determination of alkylresorcinols and their metabolites in biological samples by gas chromatography-mass spectrometry.

High throughput GC-MS methods for quantification of alkylresorcinols (AR), biomarkers of whole grain wheat and rye intake, in plasma and adipose tissue and their metabolites in urine were developed and optimised. Alkylresorcinols in plasma (200µL) and adipose tissues (10-50mg) were extracted with diethyl ether, whereas main AR metabolites such as DHBA and DHPPA and newly identified metabolites in urine (50µL) were extracted with ethyl acetate after enzymatic deconjugation. All extracts were purified on OASIS-MAX solid phase extraction cartridges. Plasma and adipose tissue sample extracts were then derivatised with trifluoroacetic anhydride and reconstituted in undecane, whereas AR metabolites in urine samples were derivatised with BSTFA+TMCS (99:1, v/v, 100µL). Prepared samples were quantified by GC-MS (EI-SIM). Analysis of all compounds in the different matrices showed good selectivity, sensitivity, linearity, precision (<15% within and between batches), adequate recovery (75-108%), and short total run time (10-12min). The methods developed are applicable to large-scale sample sets such as epidemiological studies.