HPLC-DAD–MS Analysis of Anthocyanins in Rose Wine Made From cv. Öküzgözü Grapes,and Effect of Maceration Time on Anthocyanin Content

The anthocyanin composition of rose wine made from cv. Öküzgözü, a variety of grape native to Turkey, and the effect of maceration time (3, 6, 12, 18, or 24 h) on the anthocyanin content of the wine have been investigated. High-performance liquid chromatography with diode-array detection (HPLC-DAD) and mass spectrometry (MS) were used for anthocyanin analysis. Thirteen different anthocyanins, including five glucosides, five acetyl glucosides, and three coumaroyl glucosides were identified and quantified. It was found that an important characteristic of Öküzgözü rose wine was the presence of a large amount of malvidin-3-glucoside and its acylated esters. The total anthocyanin content of the wine increased with increasing maceration time. On the basis of sensory analysis, the most preferred wine was that produced with 3-h maceration, because of its better color and general flavor attributes.     

Determination of anthocyanins in wine based on flow-injection, liquid--solid extraction, continuous evaporation and high-performance liquid chromatography--photometric detection

A continuous method, easy to automate, for the determination of anthocyanins in wine based on the coupling of continuous liquid–solid extraction, evaporation, HPLC individual separation and photometric detection is proposed. The target analytes are removed from the wine in a continuous fashion using a C₁₈ minicolumn and eluted with an aqueous solution (pH 2) with 16% acetonitrile. The eluted fraction is concentrated by solvent evaporation assisted by heat and dragging off the vapour using a flow of N₂. For in-line preconcentration, a continuous evaporation module was designed and located in the manifold between the solid-phase minicolumn and the injection valve of the chromatograph. In this way, injection of the sample into the dynamic system leads the plug through it for liquid–solid extraction of the anthocyanins, partial evaporation of the eluent (with a preconcentration factor as required) and transport to the high-pressure injection valve of the chromatograph, where individual separation and subsequent photometric detection take place. The method thus developed for the determination of malvidin-3-glucoside, cyanidin-3-glucoside and peonidin-3-glucoside anthocyanins in Spanish red wines is more sensitive than the batch manual method based on the same steps, has better linearity of the calibrations curves with lower detection limits and much wider determination range for the most abundant anthocyanins in wine. In addition, the method can be fully automated with low acquisition and maintenance costs.     

Stability of anthocyanins extracted from grape skins

Summary The extractability and stability of anthocyanins from the skins ofVitis vinifera were determined at different pH values. Anthocyanins were extracted using acetone, partitioned with chloroform and pre-purified by solid-phase extraction (SPE). They were analysed by RP-HPLC, and the kinetic parameters of decomposition were calculated. The total monomeric anthocyanin content was determined by spectrophotometry. Anthocianins were well separated by RP-HPLC. The efficiency of extraction depended strongly on the pH of the extracting agent and on the character of the pigment to be extracted. The amount of anthocyanins decreased with increasing duration of storage, more so at elevated temperatures. Presented at Balaton Symposium '01 on High-Performance Separation Methods Siófok, Hungary, September 2–4, 2001.     

Optimization of microwave‐assisted extraction for anthocyanins,polyphenols, and antioxidants from raspberry (Rubus Coreanus Miq.) using response surface methodology

Anthocyanins (Acys), polyphenols, and antioxidants were extracted from raspberry (Rubus Coreanus Miq.) using a highly efficient microwave‐assisted extraction technique. Different solvents, including methanol, ethanol, and acetone, were tested. The colors of the extracts varied from light yellow to purple red or dark red. SEM and other nutrient analyses verified that ethanol was the most favorable medium for the microwave‐assisted extraction of raspberry due to its high output and low toxicity. Effects of process parameters, including microwave power, irradiation time, and solvent concentration, were investigated through response surface methodology. Canonical analysis estimated that the highest total Acys content, total polyphenols content, and antioxidant activity of raspberry were 17.93 mg cyanidin‐3‐O‐glucoside equivalents per gram dry weight, 38.57 mg gallic acid equivalents per gram dry weight, and 81.24%, respectively. The polyphenol compositions of raspberry extract were identified by HPLC with diode array detection, and nine kinds of polyphenols were identified and quantified, revealing that chlorogenic acid, syringic acid, and rutin are the major polyphenols contained in raspberry fruits. Compared with other fruits and vegetables, raspberry contains higher Acy and polyphenol contents with stronger antioxidant activity, suggesting that raspberry fruits are a good source of natural food colorants and antioxidants.     

Rapid quantitative analysis of individual anthocyanin content based on high‐performance liquid chromatography with diode array detection with the pH differential method

A new quantitative technique for the simultaneous quantification of the individual anthocyanins based on the pH differential method and high‐performance liquid chromatography with diode array detection is proposed in this paper. The six individual anthocyanins (cyanidin 3‐glucoside, cyanidin 3‐rutinoside, petunidin 3‐glucoside, petunidin 3‐rutinoside, and malvidin 3‐rutinoside) from mulberry (Morus rubra) and Liriope platyphylla were used for demonstration and validation. The elution of anthocyanins was performed using a C₁₈ column with stepwise gradient elution and individual anthocyanins were identified by high‐performance liquid chromatography with tandem mass spectrometry. Based on the pH differential method, the high‐performance liquid chromatography peak areas of maximum and reference absorption wavelengths of anthocyanin extracts were conducted to quantify individual anthocyanins. The calibration curves for these anthocyanins were linear within the range of 10–5500 mg/L. The correlation coefficients (r²) all exceeded 0.9972, and the limits of detection were in the range of 1–4 mg/L at a signal‐to‐noise ratio ≥5 for these anthocyanins. The proposed quantitative analysis was reproducible with good accuracy of all individual anthocyanins ranging from 96.3 to 104.2% and relative recoveries were in the range 98.4–103.2%. The proposed technique is performed without anthocyanin standards and is a simple, rapid, accurate, and economical method to determine individual anthocyanin contents.     

Characterization and quantification of anthocyanins in selected artichoke (Cynara scolymus L.) cultivars by HPLC–DAD–ESI–MS n

The anthocyanin pattern of artichoke heads (Cynara scolymus L.) has been investigated by high-performance liquid chromatography–electrospray ionization mass spectrometry. For this purpose a suitable extraction and liquid chromatographic method was developed. Besides the main anthocyanins—cyanidin 3,5-diglucoside, cyanidin 3-glucoside, cyanidin 3,5-malonyldiglucoside, cyanidin 3-(3′′-malonyl)glucoside, and cyanidin 3-(6′′-malonyl)glucoside—several minor compounds were identified. Among these, two peonidin derivatives and one delphinidin derivative were characterized on the basis of their fragmentation patterns. To the best of our knowledge this is the first report on anthocyanins in artichoke heads consisting of aglycones other than those of cyanidin. Quantification of individual compounds was performed by external calibration. Cyanidin 3-(6′′-malonyl)glucoside was found to be the major anthocyanin in all the samples analyzed. Total anthocyanin content ranged from 8.4 to 1,705.4 mg kg⁻¹ dry mass.      

CZE separation of strawberry anthocyanins with acidic buffer and comparison with HPLC

Anthocyanins, the major colourants of strawberries, are polar pigments that are positively charged at low pH. Herein, we have assessed a new analytical method for the separation of anthocyanins using CZE. Acidic buffer solutions (pH <2) were employed in order to maintain pigments in the cation flavylium form and achieve high molar absorptivity at 510 nm. These spectral properties enabled us to identify strawberry anthocyanins in a preliminary stage by detection in the visible range, although the method was optimised at 280 nm to obtain the best S/N. The effects of buffer composition highlighted the necessity of adding an organic modifier to the running buffer to obtain a suitable separation. The electrophoretic method permitted the separation of the three main anthocyanins of strawberry extracts, namely pelargonidin 3-glucoside (Pg-glu), pelargonidin 3-rutinoside and cyanidin 3-glucoside. The electrophoretic results, expressed as retention time and separation efficiency of the major anthocyanin (Pg-glu), were compared to those achieved in HPLC, the analytical technique traditionally used for the investigation of anthocyanins in vegetable matrix. The content of Pg-glu in strawberries (cv. Camarosa), calculated with HPCE and HPLC methods, resulted respectively in 11.41 mg/L and 11.37 mg/L.     

High performance liquid chromatography analysis of wine anthocyanins revisited: Effect of particle size and temperature

The complex anthocyanin fraction of red wines poses a demanding analytical challenge. We have found that anthocyanins are characterised by extremely low optimal chromatographic velocities, and as a consequence generic HPLC methods suffer from limited resolving power. Slow on-column inter-conversion reactions, particularly between carbinol and flavylium species, are shown to occur on the same time scale as chromatographic separation, leading to increased plate heights at normal chromatographic velocities. In order to improve current routine HPLC separations, the use of small (1.7 μm) particles and high temperature liquid chromatography (HTLC) were investigated. 1.7 μm particles provide better efficiency and higher optimal linear velocities, although column lengths of ∼20 cm should be used to avoid the detrimental effects of conversion reactions. More importantly, operation at temperatures up to 50 °C increases the kinetics of inter-conversion reactions, and implies significantly improved efficiency under relatively mild analysis conditions. It is further demonstrated using relevant kinetic data that no on-column thermal degradation of these thermally labile compounds is observed at 50 °C and analysis times of <2 h.