The palm oil industry generates several byproducts, and more than half of the dry weight of the waste is of oil palm leaf whereby the tissue is underutilized. Recently, several research studies found promising potential of oil palm fronds as a source of nutraceutical due to its bioactive properties. However, the chemical composition of the tissue is still not deciphered. Using reversed-phase liquid chromatography (LC) electrospray mass spectrometry (ESI-MS), glycosylated apigenin and luteolin were separated and identified from oil palm (Elaeis guineensis Jacq.) leaf and structures of the constituents were elucidated by collision-induced dissociation (CID) tandem MS. From 28 derivatives of the flavones, 9 compounds were conjugated with hydroxymethylglutaric (HMG) acid. Improved knowledge on oil palm especially on bioactive component of the leaf tissue will allow correlation of its beneficial effects and further promotes efficient utilization of this agriculture byproduct.
The main challenges in the purification of αS2-casein are due to the low quantity in milk and high homology with other casein subunits, i.e., αS1-casein, β-casein, and κ-casein. To overcome these challenges, the aim of this study was to develop a two-step purification to isolate native αS2-casein in goat milk from five different breeds; British Alpine, Jamnapari, Saanen, Shami, and Toggenburg. The first step of the purification was executed by anion-exchange chromatography under optimal elution conditions followed by size exclusion chromatography. Tryptic peptides from in-gel digestion of purified αS2-casein were sequenced and analyzed by LC-ESI-MS/MS. From 1.05 g of whole casein, the highest yield of αS2-casein (6.7 mg/mL) was obtained from Jamnapari and the lowest yield (2.2 mg/mL) was from Saanen. A single band of pure αS2-casein was observed on SDS-PAGE for all breeds. The αS2-casein showed coverage percentage of amino acid sequence from 76.68 to 92.83%. The two-step purification process developed herein was successfully applied for isolating native αS2-casein from goat milk with high purity, which will allow for future in vitro studies to be conducted on this protein.
A confirmatory and quantitative HPLC-tandem mass spectrometry (MS-MS) method for human chorionic gonadotropin hormone (hCG) at concentrations as low as 5 IU/l following immunoaffinity extraction of the glycoprotein from urine was developed. The extraction method involved retention of urinary hCG in the immunoaffinity column via specific antigen-antibody interaction. A variety of eluents were then used to quantitatively elute hCG from the immunoaffinity column. Qualitative and quantitative analysis of hCG were undertaken using MS-MS by identifying the amino acid sequence of the marker peptide betaT5 obtained from hCG by tryptic digestion and the peak areas of three product ions b(6)(+), b(9)(+) and y(11)(+), respectively.
A simple, rapid, specific and reliable UFLC coupled with ESI-MSMS assay method to simultaneously quantify sildenafil and N-desmethyl sildenafil, with loperamide as internal standard, was developed. Chromatographic separation was performed on a Thermo Scientific Accucore C18 column with an isocratic mobile phase composed of 0.1% v/v formic acid in purified water-methanol (20:80, v/v), at a flow rate of 0.3 mL/min. Sildenafil, N-desmethyl sildenafil and loperamide were detected with proton adducts at m/z 475.4 > 58.2, 461.3 > 85.2 and 477.0 > 266.1 in multiple reaction monitoring positive mode, respectively. Both analytes and internal standard were extracted by diethyl ether. The method was validated over a linear concentration range of 10-800 ng/mL for sildenafil and 10-600 ng/mL for N-desmethyl sildenafil with correlation coefficient (r(2) ) ≥0.9976 for sildenafil and (r(2) ) ≥0.9992 for N-desmethyl sildenafil. The method was precise, accurate and stable. The proposed method was applied to study the bioequivalence between a 100 mg dose of two pharmaceutical products: Viagra (original) and Edyfil (generic) products. AUC0-t , Cmax and Tmax were 2285.79 ng h/mL, 726.10 ng/mL and 0.94 h for Viagra and 2363.25 ng h/mL, 713.91 ng/mL and 0.83 hour for Edyfil. The 90% confidence interval of these parameters of this study fall within the regulatory range of 80-125%, hence they are considered as bioequivalent.
An analytical method that facilitated the analysis of 11 pharmaceuticals residue (caffeine, prazosin, enalapril, carbamazepine, nifedipine, levonorgestrel, simvastatin, hydrochlorothiazide, gliclazide, diclofenac-Na, and mefenamic acid) with a single pre-treatment protocol was developed. The proposed method included an isolation and concentration procedure using solid phase extraction (Oasis HLB), a separation step using high-performance liquid chromatography, and a detection procedure that applies time-of-flight mass spectrometry. The method was validated for drinking water (DW), surface water (SW), sewage treatment plant (STP) influent and effluent, and hospital (HSP) influent and effluent. The limits of quantification were as low as 0.4, 1.6, 5, 3, 2.2 and 11 ng/L in DW, SW, HSP influent and effluent, STP effluent, and STP influent, respectively. On average, good recoveries higher than 75% were obtained for most of the target analytes in all matrices. Matrix effect was evaluated for all samples matrices. The proposed method successfully determined and quantified the target compounds in raw and treated wastewater of four STPs and three hospitals in Malaysia, as well as in two SW sites. The results showed that a number of the studied compounds pose moderate to high persistency in sewage treatment effluents as well as in the recipient rivers, namely; caffeine, simvastatin, and hydrochlorothiazide. Ten out of 11 compounds were detected and quantified in 13 sampling points. Caffeine was detected with the highest level, with concentrations reaching up to 9099 ng/L in STP influent.
An extensive comparative study on the electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) mass spectrometry using automated flow injection analysis (FIA), was performed on eurycomanone (1), 13α(21)-epoxyeurycomanone (2), eurycomanol (3), eurycomanol-2-O-β-d-glucopyranoside (4), and 13,21-dihydroeurycomanone (5), the bioactive markers isolated from Eurycoma longifolia. The effects of eluent mixture (methanol or acetonitrile in water) and acidic modifiers (acetic acid, formic acid and trifluoroacetic acid) on the ionization efficiency of the markers were also investigated. The ESI in the positive ion mode with methanol containing 0.1% (v/v) acetic acid was selected for the subsequent optimization of nebulizer pressure, dry gas flow, dry gas temperature and capillary voltage to improve the sensitivity of the total ion chromatogram (TIC). Fragmentation of the analytes was further investigated by varying the capillary exit offset voltage and fragmentation amplitude in positive mode of ESI. The detection limits (LODs) were determined in isolation mode (selected ion monitoring, SIM). Their limits of detection (LODs) ranged between 0.03 and 0.1μgmL(-1) while the intra-day and inter-day precisions were less than 5.72% and 4.82%, respectively. The method was next applied for the simultaneous analysis of the markers to standardize various batches of manufactured extracts of E. longifolia for potential use as antimalarial products. Multiple Reaction Monitoring (MRM) mode was used for the quantification of analytes which gave protonated molecular ion, [M+H](+). For those without pseudo-molecular ions, SIM mode was used to quantify the analytes. The batches contained 5.65-9.95% of eurycomanone (1), 5.21-19.75% of eurycomanol (3) and 7.59-19.95% of eurycomanol-2-O-β-d-glucopyranoside (4) as major quassinoids whereas, 13α(21)-epoxyeurycomanone (2), and 13,21-dihydroeurycomanone (5) were much lower in concentrations of 0.78-3.90% and 0.47-1.76%, respectively.
The development of a reversed phase high performance liquid chromatography fluorescence method for the determination of the mycotoxins fumonisin B(1) and fumonisin B(2) by using silica-based monolithic column is described. The samples were first extracted using acetonitrile:water (50:50, v/v) and purified by using a C(18) solid phase extraction-based clean-up column. Then, pre-column derivatization for the analyte using ortho-phthaldialdehyde in the presence of 2-mercaptoethanol was carried out. The developed method involved optimization of mobile phase composition using methanol and phosphate buffer, injection volume, temperature and flow rate. The liquid chromatographic separation was performed using a reversed phase Chromolith(®) RP-18e column (100 mm × 4.6 mm) at 30 °C and eluted with a mobile phase of a mixture of methanol and phosphate buffer pH 3.35 (78:22, v/v) at a flow rate of 1.0 mL min(-1). The fumonisins separation was achieved in about 4 min, compared to approximately 20 min by using a C(18) particle-packed column. The fluorescence excitation and emission were at 335 nm and 440 nm, respectively. The limits of detections were 0.01-0.04 μg g(-1) fumonisin B(1) and fumonisin B(2), respectively. Good recoveries were found for spiked samples (0.1, 0.5, 1.5 μg g(-1) fumonisins B(1) and B(2)), ranging from 84.0 to 106.0% for fumonisin B(1) and from 81.0 to 103.0% for fumonisin B(2). Fifty-three samples were analyzed including 39 food and feeds and 14 inoculated corn and rice. Results show that 12.8% of the food and feed samples were contaminated with fumonisin B(1) (range, 0.01-0.51 μg g(-1)) and fumonisin B(2) (0.05 μg g(-1)). The total fumonisins in these samples however, do not exceed the legal limits established by the European Union of 0.8 μg g(-1). Of the 14 inoculated samples, 57.1% contained fumonisin B(1) (0.16-41.0 μg g(-1)) and fumonisin B(2) (range, 0.22-50.0 μg g(-1)). Positive confirmation of selected samples was carried out using liquid chromatography-tandem mass spectrometry, using triple quadrupole analyzer and operated in the multiple reaction monitoring mode.
The scarcity of data about the occurrence of pharmaceuticals in water bodies in Malaysia prompted us to develop a suitable analytical method to address this issue. We therefore developed a method based on solid-phase extraction combined with liquid chromatography-time of flight/mass spectrometry (SPE-LC-TOF/MS) for the analysis of sixteen prescribed and two nonprescribed pharmaceuticals that are potentially present in water samples. The levels of these pharmaceuticals, which were among the top 50 pharmaceuticals consumed in Malaysia during the period 2011-2014, in influent and effluent of five sewage treatment plants (STPs) in Bangi, Malaysia, were then analyzed using the developed method. All of the pharmaceuticals were separated chromatographically using a 5 μm, 2.1 mm × 250 mm C18 column at a flow rate of 0.3 mL/min. Limits of quantification (LOQs) were 0.3-8.2 ng/L, 6.5-89 ng/L, and 11.1-93.8 ng/L in deionized water (DIW), STP effluent, and STP influent, respectively, for most of the pharmaceuticals. Recoveries were 51-108%, 52-118%, and 80-107% from the STP influent, STP effluent, and DIW, respectively, for most of the pharmaceuticals. The matrix effect was also evaluated. The signals from carbamazepine, diclofenac sodium, and mefenamic acid were found to be completely suppressed in the STP influent. The signals from other compounds were found to be influenced by matrix effects more strongly in STP influent (enhancement or suppression of signal ≤180%) than in effluent (≤94%). The signal from prednisolone was greatly enhanced in the STP influent, indicating a matrix effect of -134%. Twelve pharmaceuticals were frequently detected in all five STPs, and caffeine, prazosin, and theophylline presented the highest concentrations among all the pharmaceuticals monitored: up to 7611, 550, and 319 ng/L in the STP influent, respectively. To the best of our knowledge, this is the first time that prazosin has been detected in a water matrix in Malaysia. Graphical abstract ᅟ.
Phyllagathis rotundifolia (Jack) Bl. (Melastomataceae) is a creeping herb found in Peninsular Malaysia and Sumatra. Traditionally, a decoction of the leaves is used in the treatment of malaria, fever and stomach ache.
Pollutants such as human pharmaceuticals and synthetic hormones that are not covered by environmental legislation have increasingly become important emerging aquatic contaminants. This paper reports the development of a sensitive and selective multi-residue method for simultaneous determination and quantification of 23 pharmaceuticals and synthetic hormones from different therapeutic classes in water samples. Target pharmaceuticals include anti-diabetic, antihypertensive, hypolipidemic agents, β2-adrenergic receptor agonist, antihistamine, analgesic and sex hormones. The developed method is based on solid phase extraction (SPE) followed by instrumental analysis using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) with 30 min total run time. River water samples (150 mL) and (sewage treatment plant) STP effluents (100 mL) adjusted to pH 2, were loaded into MCX (3 cm(3), 60 mg) cartridge and eluted with four different reagents for maximum recovery. Quantification was achieved by using eight isotopically labeled internal standards (I.S.) that effectively correct for losses during sample preparation and matrix effects during LC-ESI-MS/MS analysis. Good recoveries higher than 70% were obtained for most of target analytes in all matrices. Method detection limit (MDL) ranged from 0.2 to 281 ng/L. The developed method was applied to determine the levels of target analytes in various samples, including river water and STP effluents. Among the tested emerging pollutants, chlorothiazide was found at the highest level, with concentrations reaching up to 865 ng/L in STP effluent, and 182 ng/L in river water.