Determination of caffeine in dietary supplements by miniaturized portable liquid chromatography.

In this study three miniaturized liquid chromatography (LC) instruments have been evaluated and compared for the analysis of caffeine in dietary supplements, namely a benchtop capillary LC (capLC) system, a benchtop nano LC (nanoLC)system and a portable LC system. Commercial products derived from different sources of caffeine have been analyzed. Under optimized conditions, the methods based on benchtop systems were superior in terms of sensitivity. The limits of detection (LODs) found with the capLC and nanoLC systems were 0.01 and 0.003 µg mL-1, respectively, whereas the LOD obtained with the portable LC instrument was of 1 µg mL-1. The portable LC-based method was superior in terms of simplicity and throughput (total analysis time < 15 min). On the basis of the results obtained, a new method for the rapid measurement of caffeine in dietary supplements by portable miniaturized LC is presented. This method provided good linearity within the 1-20 µg mL-1 interval, and it allowed the quantification of caffeine even in products derived from decaffeinated green coffee extracts. The contents of caffeine found with the proposed portable LC method in the real samples analyzed ranged from 1.38 to 7 mg per gram of product, which were values statistically equivalent to those found with the benchtop capLC and nanoLC methods, being the precision, expressed as relative standard deviation (RDS), of 2 -14% (n = 3). The proposed portable LC based method can be used as a simple and rapid alternative to estimate the quality, effectiveness and safety of dietary supplements, regarding their caffeine content.

Exploring hand-portable nano-liquid chromatography for in place water analysis: Determination of trimethylxanthines as a use case.

Analytical performance and optimization of figures of merit of a portable nano liquid chromatograph (NanoLC) with UV detection at 255 nm have been established for in place analysis. Methylxanthines: caffeine, theophylline and theobromine were selected as target analytes. A fast lab method based on IT-SPME coupled on line with capillary liquid chromatograph (CapLC) with diode array detection (DAD) was employed for comparative studies. IT-SPME and solid phase extraction were coupled off-line to NanoLC for improving instrumental parameters, mainly detection capacity and selectivity. IT-SPME or SPE/portable NanoLC based methods were superior in terms of chromatographic resolution and organic solvent consumption per sample, around 200 µL vs 10 mL for IT-SPME-CapLC-DAD. Limits of detection (LODs) obtained with the SPE/portable NanoLC were 2-10 ng/mL, which can be suitable for testing the presence of the analytes in several environmental waters in the field. As predictable, the lab method provided better LODs, between 0.1 and 0.5 ng/mL. Good linearity was achieved for both methods and precision was similar for them (≤7%). Both systems were tested for the analysis of real water samples with suitable results.

On-line in-tube solid phase microextraction coupled to capillary liquid chromatography-diode array detection for the analysis of caffeine and its metabolites in small amounts of biological samples.

In-tube solid phase microextraction (IT-SPME) coupled on-line to capillary liquid chromatography with diode array detection provides a simple and fast analytical methodology for the simultaneous quantitation of caffeine and its three primary metabolites (theobromine, paraxanthine and theophylline) in micro samples of serum, saliva and urine matrices. The sample amount required for one analysis was only 2.5 µL of saliva, 6.25 µL of serum or 40 µL of urine, a requirement for its implementation in a hospital laboratory for preterm newborns, where sample availability is a major problem. In standard conditions, 25 µL of diluted saliva or serum (or 100 µL of urine) were processed by IT-SPME in 30 cm of commercially available capillary GC column coated with ZB-FFAP (100% nitroterephthalic modified polyethylene glycol). The retained compounds were desorbed from the IT-SPME capillary by the mobile phase (a gradient mixture of water and methanol) and the separation was carried out in a C18 column (150 mm × 0.5 mm i.d., 5 µm particle size). Analytes eluted before 14 min, at a flow rate of 15 µL min-1, and were detected by absorbance at 275 nm. The calibration graphs presented good linearity (R2 > 0.99), without the presence of matrix effect, and recoveries between 84 and 112% were obtained. Limits of detection (S/N = 3) were 0.1 µg·mL-1 in serum and 0.5 µg·mL-1 in saliva and urine samples, for all compounds, and the intra- and inter-day variation coefficients (n = 3) were between 3 and 17%. Analytical figures of merit were similar to those proposed by other methodologies, but using lower sample volume and a faster and simpler sample treatment and analysis. Paired samples of serum and saliva from preterm newborns treated with caffeine at the pediatric intensive care unit were analyzed by the method, with statistically equivalent results for caffeine concentrations.

Quantitative Analysis of Terpenic Compounds in Microsamples of Resins by Capillary Liquid Chromatography.

A method has been developed for the separation and quantification of terpenic compounds typically used as markers in the chemical characterization of resins based on capillary liquid chromatography coupled to UV detection. The sample treatment, separation and detection conditions have been optimized in order to analyze compounds of different polarities and volatilities in a single chromatographic run. The monoterpene limonene and the triterpenes lupeol, lupenone, ß-amyrin, and α-amyrin have been selected as model compounds. The proposed method provides linear responses and precision (expressed as relative standard deviations) of 0.6% to 17%, within the 0.5-10.0 µg mL-1 concentration interval; the limits of detection (LODs) and quantification (LOQs) were 0.1-0.25 µg mL-1 and 0.4-0.8 µg mL-1, respectively. The method has been applied to the quantification of the target compounds in microsamples. The reliability of the proposed conditions has been tested by analyzing three resins, white copal, copal in tears, and ocote tree resin. Percentages of the triterpenes in the range 0.010% to 0.16% were measured using sample amounts of 10-15 mg, whereas the most abundant compound limonene (≥0.93%) could be determined using 1 mg portions of the resins. The proposed method can be considered complementary to existing protocols aimed at establishing the chemical fingerprint of these kinds of samples.