•The serum sample preparation uses protein precipitation alone.•Well-defined acetonitrile content releases vitamin D metabolites from the proteins.•Protein precipitation is carried out in two ...stages.•Chromatographic method enables separation of interferents and epimeric forms.
Despite the fact that more than 90% of vitamin D analysis are performed using immuno-enzymatic techniques, it is liquid chromatography coupled with tandem mass spectrometry that is currently the reference method. It allows for specific and selective analysis of all relevant vitamin D metabolites from a variety of biological materials, including serum or a dried blood spot. This paper presents development of a fast, cheap and high-throughput method of serum sample preparation using protein precipitation. For this purpose, organic solvent is used. Several substances were tested, including acetonitrile, methanol and their mixtures with zinc sulfate. However, the highest recovery values for the vitamin D metabolites were obtained for acetonitrile, with an organic solvent to serum ratio of 8:1. The preparation of a sample is carried out in 96-well plates and takes an hour and a half, together with a derivatization reaction using Cookson-type reagent 4-(4'-dimethylaminophenyl)-1,2,4-triazoline-3,5-dione. Due to the fact that vitamin D metabolites are bound to proteins, the relationship between the content of organic solvent in the sample preparation process and their release from the protein complex was examined. The results indicate that the organic solvent content should be 30-70% in order to completely release the tested compounds from the proteins. In addition, the developed chromatographic method has eliminated false positive signals for the 24,25(OH)2D3 metabolite. Total analysis time is 5.5 min., while maintaining resolution necessary to separate the analyzed compounds.
Implementing the European Foundation for Quality Management (EFQM) business excellence model in organizations is time- and cost-consuming. The integration of artificial intelligence (AI) into the ...EFQM business excellence model is a promising approach to improve the efficiency and effectiveness of excellence in organizations. This research paper’s integrated framework follows the ISO/IEC 23053 standard in addressing some of the concerns related to time and cost associated with the EFQM model, achieving higher EFQM scores, and hence operational excellence. A case study involving a UAE government organization serves as a sample to train the AI framework. Historical EFQM results from different years are used as training data. The AI framework utilizes the unsupervised machine learning technique known as k-means clustering. This technique follows the ISO/IEC 23053 standard to predict EFQM output total scores based on criteria and sub-criteria inputs. This research paper’s main output is a novel AI framework that can predict EFQM scores for organizations at an early stage. If the predicted EFQM score is not high enough, then the AI framework provides feedback to decision makers regarding the criteria that need reconsideration. Continuous use of this integrated framework helps organizations attain operational excellence. This framework is considered valuable for decision makers as it provides early predictions of EFQM total scores and identifies areas that require improvement before officially applying for the EFQM excellence award, hence saving time and cost. This approach can be considered as an innovative contribution and enhancement to knowledge body and organizational practices.
Sexual concerns are known to be common in women suffering from multiple sclerosis (MS) but definite data on the prevalence of particular sexual dysfunctions (SD) remain unclear. Previous studies ...brought inconsistent findings and rely on small groups of patients or use of unvalidated assessment methods. The aim of this research was to evaluate the prevalence of SD in women with MS using validated clinimetric scales. 137 female inpatients with MS diagnosis were interviewed, completed The Female Sexual Function Questionnaire SFQ28 and underwent neurological assessment. Only 2.2 % of patients had ever discussed their sexual concerns with a physician. 70.1 % reported sexual activity. At least one SD could be found in 82.5 % of patients, hypoactive sexual desire (57.7 %), arousal dysfunction (decreased genital sensation in 47.3 %, decreased lubrication in 48.4 %, decreased subjective arousal in 45.2 %) and orgasmic dysfunction (39.8 %) being the most probable. SD were less likely in women who assessed their relationship positively but more common in older patients and those who had a positive history of depression. The prevalence of SD was higher comparing to the majority of studies by other authors. In conclusion, SD are very common in female patients with MS and permanently overlooked by medical professionals. Therefore, the assessment of sexual function should be implemented in all patients after the diagnosis of MS. Further research is needed for better understanding of the sexuality of this particular population in order to establish targets for therapeutic intervention.
There are two forms of vitamin D which are essential to the human body, i.e. vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol). The inactive metabolites of vitamin D are commonly used for ...quantitative analysis because of their longer half-life, stability, and relatively high blood concentrations. This paper presents the development of a high-throughput and sensitive method for determining four vitamin D metabolites in dried blood spots using liquid chromatography coupled with tandem mass spectrometry. This method allows for the determination of 25(OH)D2 and 25(OH)D3 concentrations, as well as the epimeric form 3-epi-25(OH)D3 and 24,25(OH)2D3. The analyzed material is capillary blood taken from the fingertip, deposited on filter paper. Four different chromatographic columns were tested to separate all compounds, in particular, the epimeric form. The column of choice was F5 (Phenomenex, Torrance, CA, USA). In order to prove the consistency between the results for DBS, used as an alternative biological matrix, and serum, comparative studies of these two materials were carried out in nearly 100 individuals. The results indicated their positive correlation. The evaluation of short-term stability of metabolites in DBS within the month showed no change in metabolite concentration. During the validation, the impact of the matrix on the ionization of the tested compounds was evaluated. Capillary blood and venous blood collected for different anticoagulants were also compared. The smallest differences in the results were obtained for citrate. In order to achieve a limit of quantitation of 0.2 ng ml-1, sample preparation involved derivatization using a Cookson-type reagent, 4-(4'-dimethylaminophenyl)-1,2,4-triazoline-3,5-dione (DAPTAD).
Purpose
The relationship between Hashimoto's thyroiditis and vitamin D concentration was already presented in many studies. The aim of this study was to analyze the differences in the concentration ...of vitamin D metabolites between healthy women and women with Hashimoto's thyroiditis (HT).
Methods
The quantitative analysis of five vitamin D metabolites was carried out using liquid chromatography coupled with tandem mass spectrometry. The analyzed materials were serum and dried blood spots (DBS). The results obtained for the two materials were also compared.
Results
No statistically significant differences were found in the mean concentration of the 25(OH)D
3
, 25(OH)D
2
, 24,25(OH)
2
D
3
, 1,25(OH)
2
D
3
metabolites between the test and the control groups. However, a strong correlation was found between the 25(OH)D
3
and 24,25(OH)
2
D
3
metabolites.
Conclusion
The study showed that healthy women and women with Hashimoto's disease had similar concentration of vitamin D metabolites. Research also proved that DBS is a good alternative to serum. The differences in 25(OH)D concentration were not statistically significant (17.0 and 15.5 ng mL
−1
for serum and DBS, respectively). DBS can be successfully used in research on a large group of people, since the process of material collection, as well as sample preparation, is fast and simple. It is also easy to transport and store, and requires small volume of blood.
•Silica-based stationary phases of great efficiency were synthesized for IC.•New type of dendrimeric anion-exchanger was applied for the separation of nucleotides.•Novel anion-exchanger was applied ...for analysis of nucleotides in milk in 6min.
The main aim of the present study was to develop a method for the separation of 5′-monophosphate nucleotides with the use of ion chromatography. Novel dendrimeric stationary phases were used for this purpose. The effects exerted by the type of anion-exchanger support (silica or polymeric) and the number of stationary phase layers on nucleotide retention were studied. A silica-based dendrimeric anion-exchanger was most suitable for analyzing the studied compounds. An increase in the number of layers enhanced nucleotide retention inside the column. The separation efficiency of the studied compounds was tested at various concentrations of the mobile phase buffer. At higher phosphate buffer concentrations, nucleotide resolutions were achieved in 6min. Three commercially available infant milk formulas were analyzed to verify the applicability of the studied method. Solid phase extraction was used for sample cleanup and concentration. The limit of quantification of nucleotides was 0.40μg/ml, and the method was linear in the concentration range of 0.40–20.6μgml−1.
To develop a reliable and valid clinical scale measuring the severity of ataxia.
The authors devised the Scale for the Assessment and Rating of Ataxia (SARA) and tested it in two trials of 167 and ...119 patients with spinocerebellar ataxia.
The mean time to administer SARA in patients was 14.2 +/- 7.5 minutes (range 5 to 40). Interrater reliability was high, with an intraclass coefficient (ICC) of 0.98. Test-retest reliability was high with an ICC of 0.90. Internal consistency was high as indicated by Cronbach's alpha of 0.94. Factorial analysis revealed that the rating results were determined by a single factor. SARA ratings showed a linear relation to global assessments using a visual analogue scale, suggesting linearity of the scale (p < 0.0001, r(2) = 0.98). SARA score increased with the disease stage (p < 0.001) and was closely correlated with the Barthel Index (r = -0.80, p < 0.001) and part IV (functional assessment) of the Unified Huntington's Disease Rating Scale (UHDRS-IV) (r = -0.89, p < 0.0001), whereas it had only a weak correlation with disease duration (r = 0.34, p < 0.0002).
The Scale for the Assessment and Rating of Ataxia is a reliable and valid measure of ataxia, making it an appropriate primary outcome measure for clinical trials.
Although ataxia is by definition the prominent symptom of ataxia disorders, there are various neurological signs that may accompany ataxia in affected patients. Reliable and quantitative assessment ...of these signs is important because they contribute to disability, but may also interfere with ataxia. Therefore we devised the Inventory of Non-Ataxia Signs (INAS), a list of neurological signs that allows determining the presence and severity of non-ataxia signs in a standardized way. INAS underwent a rigorous validation procedure that involved a trial of 140 patients with spinocerebellar ataxia (SCA) for testing of inter-rater reliability and another trial of 28 SCA patients to assess short-term intra-rater reliability. In addition, data of the ongoing EUROSCA natural history study were used to determine the reproducibility, responsiveness and validity of INAS. Inter-rater reliability and short-term test–retest reliability was high, both for the total count and for most of the items. However, measures of responsiveness, such as the smallest detectable change and the clinically important change were not satisfactory. In addition, INAS did not differentiate between subjects that were subjectively stable and those that worsened in the 2-year observation period. In summary, INAS and INAS count showed good reproducibility, but unsatisfactory responsiveness. The present analysis and published data from the EUROSCA natural history study suggest that INAS is a valid measure of extracerebellar involvement in progressive ataxia disorders. As such, it is useful as a supplement to the measures of ataxia, but not as a primary outcome measure in future interventional trials.
To obtain quantitative data on the progression of the most common spinocerebellar ataxias (SCAs) and identify factors that influence their progression, we initiated the EUROSCA natural history study, ...a multicentric longitudinal cohort study of 526 patients with SCA1, SCA2, SCA3, or SCA6. We report the results of the 1- and 2-year follow-up visits.
As the primary outcome measure we used the Scale for the Assessment and Rating of Ataxia (SARA, 0-40), and as a secondary measure the Inventory of Non-Ataxia Symptoms (INAS, 0-16) count.
The annual increase of the SARA score was greatest in SCA1 (2.18 ± 0.17, mean ± SE) followed by SCA3 (1.61 ± 0.12) and SCA2 (1.40 ± 0.11). SARA progression in SCA6 was slowest and nonlinear (first year: 0.35 ± 0.34, second year: 1.44 ± 0.34). Analysis of the INAS count yielded similar results. Larger expanded repeats and earlier age at onset were associated with faster SARA progression in SCA1 and SCA2. In SCA1, repeat length of the expanded allele had a similar effect on INAS progression. In SCA3, SARA progression was influenced by the disease duration at inclusion, and INAS progression was faster in females.
Our study gives a comprehensive quantitative account of disease progression in SCA1, SCA2, SCA3, and SCA6 and identifies factors that specifically affect disease progression.
To determine the longitudinal metric properties of recently developed clinical assessment tools in spinocerebellar ataxia (SCA).
A subset of 171 patients from the EUROSCA natural history study cohort ...(43 SCA1, 61 SCA2, 37 SCA3, and 30 SCA6) were examined after 1 year of follow-up. Score changes and effect size indices were calculated for clinical scales (Scale for the Assessment and Rating of Ataxia SARA, Inventory of Non-Ataxia Symptoms INAS), functional tests (SCA Functional Index SCAFI and components), and a patient-based scale for subjective health status (EQ-5D visual analogue scale EQVAS). Responsiveness was determined in relation to the patient's global impression (PGI) of change and reproducibility described as retest reliability for the stable groups and smallest detectable change.
Within the 1-year follow-up period, SARA, INAS, and SCAFI but not EQVAS indicated worsening in the whole group and in the groups with subjective (PGI) worsening. SCAFI and its 9-hole pegboard (9HPT) component also deteriorated in the stable groups. Standardized response means were highest for 9HPT (-0.67), SARA (0.50), and SCAFI (-0.48) with accordingly lower sample size estimates of 143, 250, or 275 per group for a 2-arm interventional trial that aims to reduce disease progression by 50%. SARA and EQVAS performed best to distinguish groups classified as worse by PGI. All scales except EQVAS reached the criterion for retest reliability.
While both the Scale for the Assessment and Rating of Ataxia and the SCA Functional Index (SCAFI) (and its 9-hole pegboard component) had favorable measurement precision, the clinical relevance of SCAFI and 9-hole pegboard score changes warrants further exploration. The EQ-5D visual analogue scale proved insufficient for longitudinal assessment, but validly reflected patients' impression of change.
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