RESUMEN
INTRODUCTION AND OBJECTIVE: Fatty acids (FAs) influence cell and tissue metabolism, function, responsiveness to hormonal and other signals in addition to maintenance of membrane integrity of cells. ß-Thalassemia is a prevalent inherited blood disorder characterized by abnormal red cell membrane structure and function. Induction of HbF by hydroxyurea (HU) is an enduring therapeutic intervention to manage this. Therefore, in the present study we have carried out the quantification of thirteen free fatty acids to disclose the prognosis of HU in ß-thalassemia. METHODS: FAs quantification was carried out using GC-MRM-MS method in the serum of 98 cases of ß-thalassemia patients and out of which samples from 34 patients were collected before and after treatment with HU in addition to healthy controls (n = 31). RESULTS: Using the combination of random forest (RF) with GC-MRM-MS we were able to establish a classification and prediction model that can discriminate the ß-thalassemia from healthy as well as from HU treated group. Docosanoic acid (C-22:0) was most significantly altered in ß-thalassemia as compared to healthy at p-value of 8.3 × 10-09 while erucic acid (C-22:1 Δcis-13) can be used as potential marker of HU prognosis because its level became significantly dissimilar at p-value of 3.7 × 10-04 in same patients in response to HU. However, nervonic acid (C-24:1 Δcis-15) was found to be the key player in effectively separating three groups. CONCLUSION: In inference, we have noticed that HU therapy also rectifies the serum fatty acid profile in addition to its reported affect i.e. HbF induction in ß-thalassemia patients.
Asunto(s)
Ácidos Grasos/sangre , Hidroxiurea/uso terapéutico , Talasemia beta/sangre , Biomarcadores/sangre , Humanos , Talasemia beta/tratamiento farmacológicoRESUMEN
An exhaustive GC-MS sample preparation, derivatization, mass fragmentation and acquisition study was performed, for the simultaneous analysis of chlorophenols (CPs). Selected species were 2-CP, 3-CP, 4-CP, 3,5-dichlorophenol (diCP), 2,5-diCP, 2,6-diCP, 2,4-diCP, 2,3-diCP, 3,4-diCP 2,4,6-trichlorophenol (triCP), 2,4,5-triCP, 2,3,4-triCP, 2,3,4,6-tetrachlorophenol (tetraCP) and pentachlorophenol (pentaCP), in total 14 compounds. As novelties to the field, basic researches, like systematic derivatization, mass fragmentation and acquisition methods have been optimized for the trimethylsilyl (TMS) ether derivatives of CPs. The reactivity of chlorophenols with silylating agents has not been systematically analyzed. Here, we studied the reactivity of 14 chlorophenols with five silylating reagents. The three acquisition techniques, the full scan (FS), the multiple ion monitoring (MIM), and the currently optimized multiple reaction monitoring (MRM) methods, have been compared. We developed a new analytical approach, simultaneously monitoring the fragmentation pattern of the (35)Cl and the (37)Cl containing fragment ions both as precursor and as product ions. This principle resulted in remarkable specificity and sensitivity of detection and quantification; particularly in the cases of the tetraCP and pentaCP derivatives containing the (35)Cl and the (37)Cl fragment ions at an approximate ratio of <1:1. Detailed documentation of the loss of HCl via fragmentation processes, without decomposition of the benzene ring, was attributed to the "ring-walk" mechanism described first for monochlorophenol. Critical evaluation of the derivatization and acquisition protocols was collated and validated with the same characteristics. Data of six point calibration along with the corresponding relative standard deviation percentage (RSD%) values, in the line of FS, MIM and MRM methods (r(2): 0.9987, 0.9992, 0.9989; RSD%: 8.7, 5.6, 8.1), proved to be independent on the acquisition processes. The practical utility of the optimized MRM acquisition techniques was confirmed by the quantitation of the CP contents of Danube River, tap water and distilled water samples. Results confirmed at the first time the primary importance of the MRM acquisition method, even in comparison to the MIM one: we revealed that distilled water contains higher chlorophenol content than tap water, which might have a great significance for the water industry.