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Biomonitoring using dried blood spots: detection of ochratoxin A and its degradation product 2'R-ochratoxin A in blood from coffee drinkers.

ABSTRACT: In this study, human exposure to the mycotoxin ochratoxin A (OTA) and its thermal degradation product 2'R-ochratoxin A (2'R-OTA, previously named as 14R-Ochratoxin A [22]) through coffee consumption was assessed. LC-MS/MS and the dried blood spot (DBS) technique were used for the analysis of blood samples from coffee and noncoffee drinkers (n = 50), and food frequency questionnaires were used to document coffee consumption.For the detection of OTA and 2'R-OTA in blood, a new sensitive and efficient sample preparation method based on DBS was established and validated. Using this technique 2'R-OTA was for the first time detected in biological samples. Comparison between coffee drinkers and noncoffee drinkers showed for the first time that 2'R-OTA was only present in blood from the first group while OTA could be found in both groups in a mean concentration of 0.21 μg/L. 2'R-OTA mean concentration was 0.11 μg/L with a maximum concentration of 0.414 μg/L. Thus, in average 2'R-OTA was approx. half the concentration of OTA but in some cases even exceeded OTA levels. No correlation between the amounts of coffee consumption and OTA or 2'R-OTA levels was observed.The results of this study revealed for the first time a high exposure of coffee consumers to 2'R-OTA, a compound formed from OTA during coffee roasting. Since little information is available regarding toxicity and possible carcinogenicity of this compound, further OTA monitoring in blood including 2'R-OTA is advisable.

SUBMITTER: Cramer B

PROVIDER: S-EPMC4744763 | biostudies-other | 2015 Sep

REPOSITORIES: biostudies-other

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Multi-mycotoxin analysis using dried blood spots and dried serum spots.

Project description:In this study, a rapid multi-mycotoxin approach was developed for biomonitoring and quantification of 27 important mycotoxins and mycotoxin metabolites in human blood samples. HPLC-MS/MS detection was used for the analysis of dried serum spots (DSS) and dried blood spots (DBS). Detection of aflatoxins (AFB1, AFB2, AFG1, AFG2, AFM1), trichothecenes (deoxynivalenol, DON; DON-3-glucoronic acid, DON-3-GlcA; T-2; HT-2; and HT-2-4-GlcA), fumonisin B1 (FB1), ochratoxins (OTA and its thermal degradation product 2'R-OTA; OTα; 10-hydroxychratoxin A, 10-OH-OTA), citrinin (CIT and its urinary metabolite dihydrocitrinone, DH-CIT), zearalenone and zearalanone (ZEN, ZAN), altenuene (ALT), alternariols (AOH; alternariol monomethyl ether, AME), enniatins (EnA, EnA1, EnB, EnB1) and beauvericin (Bea) was validated for two matrices, serum (DSS), and whole blood (DBS). HPLC-MS/MS analysis showed signal suppression as well as signal enhancement due to matrix effects. However, for most analytes LOQs in the lower pg/mL range and excellent recovery rate were achieved using matrix-matched calibration. Besides validation of the method, the analyte stability in DBS and DSS was also investigated. Stability is a main issue for some analytes when the dried samples are stored under common conditions at room temperature. Nevertheless, the developed method was applied to DBS samples of a German cohort (n = 50). Besides positive findings of OTA and 2'R-OTA, all samples were positive for EnB. This methodical study establishes a validated multi-mycotoxin approach for the detection of 27 mycotoxins and metabolites in dried blood/serum spots based on a fast sample preparation followed by sensitive HPLC-MS/MS analysis. Graphical Abstract ᅟ.

| S-EPMC5395583 | biostudies-literature

Detection of SARS-CoV-2 IgG antibodies in dried blood spots.

Project description:The COVID-19 pandemic led to development of numerous serologic tests. To obviate the need for phlebotomy services, we validated dried blood spots (DBS) for anti-SARS-CoV-2 serologic testing. Immunoglobulins were extracted from 3 mm DBS punches and the extracts were analyzed using the Euroimmun anti-SARS-CoV-2 IgG ELISA. Various pre-analytical factors were studied. Results were favorable for DBS stored for at least 67 days at or below 37°C. Comparison between paired serum and DBS specimens tested by the Euroimmun ELISA showed 96.8% and 81.3% positive and negative agreement, respectively, indicating that confirmatory testing of positive Euroimmun results on DBS extracts is necessary to achieve clinical accuracy. Our findings suggest that any SARS-CoV-2 antibody assay that requires pre-dilution of serum is amenable to DBS as an alternate specimen type that is relatively low cost, self-collectable, stable, can be shipped by standard mail and could be used to establish the seroprevalence of large populations.

| S-EPMC8116316 | biostudies-literature

Qualitative and quantitative detection of SARS-CoV-2 antibodies from dried blood spots.

Project description:IntroductionDried blood spot (DBS) sampling is a minimally invasive method for specimen collection with potential multifaceted uses, particularly for serosurveillance of previous SARS-CoV-2 infection. In this study, we assessed DBS as a potential specimen type for assessing IgG and total (including IgG and IgM) antibodies to SARS-CoV-2 in vaccinated and naturally infected patients.MethodsSix candidate buffers were assessed for eluting blood from DBS cards. The study utilized one hundred and five paired plasma specimens and DBS specimens from prospectively collected SARS-CoV-2 vaccinated individuals, remnants from those with PCR confirmed SARS-CoV-2 infections, or remnants from those without history of infection or vaccination. All specimens were tested with the Siemens SARS-CoV-2 total assay (COV2T) or IgG assay (sCOVG).ResultsThe lowest backgrounds were observed with water and PBS, and water was used for elution. Relative to plasma samples, DBS samples had a positive percent agreement (PPA) of 94.4% (95% CI: 94.9-100%) for COV2T and 79.2 (68.4-87.0) for sCOVG using the manufacturer's cutoff. The NPA was 100 % (87.1-100.0 and 85.13-100) for both assays. Dilution studies revealed 100% (95% CI: 90.8-100%) qualitative agreement between specimen types on the COV2T assay and 98.0% (88.0-99.9%) with the sCOVG using study defined cutoffs.ConclusionDBS specimens demonstrated high PPA and NPA relative to plasma for SARS-CoV-2 serological testing. Our data support feasibility of DBS sampling for SARS-CoV-2 serological testing.

| S-EPMC8721924 | biostudies-literature

Biomonitoring of ochratoxin A, 2'R-ochratoxin A and citrinin in human blood serum from Switzerland.

Project description:Biomonitoring of mycotoxins and their metabolites in biological fluids is increasingly used to assess human exposure. In this study, biomarkers of ochratoxin A (OTA) and citrinin (CIT) exposure were determined in a large number of serum samples from healthy blood donors in Switzerland. In 2019, 700 samples from different regions were obtained. From 240 donors, a second sample (taken 2-9 months later) was available for analysis. Moreover, 355 blood donor samples from 2005 from all regions in Switzerland and 151 additional samples from the southern Swiss region of Ticino from 2005 could be analysed.OTA, 2'R-ochratoxin A (2'R-OTA), ochratoxin alpha (OTα), CIT and dihydrocitrinone (DH-CIT) were analysed using validated targeted methods including precipitation and online SPE clean-up.OTA and 2'R-OTA were frequently detected (OTA in 99%; 2'R-OTA in 51% of the tested samples). The mean concentration in all positive samples was 0.4 ng/mL for OTA and 0.2 ng/mL for 2'R-OTA. OTα was not detected in any sample above the limit of quantification (LOQ). In contrast to OTA, CIT and DH-CIT were only quantifiable in 2% and 0.1% of the samples, respectively. No significant trend was observed between the samples from 2005 and the more recent samples, but OTA concentrations were usually higher in serum samples from the southern Swiss region of Ticino and in males compared to females.Our extensive data fit well within the framework of previously published values for the healthy adult European population.

| S-EPMC9038883 | biostudies-literature

Analysis of lyso-globotriaosylsphingosine in dried blood spots.

Project description:Recently, lyso-globotriaosylsphingosine (lyso-Gb3) was found to be elevated in plasma of treatment naive male patients and some female patients with Fabry Disease (FD). This study tested whether lyso-Gb3 could be analyzed in dried blood spots (DBS) from filter cards and whether concentrations are elevated in newborn infants with FD. Lyso-Gb3 concentrations were analyzed in DBS following extraction using a novel HPLC-mass spectrometry (MS)/MS method. Lyso-Gb3 levels in DBS were above the lower limit of quantitation (0.28 ng/mL) in 5/17 newborn FD infants (16 males; range: 1.02-8.81 ng/mL), but in none of the newborn controls, in all 13 patients (4 males) with classic FD (range: 2.06-54.1 ng/mL), in 125/159 Taiwanese individuals with symptomatic or asymptomatic FD who carry the late onset ?-galactosidase A (GLA) mutation c.936+919G>A (IVS4+919G>A) (3.75±0.69 ng/mL; range: 0.418-3.97 ng/mL) and in 20/29 healthy controls (0.77±0.24 ng/mL; range: 0.507-1.4 ng/mL). The HPLC-MS/MS method for analysis of lyso-Gb3 is robust and yields reproducible results in DBS in patients with FD. However, concentrations of lyso-Gb3 were below the limit of quantitation in most newborn infants with FD rendering this approach not suitable for newborn screening. In addition, most females with the late onset mutation have undetectable lyso-Gb3 concentrations.

| S-EPMC3698306 | biostudies-literature

Quantitation of 5-Methyltetrahydrofolic Acid in Dried Blood Spots and Dried Plasma Spots by Stable Isotope Dilution Assays.

Project description:Because of minimal data available on folate analysis in dried matrix spots (DMSs), we combined the advantages of stable isotope dilution assays followed by LC-MS/MS analysis with DMS sampling to develop a reliable method for the quantitation of plasma 5-methyltetrahydrofolic acid in dried blood spots (DBSs) and dried plasma spots (DPSs) as well as for the quantitation of whole blood 5-methyltetrahydrofolic acid in DBSs. We focused on two diagnostically conclusive parameters exhibited by the plasma and whole blood 5-methyltetrahydrofolic acid levels that reflect both temporary and long-term folate status. The method is performed using the [2H4]-labeled isotopologue of the vitamin as the internal standard, and three steps are required for the extraction procedure. Elution of the punched out matrix spots was performed using stabilization buffer including Triton X-100 in a standardized ultrasonication treatment followed by enzymatic digestion (whole blood only) and solid-phase extraction with SAX cartridges. This method is sensitive enough to quantify 27 nmol/L whole blood 5-methyltetrahydrofolic acid in DBSs and 6.3 and 4.4 nmol/L plasma 5-methyltetrahydrofolic acid in DBSs and DPSs, respectively. The unprecedented accurate quantification of plasma 5-methyltetrahydrofolic acid in DBSs was achieved by thermal treatment prior to ultrasonication, inhibiting plasma conjugase activity. Mass screenings are more feasible and easier to facilitate for this method in terms of sample collection and storage compared with conventional clinical sampling for the assessment of folate status.

| S-EPMC4659665 | biostudies-literature

Sensitive and Stable Molecular Detection of Dengue, Chikungunya, and Zika Viruses from Dried Blood Spots.

Project description:Standard molecular detection of many pathogens, in particular RNA viruses, requires appropriate handling in the field for preserving the quality of the sample until processing. This could be challenging in remote tropical areas. Dengue virus (DENV), chikungunya virus (CHIKV), and Zika virus (ZIKV) are RNA viruses, prominent among the causes of fever in the tropics. We aimed to test the stability of arboviral RNA in contrived dried blood spots prepared on Whatman 903 Protein saver cards as a means of sample collection and storage. We were able to detect DENV, CHIKV, and ZIKV by real-time RT-PCR up to 180 days after card inoculation with stable Ct values across the study period. Our study supports dried blood spots (DBS) on protein saver cards as a platform for stable detection of arboviral RNA of sufficient quality to be used in diagnostic RT-PCR assays and next generation sequencing.

| S-EPMC9393429 | biostudies-literature

Validation and clinical application of a method to quantify nevirapine in dried blood spots and dried breast-milk spots.

Project description:OBJECTIVES:The validation and clinical application of an LC-MS/MS method for the quantification of nevirapine in dried blood spots (DBS) and dried breast-milk spots (DBMS) are presented. METHODS:DBS and DBMS were prepared from 50 and 30 ?L of nevirapine-spiked whole blood and human breast milk, respectively. Chromatographic separation was achieved on a reverse-phase C18 column with 0.1% formic acid in water/acetonitrile using a solvent gradient programme at a flow rate of 400 ?L/min, and detection was by a TSQ Quantum Access triple quadrupole mass spectrometer. The clinical application was evaluated in HIV-positive nursing mothers and their breastfed infants. RESULTS:The assay was validated over the concentration range 50-10,000 ng/mL. Accuracy ranged from 93.3% to 113.4% and precision ranged from 1.9% to 12.0%. The mean (percentage coefficient of variation) recovery of nevirapine from DBS and DBMS was ? 70.7% (? 8.2) and the matrix effect was ? 1.04 (? 6.1). Nevirapine was stable in DBS and DBMS for ? 15 months at room temperature and -80°C. Mean (SD) AUC0-12, Cmax and Cmin in maternal plasma versus breast milk were 57,808 ng · h/mL (24,315) versus 55,817 ng · h/mL (22,368), 6140 ng/mL (2605) versus 5231 ng/mL (2215) and 4334 ng/mL (1880) versus 4342 ng/mL (2245), respectively. The milk-to-plasma concentration ratio over the dosing interval was 0.94 (0.15). Infant plasma concentrations 2 and 8 h after maternal dosing were 580.6 ng/mL (464.7-1607) and 584.1 ng/mL (381.5-1570), respectively. CONCLUSIONS:These methods further extend opportunities for conducting clinical pharmacokinetic studies in nursing mother-infant pairs, especially in resource-limited settings.

| S-EPMC6538530 | biostudies-literature

Quantification of Zonisamide in Dried Blood Spots and Dried Plasma Spots by UPLC-MS/MS: Application to Clinical Practice.

Project description:In this research, a UHPLC-MS/MS method was developed and validated for the determination of zonisamide in dried plasma spots (DPS) and dried blood spots (DBS). Detection of zonisamide and internal standard, 1-(2,3-dichlorphenyl)piperazine, was carried out in ESI+ mode by monitoring two MRM transitions per analyte. Total run time, less than 2.5 min, was achieved using Acquity UPLC BEH Amide (2.1 × 100 mm, 1.7 µm particle size) column with mobile phase comprising acetonitrile-water (85:15%, v/v) with 0.075% formic acid. The flow rate was 0.225 mL/min, the column temperature was 30 °C and the injection volume was 3 µL. Desolvation temperature, desolvation gas flow rate, ion source temperature and cone gas flow rate were set by the IntelliStart software tool in combination with tuning. All of the Guthrie cards were scanned, and DPS/DBS areas were determined by the image processing tool. The influence of hematocrit values (20-60%) on accuracy and precision was evaluated to determine the range within which method for DBSs is free from Hct or dependency is within acceptable limits. The validated method was applied to the determination of zonisamide levels in DPS and DBS samples obtained from patients confirming its suitability for clinical application. Finally, the distribution of zonisamide into the red blood cells was estimated by correlating its DPS and DBS levels.

| S-EPMC9369825 | biostudies-literature

Hepatitis C Core-Antigen Testing from Dried Blood Spots.

Project description:In order to expand hepatitis C virus (HCV) screening, a change in the diagnostic paradigm is warranted to improve accessibility and decrease costs, such as utilizing dried blood spot (DBS) collection. In our study, blood from 68 patients with chronic HCV infection was spotted onto DBS cards and stored at the following temperatures for one week: -80 °C, 4 °C, 21 °C, 37 °C, and alternating 37 °C and 4 °C; to assess whether temperature change during transportation would affect sensitivity. Sample was eluted from the DBS cards and tested for HCV antibodies (HCV-Ab) and HCV core antigen (core-Ag). HCV-Abs were detected from 68/68 DBS samples at -80 °C, 4 °C, 21 °C, and 67/68 at 37 °C and alternating 37 °C and 4 °C. Sensitivity of core-Ag was as follows: 94% (-80 °C), 94% (4 °C), 91% (21 °C), 93% (37 °C), and 93% (37 °C/4 °C). Not only did temperature not greatly affect sensitivity, but sensitivities are higher than previously reported, and support the use of this assay as an alternative to HCV RNA. We then completed a head-to-head comparison (n = 49) of venous versus capillary samples, and one versus two DBS. No difference in core-Ag sensitivity was observed by sample type, but there was an improvement when using two spots. We conclude that HCV-Abs and core-Ag testing from DBS cards has high diagnostic accuracy and could be considered as an alternative to HCV RNA in certain settings.

| S-EPMC6784259 | biostudies-literature

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