Urinary Excretion Of Metabolites Research Articles

A Phase 1 Randomized, Placebo-Controlled Study Evaluating the Safety, Tolerability, and Pharmacokinetics of Enteric-Coated Stabilized Sulforaphane (SFX-01) in Male Participants.

Sulforaphane (SFN) is a naturally occurring isothiocyanate associated with various health benefits, including reduced cancer risk, and has been extensively explored as a potential therapeutic. However, its inherent instability presents challenges in formulation, storage, and administration as a medicinal product. SFX-01 (Sulforadex®) is a patented synthetic form of d,l-SFN stabilized within a biologically inert alpha-cyclodextrin complex. The safety, tolerability, and pharmacokinetics of an enteric-coated tablet formulation of SFX-01 were evaluated in a randomized, double-blind, placebo-controlled, dose-escalation study [300mg once daily (46.2mg SFN), 300mg twice daily or 600mg once daily (92.4mg SFN)] over 7days in healthy male participants. Treatment-emergent adverse events (TEAEs) occurred in 94% of participants who received SFX-01 and were most commonly gastrointestinal events, which were mild in severity and related to treatment. Following ingestion of SFX-01 tablets, SFN was rapidly absorbed, with a timescale consistent with the enteric coating, and subsequently metabolized. The observed peak blood concentration (Cmax) for the sum of SFN and metabolites (total thiol) across all treatment cohorts ranged from 0.43 to 2.12µmol/L in 3-6h. Cmax data were considered inconclusive with respect to dose-proportionality and there was minimal evidence of accumulation of SFN and metabolites. Urinary excretion of SFN and individual metabolites ranged from<1 to 41%, and the proportion excreted did not appear to be influenced by the dose. This study demonstrated the safety and tolerability of SFX-01 over 7days and indicated that the pharmacokinetic behavior of SFX-01 enteric-coated tablets was in line with expectations. European Union Drug Regulating Authorities Clinical Trials Database (EudraCT) number: 2022-001601-43; ISRCTN Study Registration number: ISRCTN9628565.

Renal excretion of 1,2-dihydroxynaphthalene (DHN) in firefighting instructors after exposure to polycyclic aromatic hydrocarbons (PAHs) during live fire training

Exposure of firefighting instructors to polycyclic aromatic hydrocarbons (PAHs) such as naphthalene is unavoidable during live fire training. The study aimed to investigate naphthalene uptake by measuring the urinary excretion of the naphthalene metabolite 1,2-dihydroxynaphthalene (DHN), to describe the DHN elimination kinetics and to evaluate the results by comparison to further biomarkers of PAH exposure. N = 6 male non-smoking firefighting instructors completed five training sessions each in a residential fire simulation unit under respiratory protection. All participants provided two urine samples before and another seven samples within an 18-h-interval after each session. DHN was detected by gas chromatography/tandem mass spectrometry (GC–MS/MS) in all samples (n = 237) with median concentrations ranging from 3.3 µg/g crea. (range 0.9–10.2) before exposure to 134.2 µg/g crea. (43.4–380.4) post exposure. Maximum elimination found 3.3 h (median) after onset of exposure decreased with a mean half-life of 6.6 h to 27.1 µg/g crea. (15.7–139.5) 18 h after training. DHN sensitively indicated a presumed dermal naphthalene intake during training, showing similar elimination kinetics like other naphthalene metabolites. Internal exposure of the participants transiently exceeded exposures determined for non-smokers in the general population, but was lower than at other workplaces with PAH exposure. Despite limited uptake, accumulation is possible with daily exposure.

Chronic consumption of orange juice modifies urinary excretion of flavanone gut-derived metabolites through gut microbiota modulation

The metabolism and absorption of citrus flavanones are intrinsically linked to the gut microbiota, creating a bidirectional relationship where these compounds influence the microbiome, and in turn, the microbiota affects their metabolism. This study evaluates the effect of acute and chronic consumption of orange juice (OJ) on the urinary excretion of gut-derived flavanone metabolites and the gut microbiota. Health volunteers ingested 500 mL of OJ for 60 days in a single-arm human intervention study. Blood and feces were collected at baseline and after 60 days, with an additional 24-hour urine collection after a single dose on day 1 and day 63. LC-MS/MS analyzed urinary flavanone metabolites, while 16S rRNA sequencing characterized gut microbiota. Total urinary hesperetin conjugates excretion significantly decreased over 60 days, while gut-derived total phenolic acids, particularly three hydroxybenzoic acids, increased. Moreover, the heterogeneity of the total amount of flavanone conjugates, initially categorizing individuals into high-, medium- and low- urinary excretor profiles, shifted towards medium-excretor, except for five individuals who remained as low-excretors. This alteration was accompanied by a decrease in intestinal β-glucosidase activity and a shift in the relative abundance of specific genera, such as decreases in Blautia, Eubacterium hallii, Anaerostipes, and Fusicatenibacter, among which, Blautia was associated with higher urinary flavanone conjugates excretion. Conversely, an increase in Prevotella was observed. In summary, chronic OJ consumption induced transient changes in gut microbiota and altered the metabolism of citrus flavanones, leading to distinct urinary excretion profiles of flavanone metabolites.

Urinary Prostaglandin E2 Excretion and the Risk of Cardiovascular and Kidney Disease.

Inhibition of prostaglandin synthesis by nonsteroidal anti-inflammatory drugs is associated with cardiovascular mortality and kidney disease. This study hypothesizes that urinary prostaglandin E2 (PGE2) and PGE2 metabolite (PGEM) excretions are markers of cardiovascular and kidney health, because they reflect both systemic and kidney-derived PGE2 production. PGE2 and PGEM were measured in spot urine samples from 2291 participants (≥55 years old)of the population-based Rotterdam Study. Urinary PGE2 and PGEM excretions were analyzed using linear regression analyses to identify cross-sectional associations with cardiovascular risk factors and baseline estimated glomerular filtration rate (eGFR). Longitudinal associations with cardiovascular mortality and kidney outcomes (eGFR <60 or <45 mL/min per 1.73 m2 and the composite outcome 40% eGFR loss or kidney failure) were assessed with Cox regression. Urinary PGE2 and PGEM excretions were higher with increasing age, lower eGFR, smoking, diabetes, and albuminuria. A 2-fold higher urinary PGE2 and PGEM excretion was associated with a higher risk of cardiovascular mortality (28 825 patient-years; 160 events; PGE2 hazard ratio [HR], 1.27, [95% CI, 1.06-1.54]; PGEM HR, 1.36 [95% CI, 1.10-1.67]). Higher PGE2 excretions were also associated with a higher risk of incident eGFR <60 mL/min per 1.73 m2 (31 530 person-years; 691 events; HR, 1.13 [95% CI, 1.02-1.25]) with similar HRs for the other kidney outcomes. Urinary PGE2 and PGEM excretions are novel markers for the presence and progression of cardiovascular and kidney disease. Future studies should address whether these associations are causal and can be targeted to improve cardiovascular and kidney outcomes.

A phase 2 trial investigating the efficacy and safety of the mPGES-1 inhibitor vipoglanstat in systemic sclerosis-related Raynaud's.

Our objective was to test the hypothesis, in a double-blind, placebo-controlled study that vipoglanstat, an inhibitor of microsomal prostaglandin E synthase-1 (mPGES-1) which decreases prostaglandin E2 (PGE2) and increases prostacyclin biosynthesis, improves RP. Patients with systemic sclerosis (SSc) and ≥7 RP attacks during the last screening week prior to a baseline visit were randomised to four weeks treatment with vipoglanstat 120 mg or placebo. A daily electronic diary captured RP attacks (duration and pain) and Raynaud's Condition Score, with change in RP attacks/week as primary end point. Cold challenge assessments were performed at baseline and end of treatment. Exploratory endpoints included patients' and physicians' global impression of change, Assessment of Scleroderma-associated Raynaud's Phenomenon questionnaire, mPGES-1 activity, and urinary excretion of arachidonic acid metabolites. Sixty-nine subjects received vipoglanstat (n = 33) or placebo (n = 36). Mean weekly number of RP attacks (baseline; vipoglanstat 14.4[SD 6.7], placebo 18.2[12.6]) decreased by 3.4[95% CI -5.8;-1.0] and 4.2[-6.5;-2.0] attacks per week (p= 0.628) respectively. All patient reported outcomes improved, with no difference between the groups. Mean change in recovery of peripheral blood flow after cold challenge did not differ between the study groups. Vipoglanstat fully inhibited mPGES-1, resulting in 57% reduction of PGE2 and 50% increase of prostacyclin metabolites in urine. Vipoglanstat was safe and well tolerated. Although vipoglanstat was safe, and well tolerated in a dose achieving full inhibition of mPGES-1, it was ineffective in SSc-related RP. Further development and evaluation of vipoglanstat will therefore be in other diseases where mPGES-1 plays a pathogenetic role.

Urine steroid metabolomics as a diagnostic tool in primary aldosteronism

Primary aldosteronism (PA) causes 5–10% of hypertension cases, but only a minority of patients are currently diagnosed and treated because of a complex, stepwise, and partly invasive workup. We tested the performance of urine steroid metabolomics, the computational analysis of 24-hour urine steroid metabolome data by machine learning, for the identification and subtyping of PA. Mass spectrometry-based multi-steroid profiling was used to quantify the excretion of 34 steroid metabolites in 24-hour urine samples from 158 adults with PA (88 with unilateral PA [UPA] due to aldosterone-producing adenomas [APAs]; 70 with bilateral PA [BPA]) and 65 sex- and age-matched healthy controls. All APAs were resected and underwent targeted gene sequencing to detect somatic mutations associated with UPA. Patients with PA had increased urinary metabolite excretion of mineralocorticoids, glucocorticoids, and glucocorticoid precursors. Urine steroid metabolomics identified patients with PA with high accuracy, both when applied to all 34 or only the three most discriminative steroid metabolites (average areas under the receiver-operating characteristics curve [AUCs-ROC] 0.95–0.97). Whilst machine learning was suboptimal in differentiating UPA from BPA (average AUCs-ROC 0.65–0.73), it readily identified APA cases harbouring somatic KCNJ5 mutations (average AUCs-ROC 0.79–85). These patients showed a distinctly increased urine excretion of the hybrid steroid 18-hydroxycortisol and its metabolite 18-oxo-tetrahydrocortisol, the latter identified by machine learning as by far the most discriminative steroid. In conclusion, urine steroid metabolomics is a non-invasive candidate test for the accurate identification of PA cases and KCNJ5-mutated APAs.

Niacin - a scoping review for Nordic Nutrition Recommendations 2023.

Niacin is the precursor to pyridine nucleotides NAD (nicotinamide adenine dinucleotide) and NADP (nicotinamide adenine dinucleotide phosphate). Niacin (vitamin B3) is the common term for nicotinic acid, nicotinamide and derivatives that exhibit the biological activity of nicotinamide. Furthermore, the indispensable amino acid tryptophan is the substrate for de novo synthesis of NAD. Thus, the requirements and intake of niacin are expressed as niacin equivalents (NE). The focus of interest for niacin over the last decade has primarily been on pharmacological doses of nicotinic acid as a lipid-lowering agent and other NAD precursors as potential enhancers of cellular NAD+ concentrations. None of these studies, however, makes a useful contribution to understanding dietary requirements in healthy populations. The requirement for niacin is estimated based on the relationship between intake and biochemical indices of niacin status, primarily urinary excretion of nicotinamide metabolites.

A-041 Measurement of Plasma 3-Hydroxyisovalerylcarnitine by LC-MS/MS to Predict Marginal Biotin Deficiency in Inflammatory Bowel Disease

Abstract Background Biotin’s role in the immune response have emergently revealed that biotin is related to inflammation and that its deficiency leads to an increase in the level of proinflammatory cytokines. A biotin-deficient diet was recently shown to induce a colitis-like phenotype in mice, alleviated by biotin substitution. Mice with dextran sulphate sodium (DSS)-induced colitis showed biotin deficiency and significantly reduced biotin absorption. Oral biotin substitution reversed DSS colitis and induced remission. Diagnosis of biotin deficiency is rather challenging since based solely on measurement of circulating biotin levels has been shown to be insufficiently sensitive for clinical purposes. Increased urinary excretion of biotin metabolites, is suggested as a sensitive indicator of biotin deficiency. However, 24 h urine collection is time consuming, expensive, and the test relies on correct collection of samples by the patient, thus if this is not done properly the results may be inaccurate. 3HIVc increases in the case of biotin deficiency as a response to the decreased activity of biotin dependent enzyme methyl crotonyl-coenzyme. Analysis of circulating 3HIVc levels by LC-MS/MS has been found to be one of the most sensitive markers of biotin depletion. Methods Study cohort conducted with 100 inflammatory bowel disease (IBD) patients (20–60 years, 40 females) whom diagnosed according to standard clinical, radiological and pathological criteria and 100 healthy controls (20–60 years, 40 females). Of the patients 58/100 had Crohn’s Disease (CD), 42/100 had ulcerative colitis (UC) and 29/100 were in inflammatory state (serum high sensitivity C reactive protein (hsCRP) concentrations &amp;gt;5 mg/L). Standard serum and citrate-plasma samples were withdrawn between 8:00–10:00 h in the clinic after overnight fasting and stored at −20 °C until the analysis. Complete blood count (CBC), high sensitivity C reactive protein (hsCRP), fecal calprotectin (fCal) were determined with standard methods in serum matrix. 3-Hydroxyisovalerylcarntine (3HIVc) levels were determined by a commerical LC-MS/MS Kit in citrate plasma (KM3200, Immundiagnostik AG, Germany). Results Patients with IBD were found to have significantly higher 3HIVc levels than controls (6.1 ± 2.4 ng/mL vs 5.1 ± 2.1 ng/mL respectively, P = 0.004). 3HIVc concentrations were compared according to different disease characteristics within the patient group: circulating 3HIVc levels were found to be similar in patients with CD and UC (5.2 ± 2.1 ng/mL vs 4.9 ± 2.1, P = 0.451). Patients were also found to have similar circulating 3HIVc concentrations irrespective of whether or not inflammatory activity was present (5.1 ± 2.0 ng/mL in inflammatory vs 5.7 ± 2.4 ng/mL in noninflammatory conditions, P = 0.131). Conclusion The clinical findings of this study are in line with the existing preclinical evidence indicating that biotin deficiency is more common in patients with IBD in comparison to healthy controls in a clinical setting. However, surprisingly, the biotin status of patients with IBD was not found to differ according to inflammatory status. Therefore, there is still a room for more comprehensive look into the relationship between IBD related inflammation and biotin deficiency. Moreover, LC-MS/MS analysis of plasma 3HIVc has the potential to help clinicians to define biotin repletion and help to clearify the latter relation.

First Report of a Low-Frequency Mosaic Mutation in the Hydroxymethylbilane Synthase Gene Causing Acute Intermittent Porphyria.

Acute porphyrias are a group of monogenetic inborn errors of heme biosynthesis, characterized by acute and potentially life-threatening neurovisceral attacks upon exposure to certain triggering factors. Biochemical analyses can determine the type of acute porphyria, and subsequent genetic analysis allows for the identification of pathogenic variants in the specific gene, which provides information for family counselling. In 2017, a male Swiss patient was diagnosed with an acute porphyria while suffering from an acute attack. The pattern of porphyrin metabolite excretion in urine, faeces, and plasma was typical for an acute intermittent porphyria (AIP), which is caused by inherited autosomal dominant mutations in the gene for hydroxymethylbilane synthase (HMBS), the third enzyme in the heme biosynthetic pathway. However, the measurement of HMBS enzymatic activity in the erythrocytes was within the normal range and Sanger sequencing of the HMBS gene failed to detect any pathogenic variants. To explore the molecular basis of the apparent AIP in this patient, we performed third-generation long-read single-molecule sequencing (nanopore sequencing) on a PCR product spanning the entire HMBS gene, including the intronic sequences. We identified a known pathogenic variant, c.77G>A, p.(Arg26His), in exon 3 at an allelic frequency of ~22% in the patient's blood. The absence of the pathogenic variant in the DNA of the parents and the results of additional confirmatory studies supported the presence of a de novo mosaic mutation. To our knowledge, such a mutation has not been previously described in any acute porphyria. Therefore, de novo mosaic mutations should be considered as potential causes of acute porphyrias when no pathogenic genetic variant can be identified through routine molecular diagnostics.

Prognostic markers of recurrence in adrenocortical carcinoma patients after surgery

BACKGROUND: Adrenocortical carcinoma is a rare and aggressive disease. Tumor recurrence prevention is vital for increasing patients survival rate. Therefore, the identification of prognostic markers is of particular importance.&#x0D; AIM: To evaluate clinical, laboratory and chromatographic criteria for adrenocortical carcinoma recurrence using gas chromatography-mass spectrometry to optimize patients follow-up.&#x0D; MATERIALS AND METHODS: 40 patients [10 (25%) men and 30 (75%) women] with histologically confirmed adrenocortical carcinoma (according to the European Network for the Study of Adrenal Tumors stage I 3, II 29, III 8 patients at presentation) have been recruited. Measurement of 24-hour urinary steroid metabolite excretion has been carried out by gas chromatography-mass spectrometry (GCMS-ТQ8050, Shimadzu) in preoperative period. The survival distribution has been assessed according to the KaplanMeier method. Cox proportional hazards regression methods have been used to determine predictive factors on recurrence-free survival.&#x0D; RESULTS: The patients with early stages of disease (III versus III) had a significantly longer recurrence-free survival, overt hypercortisolism was associated with significantly shorter recurrence-free survival. A significant correlation between pregnenediol urinary excretion and tumor diameter, stage has been found. Increased urinary excretion of tetrahydro-11-deoxycortisol, pregnenediol was associated with a decreased recurrence-free survival in the patients with overt hypercortisolism. Increased urinary excretion of 16-OH-dehydroepiandrosterone was a marker of a shorter recurrence-free survival in the patients without Cushing syndrome. Increased urinary excretion of pregnenediol, pregnenetriol, 16-oxo-androstenediol in all the adrenocortical carcinoma patients was associated with a recurrence risk. In the multivariate analysis pregnenediol urinary excretion, stage and overt cortisol excess were significantly and independently associated with a shorter recurrence-free survival.&#x0D; CONCLUSIONS: Adrenocortical carcinoma recurrence prognostic markers have been found by gas chromatography-mass spectrometry. The application of these findings may improve patient-centered outcomes.

Tissue Glucocorticoid Metabolism in Adrenal Insufficiency: A Prospective Study of Dual-release Hydrocortisone Therapy.

Patients with adrenal insufficiency (AI) require life-long glucocorticoid (GC) replacement therapy. Within tissues, cortisol (F) availability is under the control of the isozymes of 11β-hydroxysteroid dehydrogenase (11β-HSD). We hypothesize that corticosteroid metabolism is altered in patients with AI because of the nonphysiological pattern of current immediate release hydrocortisone (IR-HC) replacement therapy. The use of a once-daily dual-release hydrocortisone (DR-HC) preparation, (Plenadren®), offers a more physiological cortisol profile and may alter corticosteroid metabolism in vivo. Prospective crossover study assessing the impact of 12 weeks of DR-HC on systemic GC metabolism (urinary steroid metabolome profiling), cortisol activation in the liver (cortisone acetate challenge test), and subcutaneous adipose tissue (microdialysis, biopsy for gene expression analysis) in 51 patients with AI (primary and secondary) in comparison to IR-HC treatment and age- and BMI-matched controls. Patients with AI receiving IR-HC had a higher median 24-hour urinary excretion of cortisol compared with healthy controls (72.1 µg/24 hours [IQR 43.6-124.2] vs 51.9 µg/24 hours [35.5-72.3], P = .02), with lower global activity of 11β-HSD2 and higher 5-alpha reductase activity. Following the switch from IR-HC to DR-HC therapy, there was a significant reduction in urinary cortisol and total GC metabolite excretion, which was most significant in the evening. There was an increase in 11β-HSD2 activity. Hepatic 11β-HSD1 activity was not significantly altered after switching to DR-HC, but there was a significant reduction in the expression and activity of 11β-HSD1 in subcutaneous adipose tissue. Using comprehensive in vivo techniques, we have demonstrated abnormalities in corticosteroid metabolism in patients with primary and secondary AI receiving IR-HC. This dysregulation of pre-receptor glucocorticoid metabolism results in enhanced glucocorticoid activation in adipose tissue, which was ameliorated by treatment with DR-HC.

Development, testing, parameterisation, and calibration of a human PBPK model for the plasticiser, di-(2-ethylhexyl) terephthalate (DEHTP) using in silico, in vitro and human biomonitoring data.

A physiologically based pharmacokinetic model for di-(2-ethylhexyl) terephthalate (DEHTP) based on a refined model for di-(2-propylheptyl) phthalate (DPHP) was developed to interpret the metabolism and biokinetics of DEHTP following a single oral dose of 50mg to three male volunteers. In vitro and in silico methods were used to generate parameters for the model. For example, measured intrinsic hepatic clearance scaled from in vitro to in vivo and plasma unbound fraction and tissue:blood partition coefficients (PCs) were predicted algorithmically. Whereas the development and calibration of the DPHP model was based upon two data streams, blood concentrations of parent chemical and first metabolite and the urinary excretion of metabolites, the model for DEHTP was calibrated against a single data stream, the urinary excretion of metabolites. Despite the model form and structure being identical significant quantitative differences in lymphatic uptake between the models were observed. In contrast to DPHP the fraction of ingested DEHTP entering lymphatic circulation was much greater and of a similar magnitude to that entering the liver with evidence for the dual uptake mechanisms discernible in the urinary excretion data. Further, the absolute amounts absorbed by the study participants, were much higher for DEHTP relative to DPHP. The in silico algorithm for predicting protein binding performed poorly with an error of more than two orders of magnitude. The extent of plasma protein binding has important implications for the persistence of parent chemical in venous blood-inferences on the behaviour of this class of highly lipophilic chemicals, based on calculations of chemical properties, should be made with extreme caution. Attempting read across for this class of highly lipophilic chemicals should be undertaken with caution since basic adjustments to PCs and metabolism parameters would be insufficient, even when the structure of the model itself is appropriate. Therefore, validation of a model parameterized entirely with in vitro and in silico derived parameters would need to be calibrated against several human biomonitoring data streams to constitute a data rich source chemical to afford confidence for future evaluations of other similar chemicals using the read-across approach.

Wound Healing Process as the Most Appropriate Modality of Cancer Therapy

President Biden of USA declared cancer moonshot initiative on Sept. 12, 2022 to urge the health profession to come up solutions to save 50% of cancer patients in the following 25 years. Cancer therapies were dominated by strategy based on killing of cancer cells in the past, which have been drilled through as a presidential project, but failed to achieve the goal to win the war on cancer. Obviously, a drastic change of strategy is necessary to fulfill moonshot of President Biden. Wound healing requires the proliferation and the terminal differentiation of progenitor stem cells (PSCs). Wound healing comes naturally, because the nature creates chemo-surveillance to ensure perfection of wound healing. Healthy people are able to maintain a steady level of wound heading metabolites active as differentiation inducers (DIs) and differentiation helper inducers (DHIs) to promote terminal differentiation of PSCs through destabilization of abnormal methylation enzymes (MEs). The functionality of chemo-surveillance can be damaged under pathological conditions triggering the production of tumor necrotic factor (TNF) to cause cachexia symptoms. A manifestation of cachexia symptoms is the excessive urinary excretion of wound healing metabolites to affect terminal differentiation of PSCs to heal wound, allowing PSCs to evolve into cancer stem cells (CSCs), and then to progress to faster growing cancer cells (CCs) through activation of oncogenes and/or inactivation of suppressor genes. Since cancer is caused by wound unhealing. Thus, wound healing process offers the most appropriate modality of cancer therapy. CDA-2 is a preparation of wound healing metabolites purified from urine, which has superb therapeutic efficacy against myelodysplastic syndrome (MDS). MDS is a disease caused by the build up of CSCs unable to undergo terminal differentiation. Apparently, wound healing metabolites are the best medicine to eradicate CSCs. For the therapy of cancer, we propose to add another set of DI + DHI consisting of synthetic products to target CCs, since CCs are known to express high levels of salvage enzymes to support their faster growth. CDA formulations definitely can fulfill cancer moonshot. Perpetual replication of CCs is the most outstanding feature of cancer. Naturally, killing of CCs becomes the choice of cancer establishments to combat cancer in the past. This modality of cancer therapy has been drilled through as a presidential project, but failed to achieve the goal to win the war on cancer. The contribution to damage chemo-surveillance and the inability to eradicate CSCs are responsible for the failure to save cancer patients. A combination therapy with CDA formulations can eliminate such deficit to fulfill cancer moonshot.

Refinement and calibration of a human PBPK model for the plasticiser, Di-(2-propylheptyl) phthalate (DPHP) using in silico, in vitro and human biomonitoring data.

An existing physiologically based pharmacokinetic model for Di-(2-propylheptyl) phthalate (DPHP) was refined to improve the simulations of the venous blood concentrations of the primary monoester metabolite, mono-(2-propylheptyl) phthalate (MPHP). This was considered a significant deficiency that should be addressed because the primary metabolite of other high molecular weight phthalates has been associated with toxicity. The various processes that influence the concentration of DPHP and MPHP in blood were re-evaluated and modified. A few simplifications of the existing model were made, including the removal of enterohepatic recirculation (EHR) of MPHP. However, the primary development was describing the partial binding of MPHP to plasma proteins following uptake of DPHP and metabolism in the gut affording better simulation of the trends observed in the biological monitoring data. Secondly, the relationship between blood concentrations and the urinary excretion of secondary metabolites was explored further because the availability of two data streams provides a better understanding of the kinetics than reliance on just one. Most human studies are conducted with few volunteers and generally with the absence of blood metabolite measurements which would likely imply an incomplete understanding of the kinetics. This has important implications for the "read across" approach proposed as part of the development of New Approach Methods for the replacement of animals in chemical safety assessments. This is where the endpoint of a target chemical is predicted by using data for the same endpoint from another more "data rich" source chemical. Validation of a model parameterized entirely with in vitro and in silico derived parameters and calibrated against several data streams would constitute a data rich source chemical and afford more confidence for future evaluations of other similar chemicals using the read-across approach.

Pediatric Urinary Mast Cell Mediator Levels

Increased urinary excretion of metabolites of the mast cell mediators prostaglandin D2, (2,3-dinor-11β prostaglandin F2α) (2,3-BPG), leukotriene (LT) C4 (LTE4) and histamine (N-methyl histamine (NMH)) support the diagnosis of various allergic disorders in the adult population. In the pediatric population, the reference range and clinical utility of mast cell mediators have not been well defined.

Dietary restriction and medical therapy drives PPARα-regulated improvements in early diabetic kidney disease in male rats.

The attenuation of diabetic kidney disease (DKD) by metabolic surgery is enhanced by pharmacotherapy promoting renal fatty acid oxidation (FAO). Using the Zucker Diabetic Fatty and Zucker Diabetic Sprague Dawley rat models of DKD, we conducted studies to determine if these effects could be replicated with a non-invasive bariatric mimetic intervention. Metabolic control and renal injury were compared in rats undergoing a dietary restriction plus medical therapy protocol (DMT; fenofibrate, liraglutide, metformin, ramipril, and rosuvastatin) and ad libitum-fed controls. The global renal cortical transcriptome and urinary 1H-NMR metabolomic profiles were also compared. Kidney cell type-specific and medication-specific transcriptomic responses were explored through in silico deconvolution. Transcriptomic and metabolomic correlates of improvements in kidney structure were defined using a molecular morphometric approach. The DMT protocol led to ∼20% weight loss, normalized metabolic parameters and was associated with reductions in indices of glomerular and proximal tubular injury. The transcriptomic response to DMT was dominated by changes in fenofibrate- and peroxisome proliferator-activated receptor-α (PPARα)-governed peroxisomal and mitochondrial FAO transcripts localizing to the proximal tubule. DMT induced urinary excretion of PPARα-regulated metabolites involved in nicotinamide metabolism and reversed DKD-associated changes in the urinary excretion of tricarboxylic acid (TCA) cycle intermediates. FAO transcripts and urinary nicotinamide and TCA cycle metabolites were moderately to strongly correlated with improvements in glomerular and proximal tubular injury. Weight loss plus pharmacological PPARα agonism is a promising means of attenuating DKD.

RF21 | PSAT99 Urinary adrenal steroidome modifications by Ketoconazole and Metyrapone alter the urinary free cortisol immunoassay reliability in Cushing Syndrome

Abstract Introduction Twenty-four-hour urinary free cortisol (24h-UFC) is the most used test for follow-up decision-making in patients with Cushing syndrome (CS) under adrenal-directed medical treatment. In CS patients without medical treatment, 24h-UFC determinations by immunoassays (IA) are commonly overestimated because of interference by cross-reacting steroid metabolites. However, it is unknown how induced changes by Ketoconazole (KTZ) or Metyrapone (MTP) on the excretion of urinary steroid metabolites alter the IA cross-reactivity interference of 24h-UFC determinations and what are its clinical implications. Methods 24h-UFC was analyzed by IA and gas chromatography-mass spectrometry (GC-MS) in 193 samples (81 before treatment, 73 during KTZ and 39 during MTP) from 34 CS patients. In addition, 35 urinary steroids were analyzed by GC-MS on each patient before and during treatment. Results Before treatment, 24h-UFC*IA determinations were overestimated by a factor of 1.75: 1 (95% CI 1.60 - 1.94) compared to those by GC-MS. During treatment with KTZ, the cross-reactivity interference of the 24h-UFC results by IA decreased in a dose-dependent manner (p = 0.000). Thus, in patients taking KTZ &amp;gt;400mg/day, 24h-UFC*IA determinations were no longer overestimated, giving similar results to those by GC-MS (factor of 0.98: 1, 95% CI, 0.83–1.20). Baseline 24h-UFC and upper limit normal (ULN) reductions were magnified (&amp;gt;25%) when assessed by IA vs GC-MS leading to higher falsely KTZ efficacy and misleading biochemical classification of 15% of patients. Despite dosage, in patients taking MTP the IA cross-reactivity only decreased 0.55 which resulted in persistence of overestimated 24h-UFC results (factor of 1.33: 1, 95% CI, 1.09 -1.76). When comparing MTP efficacy among IA vs GC-MS results, ULN baseline reductions were &amp;gt;50% different in some patients, while in others, similar reductions were found. Acceptable 24h-UFC method agreement between GC-MS and IA before treatment (R2 = 0.954) declined in patients under KTZ (R2 = 0.632) but not in MTP (R2 = 0.917). Urinary excretion changes of 22 urinary steroid metabolites explained 86% of the 24h-UFC*IA interference. 6β-Hydroxy-cortisol (6β-OH-cortisol) was the metabolite determining most of the 24h-UFC cross-reactivity variability (R2= 48.3%, p = 0.000) followed by 20α-dihydrocortisol (20α-DHF) (R2= 24.4%, p = 0.000). Each treatment induced a unique urinary steroid signature (p &amp;lt;0.001) and larger excretion reductions of 6β-OH-cortisol, 20α-DHF and 18-Hydroxy-cortisol in patients with KTZ elucidated the higher decrement of interference in the 24h-UFC by IA, compared to MTP treated patients (p = 0.001). Conclusion KTZ and MTP alter distinctly the urinary excretion of IA cross-reactive steroid metabolites, thus decreasing the cross-reactive interference of 24h-UFC*IA determinations present before treatment in a unique manner. Consequently, this interference reduction in 24h-UFC*IA leads to loss of method agreement with GC-MS and high risk of overestimating the biochemical impact of KTZ and MTP in controlling CS because of poor reliability of ULN. Presentation: Saturday, June 11, 2022 1:00 p.m. - 3:00 p.m., Sunday, June 12, 2022 12:36 p.m. - 12:41 p.m.

Metabolic impact of infant formulas in young infants. An outlook from the urine metabolome

IntroductionAlthough breast milk is the ideal food source for newborns during the first six months of life, a high percentage of children receive infant formulas. There is evidence that specific diet habits may influence individual metabolic profile. Therefore, in newborns, such profile can be influenced by the use of infantile formulas given the composition differences that display compared to human milk. Up to now, there are no reports in the literature that address this issue. Objectivesthis work aims to compare the metabolic profile of full-term newborns that were feed with either breast milk (n = 32) or infantile formulas (n = 21). Methods: Metabolic profile was established based on urine analysis through gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance (H-NMR). Resultsour results evidenced a more gluconeogenic profile in breast-fed infants characterized by elevation of Kreb's cycle intermediaries like fumaric, succinic and ketoglutaric acids compared to infants receiving infant formula. In addition, infant formula fed infants presented urinary excretion of metabolites derived from specific compounds present in this type of diet that were not observed in breast-fed infants, for instance D-glucitol, and 4-deoxytetronic. Moreover, in infant formula fed infants there was excretion of basal levels of metabolites of clinical relevance like 3-hydroxy-3-methyl-glutaric, 2-methyl-3-keto-valeric and 3,4-dihydroxybutyric. ConclusionThese results show the importance of understanding the metabolic impact of diet in newborn population in normal and pathological contexts.

Gas chromatography-mass spectrometry based steroid metabolomics in women with different phenotypes of polycystic ovarian syndrome and normal body weight

Objective: to study the steroid metabolomics in women with normal body weight and various PCOS phenotypes by gas chromatography-mass spectrometry (GC-MS). Materials and methods: forty-eight(48)women with PCOS aged 25±0,3 yearswith a BMI less than 25 kg/m2 were examined. The control group (CG) consisted of twenty-five (25) healthy women aged 26±0,6 years with a BMI of 23 (21-24) kg/m2. Immunoassays were used to determine the levels of hormones in serum. Urinary steroid profiles (USP) were studied by GC-MS method. Statistical data processing was performed using the software system STATISTICA for WINDOWS (ver. 10). Results: the article provides an analysis of the metabolism of androgens, glucocorticoids and progestogens in women with different phenotypes of polycystic ovary syndromeaccording to gas chromatography-mass spectrometry. Summary: the urinary excretion of androstenedione metabolites was increased in PCOS patients with androgen excess and anovulation (A and B phenotypes), dehydroepiandrosterone metabolites - in PCOS patients with androgen excess (A, B and C phenotypes). PCOS women with phenotype C showed raised urinary excretion of 11-oxo-pregnanetriol, pregnanetriol and 17-hydroxypregnanolone, a decrease in the ratios of the sum of tetrahydro derivatives of cortisol and cortisone to these progestogens, as well as determination of tetrahydro-21-deoxycorticol and nonclassical 5-ene-pregnenes according to GC-MS data. In fact, it indicated to deficiency of the 21-hydroxylase enzyme in these patients. It was found PCOS patients with androgen excess (A, B and C phenotypes) had the signs of insufficient 3β-hydroxysteroid dehydrogenase activity. PCOS women with phenotype A were revealed deficiency of 11β-hydroxysteroid dehydrogenase (type 1).

Hesperidin Bioavailability Is Increased by the Presence of 2S-Diastereoisomer and Micronization-A Randomized, Crossover and Double-Blind Clinical Trial.

Hesperidin is a flavanone abundantly found in citrus fruits for which health beneficial effects have been reported. However, hesperidin shows a low bioavailability among individuals. The aim of this study was to evaluate the effects of the micronization process and 2R- and 2S-hesperidin diastereoisomers ratio on hesperidin bioavailability. In a first phase, thirty healthy individuals consumed 500 mL of orange juice with 345 mg of hesperidin, and the levels of hesperidin metabolites excreted in urine were determined. In the second phase, fifteen individuals with intermediate hesperidin metabolite levels excreted in urine were randomized in a crossover, postprandial and double-blind intervention study. Participants consumed 500 mg of the hesperidin-supplemented Hesperidin epimeric mixture (HEM), the micronized Hesperidin epimeric mixture (MHEM) and micronized 2S-Hesperidin (M2SH) in each study visit with 1 week of washout. Hesperidin metabolites and catabolites were determined in blood and urine obtained at different timepoints over a 24 h period. The bioavailability—relative urinary hesperidin excretion (% of hesperidin ingested)—of M2SH (70 ± 14%) formed mainly by 2S-diastereoisomer was significantly higher than the bioavailability of the MHEM (55 ± 15%) and HEM (43 ± 8.0%), which consisted of a mixture of both hesperidin diastereoisomers. Relative urinary excretion of hesperidin metabolites for MHEM (9.2 ± 1.6%) was significantly higher compared to the HEM (5.2 ± 0.81%) and M2SH (3.6 ± 1.0%). In conclusion, the bioavailability of 2S-hesperidin extract was higher compared to the standard mixture of 2S-/2R-hesperidin extract due to a greater formation of hesperidin catabolites. Furthermore, the micronization process increased hesperidin bioavailability.