bims-supasi Biomed News
on Sulfation pathways and signalling
Issue of 2022‒04‒24
eighteen papers selected by
Jonathan Wolf Mueller
University of Birmingham

  1. Antioxidants (Basel). 2022 Mar 30. pii: 678. [Epub ahead of print]11(4):
      Mucopolysaccharidosis, type IIIB (MPS IIIB) is a rare disease caused by mutations in the N-alpha-acetylglucosaminidase (NAGLU) gene resulting in decreased or absent enzyme activity. On the cellular level, the disorder is characterized by the massive lysosomal storage of heparan sulfate (HS)-one species of glycosaminoglycans. HS is a sulfur-rich macromolecule, and its accumulation should affect the turnover of total sulfur in cells; according to the studies presented here, it, indeed, does. The lysosomal degradation of HS in cells produces monosaccharides and inorganic sulfate (SO42-). Sulfate is a product of L-cysteine metabolism, and any disruption of its levels affects the entire L-cysteine catabolism pathway, which was first reported in 2019. It is known that L-cysteine level is elevated in cells with the Naglu-/- gene mutation and in selected tissues of individuals with MPS IIIB. The level of glutathione and the Naglu-/- cells' antioxidant potential are significantly reduced, as well as the activity of 3-mercaptopyruvate sulfurtransferase (MPST, EC and the level of sulfane sulfur-containing compounds. The direct reason is not yet known. This paper attempts to identify some of cause-and-effect correlations that may lead to this condition and identifies research directions that should be explored.
    Keywords:  3-mercaptopyruvate sulfurtransferase; Sanfilippo B syndrome; cysteine; glycosaminoglycans; heparin; sulfane sulfur; sulfate; sulfurtransferases
  2. ChemMedChem. 2022 Apr 22.
      Glycosaminoglycans (GAGs) are highly negatively charged macromolecules with a large cation binding capacity, but their interaction potential with exogeneous Gd 3+ ions is under-investigated. These might be released from chelates used as Gadolinium-based contrast agents (GBCAs) for clinical MR imaging due to transmetallation with endogenous cations like Zn 2+ . Recent studies have quantified how an endogenous GAG sequesters released Gd 3+ ions and impacts the thermodynamic and kinetic stability of some GBCAs. In this study, we investigate and compare the chelation ability of two important GAGs (heparin and chondroitin sulfate), as well as the homopolysaccharides dextran and dextran sulfate that are used as models for alternative macromolecular chelators. Our combined approach of MRI-based relaxometry and isothermal titration calorimetry shows that the chelation process of Gd 3+ into GAGs is not just a long-range electrostatic interaction as proposed for the Manning model, but presumably a site-specific binding. Furthermore, our results highlight the crucial role of sulfate groups in this process and indicate that the potential of a specific GAG to engage in this mechanism increases with its degree of sulfation. The transchelation of Gd 3+ ions from GBCAs to sulfated GAGs should thus be considered as one possible explanation for the observed long-term deposition of Gd 3+ in vivo and related observations of long-term signal enhancements on T 1 -weighted MR images.
    Keywords:  GBCA; MRI contrast agent; chelation; gadolinium; glycosaminoglycans
  3. Front Med (Lausanne). 2022 ;9 841726
      Binding to antithrombin-III (ATIII) determines the anticoagulant activity of heparin. The complexes formed between heparin and ATIII result from a specific pentasaccharide sequence containing a 3-O-sulfated glucosamine in medium position. Building block analysis of heparins, following heparinase digestion, is a critical method in quality control that provides a simple structural characterization of a complex product. Hence, in these applications, study of the digestion of 3-O-sulfated moieties merits special attention. With heparinase II, specific inhibition of cleavage of the non-reducing bond of 3-O-sulfated units is observed. This specificity was erroneously generalized to other heparinases when it was observed that in exhaustive digests of heparins with the heparinase mixture, resistant 3-O-sulfated tetrasaccharides were also obtained from the specific ATIII-binding pentasaccharides. In fact, the detection of unsaturated 3-O-sulfated disaccharides in digests of heparin by heparinases I+II+III, resulting from the cleavage of the 3-O sulfated unit by heparinase I in non-conventional sequences, shows that this inhibition has exceptions. Thus, in experiments where heparinase II is selectively applied, these sequences can only be digested into tetra- or hexasaccharides where the 3-O-sulfated glucosamine is shifted on the reducing end. Heparinase I+II+III and heparinase II digests with additional tagging by reductive amination with sulfanilic acid were used to study the structural neighborhood of 3-O-sulfated disaccharides in bovine mucosal heparin fractions with increasing affinity for ATIII. The 3-O-sulfated disaccharides detected in heparinase I+II+III digests turn into numerous specific 3-O-sulfated tetrasaccharides in heparinase II digests. Additionally, ATIII-binding pentasaccharides with an extra 3-O-sulfate at the reducing glucosamine are detected in fractions of highest affinity as heparinase II-resistant hexasaccharides with two consecutive 3-O-sulfated units.
    Keywords:  3-O-sulfated disaccharides; bovine intestinal heparin; heparinase II; heparinase digestion; sulfanilic tagging
  4. Sci Rep. 2022 Apr 15. 12(1): 6294
      Spike-mediated entry of SARS-CoV-2 into human airway epithelial cells is an attractive therapeutic target for COVID-19. In addition to protein receptors, the SARS-CoV-2 spike (S) protein also interacts with heparan sulfate, a negatively charged glycosaminoglycan (GAG) attached to certain membrane proteins on the cell surface. This interaction facilitates the engagement of spike with a downstream receptor to promote viral entry. Here, we show that Mitoxantrone, an FDA-approved topoisomerase inhibitor, targets a heparan sulfate-spike complex to compromise the fusogenic function of spike in viral entry. As a single agent, Mitoxantrone inhibits the infection of an authentic SARS-CoV-2 strain in a cell-based model and in human lung EpiAirway 3D tissues. Gene expression profiling supports the plasma membrane as a major target of Mitoxantrone but also underscores an undesired activity targeting nucleosome dynamics. We propose that Mitoxantrone analogs bearing similar heparan sulfate-binding activities but with reduced affinity for DNA topoisomerases may offer an alternative therapy to overcome breakthrough infections in the post-vaccine era.
  5. Biology (Basel). 2022 Mar 25. pii: 506. [Epub ahead of print]11(4):
      Glycosaminoglycans (GAGs) are considered to be the most difficult type of glycoconjugates to analyze as they are constituted of linear long polysaccharidic chains having molecular weights reaching up to several million daltons. Bottom-up analysis of glycosaminoglycans from biological samples is a long and work-extensive procedure due to the many preparation steps involved. In addition, so far, only few research articles have been dedicated to the analysis of GAGs by means of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) because their intact ionization can be problematic due to the presence of labile sulfate groups. In this work, we had the aim of exploring the sulfation pattern of monosulfated chondroitin/dermatan sulfate (CS/DS) disaccharides in human tissue samples because they represent the most abundant form of sulfation in disaccharides. We present here an optimized strategy to analyze on-target derivatized CS/DS disaccharides via MALDI-TOF-MS using a fast workflow that does not require any purification after enzymatic cleavage. For the first time, we show that MALDI-TOF/TOF experiments allow for discrimination between monosulfated CS disaccharide isomers via specific fragments corresponding to glycosidic linkages and to cross-ring cleavages. This proof of concept is illustrated via the analysis of CS/DS disaccharides of atherosclerotic lesions of different histological origins, in which we were able to identify their monosulfation patterns.
    Keywords:  chondroitin sulfate; glycosaminoglycans; mass spectrometry
  6. Biochem Biophys Res Commun. 2022 Apr 10. pii: S0006-291X(22)00558-7. [Epub ahead of print]609 183-188
      Effective cancer immunotherapy requires physical contact of T cells with cancer cells. However, tumors often constitute special microenvironments that exclude T cells and resist immunotherapy. Cholesterol sulfate (CS) is a product of sulfotransferase SULT2B1b and acts as an endogenous inhibitor of DOCK2, a Rac activator essential for migration and activation of lymphocytes. We have recently shown that cancer-derived CS prevents tumor infiltration by effector T cells. Therefore, SULT2B1b may be a therapeutic target to dampen CS-mediated immune evasion. Here, we identified 3β-hydroxy-5-cholenoic acid (3β-OH-5-Chln) as a cell-active inhibitor of SULT2B1b. 3β-OH-5-Chln inhibited the cholesterol sulfotransferase activity of SULT2B1b in vitro and suppressed CS production from cancer cells expressing SULT2B1b. In vivo administration of 3β-OH-5-Chln locally reduced CS level in murine CS-producing tumors and increased infiltration of CD8+ T cells. When combined with immune checkpoint blockade or antigen-specific T cell transfer, 3β-OH-5-Chln suppressed the growth of CS-producing tumors. These results demonstrate that pharmacological inhibition of SULT2B1b can promote antitumor immunity through suppressing CS-mediated T cell exclusion.
    Keywords:  Cancer immunotherapy; Cholesterol sulfate; DOCK2; Immune evasion; SULT2B1b; T cell exclusion
  7. Curr Oncol. 2022 Apr 12. 29(4): 2681-2694
      Cancer-related cognitive impairment (CRCI) has been frequently reported in colorectal cancer survivors. Heparan sulfate (HS) was gradually considered to be related to cognitive disorders. The effect and potential mechanism of HS on CRCI in colorectal cancer patients were unexplored. In this study, all participants were divided into a cognitive impaired group and a cognitive normal group. The concentrations of oxidative stress factors and HS in serum were detected. Associations among HS, oxidative stress factors and CRCI were evaluated. Participants with cognitive impairment exhibited increased levels of HS, GSH, SOD and MDA, compared to the patients with normal cognitive performance. The independent significant association was found between HS and CRCI after controlling for various covariates. The higher concentrations of HS were related to the decreased cognitive performance among survivors who reported higher levels of GSH (β = 0.080, p = 0.002). Moreover, the nonlinear association between the level of HS and cognitive scores was confirmed using the restricted cubic splines (p < 0.001). These results indicated that the increased concentrations of circulating HS had a nonlinear negative connection with cognitive performance in colorectal cancer survivors, which was moderated by GSH. HS might be a new biomolecule for the identification and management of patients with CRCI.
    Keywords:  GSH; cancer-related cognitive impairment; colorectal cancer; heparan sulfate; oxidative stress
  8. Carbohydr Polym. 2022 Jul 15. pii: S0144-8617(22)00283-1. [Epub ahead of print]288 119379
      Lactose-modified chitosan (CTL) is sulfated using SO3·py or SO3·DMF as sulfating agents. The two products are characterized by elemental analysis, FT-IR, 1H,13C-DEPT-HSQC and 1H,13C-HSQC-TOCSY experiments which allow the extent and selectivity of chemical sulfation to be determined. Dynamic Light Scattering shows a pH-dependent association of the sulfated polysaccharides which are described as flexible by the Smidsrød's B parameter and the intrinsic viscosity at infinite ionic strength. Shear viscosity and intrinsic viscosity show that sulfation protocols lead to chain scission which is more pronounced when SO3·DMF is used. The sulfated samples are able to induce aggregation of human bone marrow mesenchymal stem cells, resulting in the formation of smaller nodules compared to the unmodified CTL sample. Over time, the sample with the higher degree of sulfation allows further aggregation between cell clusters while the sample with the lower degree of sulfation shows dissolution of the aggregates.
    Keywords:  Cell aggregation; Lactose-modified chitosan; Sulfation; hMSC-BM
  9. Biomolecules. 2022 Apr 05. pii: 545. [Epub ahead of print]12(4):
      In postmenopausal women, estrogen levels exclusively depend on local formation from the steroid precursors dehydroepiandrosterone sulfate and estrone sulfate (E1-S). Reduced estrogen levels are associated with menopausal symptoms. To mitigate these symptoms, more women nowadays choose medicine of natural origin, e.g., Cimicifuga racemosa (CR), instead of hormone replacement therapy, which is associated with an increased risk of breast cancer, stroke, and pulmonary embolism. Although CR treatment is considered safe, little is known about its effects on healthy endometrial and ovarian tissue and hormone-dependent malignancies, e.g., endometrial and ovarian cancers that arise during menopause. The aim of our study was to examine the effects of CR on the expression of genes encoding E1-S transporters and estrogen-related enzymes in control and cancerous endometrial and ovarian cell lines. CR affected the expression of genes encoding E1-S transporters and estrogen-related enzymes only at very high concentrations, whereas no changes were observed at physiological concentrations of CR. This suggests that CR does not exert estrogenic effects in endometrial and ovarian tissues and probably does not affect postmenopausal women's risks of endometrial or ovarian cancer or the outcomes of endometrial and ovarian cancer patients.
    Keywords:  Cimicifuga racemosa; endometrial cancer; ovarian cancer; steroid transporters
  10. Int J Biol Macromol. 2022 Apr 19. pii: S0141-8130(22)00792-9. [Epub ahead of print]
      Elevated expression of transforming growth factor β1 (TGF-β1) have been implicated in the pathogenesis of liver fibrosis, thus attenuating the excessive TGF-β1's activity by TGF-β1-binding peptide is an ideal strategy for the treatment of liver fibrosis. However, the application of small peptide as a pharmaceutical agent is obstacle due to difficult preparation and non-selectively deliver. The I-plus sequences of circumsporozoite protein (CSP-I) possesses high affinity for heparan sulfate proteoglycans, which are primarily located on liver tissues. TGF-β1-binding peptide P15 hold specific ability of binding to TGF-β1. In this study, we describe an approach to efficiently preparing liver-targeting peptide P15-CSP-I, which is conjugation of the sequences of P15 to the N-terminus of CSP-I, from the cleavage of biological macromolecule SUMO-tagged P15-CSP-I. In vitro and ex vivo binding assay showed that P15-CSP-I specifically targeted to the hepatocytes and liver tissues. Moreover, P15-CSP-I inhibited cell proliferation, migration and invasion, and decreased fibrosis-related proteins expression in TGF-β1-activated HSCs in vitro. Furthermore, P15-CSP-I ameliorated liver morphology and decreased the fibrosis responses in vivo. Taken together, P15-CSP-I may be a potential candidate for targeting therapy on liver fibrosis due to its high efficient preparation, specific liver-targeting potential and improved anti-liver fibrotic activity.
    Keywords:  Liver fibrosis; Transforming growth factor β1; Transforming growth factor β1-binding peptide
  11. Toxins (Basel). 2022 Apr 16. pii: 287. [Epub ahead of print]14(4):
      p-Cresyl sulfate (PCS) is a uremic toxin that causes cardiovascular injury and progression in patients with chronic kidney disease (CKD). Peripheral arterial stiffness (PAS) as measured using the brachial-ankle pulse wave velocity (baPWV) is considered a valuable predictor of cardiovascular event risk in the general population. The study investigated the correlation between serum PCS levels and PAS (baPWV > 18.0 m/s) in 160 patients with stage 3-5 CKD. Liquid chromatography-mass spectrometry was used to assay serum PCS levels. PAS was detected in 54 patients (33.8%), and it was linked to older age, a higher prevalence of hypertension, higher systolic and diastolic blood pressure, higher serum calcium-phosphorus product and PCS levels, and lower height and body weight. Multivariable logistic regression analysis for independent factors associated with PAS illustrated that, in addition to age and diastolic blood pressure, serum PCS levels exhibited an odds ratio (OR) of 1.098 (95% confidence interval = 1.029-1.171, p = 0.005). These findings demonstrated that serum PCS levels were associated with PAS among patients with stage 3-5 CKD.
    Keywords:  brachial-ankle pulse wave velocity; chronic kidney disease; p-cresyl sulfate; peripheral artery stiffness
  12. Int J Eat Disord. 2022 Apr 22.
      OBJECTIVE: This study was designed to determine the status of dehydroepiandrosterone (DHEA) in women with anorexia nervosa (AN) and to assess the efficacy of DHEA supplementation as a treatment for bone health in women with AN.METHOD: Studies were retrieved from the PubMed, Embase, Cochrane Library, MEDLINE, and Scopus databases from inception to February 14, 2022. Observational studies that compared serum DHEA levels between women with AN and healthy controls were included for meta-analysis, and randomized controlled trials (RCTs) that evaluated the effects of DHEA supplementation on bone mass were reviewed.
    RESULTS: Meta-analysis of 15 cross-sectional studies revealed that patients with AN had significantly elevated serum DHEA levels (mean difference (MD) = 311.63 ng/dl; 95% confidence interval (CI), 78.01-545.25) and reduced DHEAS levels (MD = -24.90 μg/dl; 95% CI, -41.72 to -8.07) compared with healthy controls. A systematic review of seven RCTs found that DHEA monotherapy does not improve bone mineral density (BMD) compared with placebo after adjusting for weight gain. While the combination of DHEA and conjugated oral contraceptives has led to increased bone strength and decreased bone loss, the beneficial effect appears to be limited to older adolescents and adults with closed physes. Potential detrimental effects on BMD were identified in younger adolescents with open physes in one study.
    DISCUSSION: Due to the lack of apparent benefit of DHEA in women with AN and its potential detrimental effect on BMD in young patients with AN, current evidence does not support the use of DHEA.
    PUBLIC SIGNIFICANCE: This study demonstrates that women with anorexia nervosa have abnormal levels of dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEAS), which have been suggested by previous studies to play a role in the development of low bone density in this condition. However, current evidence does not support the use of DHEA as a treatment to preserve bone health in patients with anorexia nervosa given the lack of clear benefit following its use and also because of a potential detrimental effect on bone mineral density in young patients with anorexia nervosa.
    Keywords:  anorexia nervosa; bone loss; clinical trails; dehydroepiandrosterone; hormone replacement
  13. Microbiome. 2022 Apr 19. 10(1): 64
      BACKGROUND: Recent evidence implicates microbial sulfidogenesis as a potential trigger of colorectal cancer (CRC), highlighting the need for comprehensive knowledge of sulfur metabolism within the human gut. Microbial sulfidogenesis produces genotoxic hydrogen sulfide (H2S) in the human colon using inorganic (sulfate) and organic (taurine/cysteine/methionine) substrates; however, the majority of studies have focused on sulfate reduction using dissimilatory sulfite reductases (Dsr).RESULTS: Here, we show that genes for microbial sulfur metabolism are more abundant and diverse than previously observed and are statistically associated with CRC. Using ~ 17,000 bacterial genomes from publicly available stool metagenomes, we studied the diversity of sulfur metabolic genes in 667 participants across different health statuses: healthy, adenoma, and carcinoma. Sulfidogenic genes were harbored by 142 bacterial genera and both organic and inorganic sulfidogenic genes were associated with carcinoma. Significantly, the anaerobic sulfite reductase (asr) genes were twice as abundant as dsr, demonstrating that Asr is likely a more important contributor to sulfate reduction in the human gut than Dsr. We identified twelve potential pathways for reductive taurine metabolism and discovered novel genera harboring these pathways. Finally, the prevalence of metabolic genes for organic sulfur indicates that these understudied substrates may be the most abundant source of microbially derived H2S.
    CONCLUSIONS: Our findings significantly expand knowledge of microbial sulfur metabolism in the human gut. We show that genes for microbial sulfur metabolism in the human gut are more prevalent than previously known, irrespective of health status (i.e., in both healthy and diseased states). Our results significantly increase the diversity of pathways and bacteria that are associated with microbial sulfur metabolism in the human gut. Overall, our results have implications for understanding the role of the human gut microbiome and its potential contributions to the pathogenesis of CRC. Video abstract.
    Keywords:  Colorectal cancer; Cysteine; Gut; Human microbiome; Hydrogen sulfide; Metagenomics; Sulfur metabolism; Taurine
  14. J Mass Spectrom. 2022 May;57(5): e4827
      24S-hydroxycholesterol (i.e., cerebrosterol, 24S-OH-Chol) is the main form of cholesterol elimination from the brain. Liquid chromatography-tandem mass spectrometry methods were developed for the quantification of the total and unesterified/unbound fractions of 24S-OH-Chol, its monosulfate, monoglucuronide, and diconjugate derivatives (24S-OH-Chol-3sulfate [3S], 24S-OH-Chol-24glucuronide [24G] and 24S-OH-Chol-3S, 24G, respectively) in human plasma. Linearity, precision, accuracy, and extraction recovery were validated within the typical physiological and pathological ranges of concentrations for each compound. The lower limit of quantifications was 2.00, 0.33, 0.26, and 0.74 ng/ml for 24S-OH-Chol, 24S-OH-Chol-24G, 24S-OH-Chol-3S, and 24-OH-Chol-3S, 24G, respectively. Extraction recovery values in total and unbound plasma fractions were also analyzed in murine and monkey plasma and varied from 73% in mouse to 113% in cynomolgus monkey. The methods could rapidly (less than 7 min) quantify individual compounds with high sensitivity, accuracy (bias ≤15%), and reproducibility (coefficient of variation [CV] ≤ 17%). Their clinical applications were validated by measuring levels of the 4 compounds in samples from 20 noncholestatic donors, 5 cholestatic patients suffering from primary biliary cirrhosis, and 10 patients suffering from biliary stenosis. Results highlight the abundance of 24S-OH-Chol in the total fraction and the abundance of 24S-OH-Chol-3S and 24G in the unbound ones. While the latter strongly accumulate in plasma fractions of cholestatic patients, levels of 24S-OH-Chol remained similar to those of healthy donors. Our results indicate that this approach is suitable for monitoring cerebrosterol and its conjugates in large-scale clinical studies.
    Keywords:  24S-hydroxycholesterol; glucuronide conjugate; liquid chromatography coupled to tandem mass spectrometry; plasma levels; sulfate conjugate
  15. Pril (Makedon Akad Nauk Umet Odd Med Nauki). 2022 Apr 22. 43(1): 29-39
      Background: Evidence for disturbances in HPA activation and abnormal HPA regulatory mechanisms in schizophrenia is accumulating. Aim: To compare serum levels of cortisol, DHEA-S and their ratio between patients with schizophrenia and healthy controls and among patients before and after treatment with different types of antipsychotics. Material and methods: In this clinical prospective study, 60 patients with schizophrenia and 40 healthy age and sex matched control subjects were included. All patients experienced an acute exacerbation of the illness (PANSS: P1 and P3 ≥ 4). Clinical evaluation of patients was performed using the Positive and Negative Symptom Scale. A questionnaire for socio-demographic and clinical data collection was used. Serum levels of cortisol, DHEA-S and their ratio were measured at baseline in all participants and after 3 and 6 weeks, respectively, of the antipsychotic treatment with different types of antipsychotics in patients with schizophrenia. Results: Patients with schizophrenia had significantly higher serum cortisol and DHEA-S levels in comparison to the control group. There was no significant difference in serum levels of cortisol, DHEA-S and their ratio between patients treated with different types of antipsychotics (typical/atypical). Serum levels of the analyzed hormones significantly reduce during the 6-week period of examination in both subgroups treated with different types of antipsychotics. Conclusion: Elevated serum cortisol and DHEA-S in schizophrenic patients might be associated with their role in the pathophysiology of the disorder. There is no significant difference in serum levels of cortisol, DHEA-S and their ratio among the patients treated with different types of antipsychotics.
    Keywords:  DHEA-S; cortisol; schizophrenia; typical/atypical antipsychotics
  16. Mar Drugs. 2022 Mar 24. pii: 223. [Epub ahead of print]20(4):
      An immunomodulatory polysaccharide (DAP4) was extracted, purified, and characterized from Durvillaea antarctica. The results of chemical and spectroscopic analyses demonstrated that the polysaccharide was a fucoidan, and was mainly composed of (1→3)-α-l-Fucp and (1→4)-α-l-Fucp residues with a small degree of branching at C-3 of (1→4)-α-l-Fucp residues. Sulfate groups were at C-4 of (1→3)-α-l-Fucp, C-2 of (1→4)-α-l-Fucp and minor C-6 of (1→4)-β-d-Galp. Small amounts of xylose and galactose exist in the forms of β-d-Xylp-(1→ and β-d-Gal-(1→. The immunomodulatory activity of DAP4 was measured on RAW 264.7 cells, the results proved that DAP4 exhibited excellent immunomodulatory activities, such as promoted the proliferation of spleen lymphocytes, increased NO production, as well as enhanced phagocytic of macrophages. Besides, DAP4 could also produce better enhancement on the vitality of NK cells. For the high immunomodulatory activity, DAP4 might be a potential source of immunomodulatory fucoidan with a novel structure.
    Keywords:  Durvillaea antarctica; fucoidan; immunomodulatory; structural characterization
  17. Int J Lab Hematol. 2022 Apr 19.
      INTRODUCTION: Global coagulation assays may be of added value to the anti-Xa assay for monitoring heparin therapy. Unlike most testing methods, the thrombin generation assay (TGA) has the ability to assess the overall function of the hemostatic system, which provides information on the anticoagulation status of patients. We compared the TGA, measured with ST Genesia® STG-DrugScreen® reagent, with the anti-Xa assay for monitoring heparin therapy in inflammatory and non-inflammatory patients. We also determined reference values for STG-DrugScreen® thrombin generation (TG) parameters.METHODS: Reference values were determined on 120 healthy donors. Furthermore, a spiking experiment with unfractionated heparin (UFH) and low molecular weight heparin (LMWH) was performed, and samples of patients receiving UFH or LMWH were analyzed with ST Genesia® and the anti-Xa assay.
    RESULTS: High discrepancy between TG parameters and anti-Xa activity was observed for low LMWH anti-Xa levels. TG parameters were affected in 36/46 (time to peak) to 42/46 (peak height) patients during UFH therapy with sub-target anti-Xa activity levels.
    CONCLUSION: TGA seems insufficiently sensitive for low concentrations of LMWH. There may be an added value of the TGA for monitoring UFH in so-called heparin-resistant patients. Therefore, the TGA has the potential to be introduced as an additional tool for monitoring heparin therapy.
    Keywords:  blood coagulation tests; heparin; reference values; thrombin
  18. Metabolites. 2022 Apr 12. pii: 345. [Epub ahead of print]12(4):
      Hop prenylated flavonoids have been investigated for their in vivo activities due to their broad spectrum of positive health effects. Previous studies on the metabolism of xanthohumol using untargeted methods have found that it is first degraded into 8-prenylnaringenin and 6-prenylnaringenin, by spontaneous cyclisation into isoxanthohumol, and subsequently demethylated by gut bacteria. Further combinations of metabolism by hydroxylation, sulfation, and glucuronidation result in an unknown number of isomers. Most investigations involving the analysis of prenylated flavonoids used surrogate or untargeted approaches in metabolite identification, which is prone to errors in absolute identification. Here, we present a synthetic approach to obtaining reference standards for the identification of human xanthohumol metabolites. The synthesised metabolites were subsequently analysed by qTOF LC-MS/MS, and some were matched to a human blood sample obtained after the consumption of 43 mg of micellarised xanthohumol. Additionally, isomers of the reference standards were identified due to their having the same mass fragmentation pattern and different retention times. Overall, the methods unequivocally identified the metabolites of xanthohumol that are present in the blood circulatory system. Lastly, in vitro bioactive testing should be applied using metabolites and not original compounds, as free compounds are scarcely found in human blood.
    Keywords:  beer; blood analysis; hops; metabolites; prenylated flavonoids; synthesis; xanthohumol