Analysis of Isoflavones in Foods.

In recent years the nutritional and bioactive properties of foods are being intensively investigated with a view to control, in addition to food quality, their possible influence on human health. Because of this, there is a growing demand for rapid, selective, sensitive, and validated methods for analysis and quantification. Bioactive plant compounds include those with weak estrogenic activity (phytoestrogens), among which are the isoflavones. Some of the beneficial activities that have been attributed to isoflavones are anticarcinogenic activity, the prevention of cardiovascular disease, the improvement of bone health, and antioxidant activity. The objective of this work is to provide an updated review of the methods used in sample preparation and subsequent analysis for the determination of isoflavones in food samples, including both soybean and soy products, as well as other foods with low isoflavone contents. The review focuses on the most common sample preparation techniques used during the last 10 years, including both conventional solvent extraction and other more recent extraction techniques. Separation and detection methods, including current trends in liquid chromatography analysis, such as the use of monolithic columns or ultra-high-pressure liquid chromatography, are also discussed.

Determination of tocopherols and sitosterols in seeds and nuts by QuEChERS-liquid chromatography.

In the present work a simple, reliable and affordable sample treatment method for the simultaneous analysis of tocopherols and free phytosterols in nuts was developed. Analyte extraction was carried out using the QuEChERS methodology and analyte separation and detection were accomplished using HPLC-DAD. The use of this methodology for the extraction of natural occurring substances provides advantages such as speed, simplicity and ease of use. The parameters evaluated for the validation of the method developed included the linearity of the calibration plots, the detection and quantification limits, repeatability, reproducibility and recovery. The proposed method was successfully applied to the analysis of tocopherols and free phytosterols in samples of almonds, cashew nuts, hazelnuts, peanuts, tiger nuts, sun flower seeds and pistachios.

Comparative study of the methodology used in the extraction of isoflavones from legumes applying a modified QuEChERS approach.

INTRODUCTION: Isoflavones are phytochemicals of great interest because of their association with a large variety of positive effects on human health. The major sources of isoflavones in the diet are plants of the Leguminosae family, especially soybeans, although many other legumes more widely consumed in the Mediterranean diet have also been reported to contain these compounds. In previous work we extracted isoflavones from samples using a modified QuEChERS (Quick Easy Cheap Effective Rugged Safe) methodology. OBJECTIVE: To compare different methods for placing the sample and the solvent in contact to optimise the extraction of isoflavones from legumes (chickpeas, lentils and white beans) using a modified QuEChERS methodology. METHOD: Five different approaches to sample agitation were tested: vortex agitation, thermostatted stirring agitation and thermostatted tray shaking, and a thermostatted ultrasound bath and an ultrasound probe. To evaluate the different methodologies a modified QuEChERS approach was used as the extraction method. The separation and quantification of isoflavones was carried out using liquid chromatography-triple quadrupole/mass spectrometry (LC-MS/MS). RESULTS: The best methods were found upon using a thermostatted shaking tray for the extraction of chickpeas and white beans and the ultrasound probe for lentil samples. These methods were chosen based on the highest amount of analytes obtained as well as the best recovery values. CONCLUSION: Determination of isoflavones in foods may be affected by the different methods used to place the sample and the solvent in contact in the extraction step. The main advantages of the proposed extraction procedures are their simplicity, speed, reliability and low cost.

Pressurized liquid extraction as a sample preparation method for the analysis of isoflavones in pulses.

In this work, we describe a rapid and simple analytical method that exploits pressurized liquid extraction (PLE) and liquid chromatography with diode array detection for the determination of isoflavones in samples of Spanish pulses. Confirmation of the analytes present was performed using ion-trap mass spectrometry. To optimize the PLE extraction, variables such as the dispersing agent, type of solvent and sample amount, and the experimental parameters, such as temperature and the number of extraction cycles, were studied. Separation was carried out using a reverse-phase C18 with polar endcapping as the stationary phase and acetonitrile/water with 0.2 % of formic acid, under a gradient regime, as the mobile phase. Optimal extraction of formononetin and biochanin-A from chickpeas with PLE was achieved using Hydromatrix as a dispersant agent, methanol/water (50:50), a temperature of 90 °C, and three cycles. The same optimal conditions-except methanol/water (75:25)-for solvent extraction were obtained for the extraction of daidzin, genistin, and formononetin from lentils. Recoveries ranged from 97 to 110 %, and standard deviations lower than 20 % were obtained. The contents obtained for daidzin in lentils using the proposed method were not significantly different from those obtained using another official method of analysis.