Mellow yellow: Crocin in food and drinks measured by HPLC

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  • Published: Feb 21, 2011
  • Author: Steve Down
  • Channels: HPLC
thumbnail image: Mellow yellow: Crocin in food and drinks measured by HPLC

Easy extraction of crocin from foods for HPLC

Natural compounds are regaining acceptance within some sectors of the food industry due to increasing health fears about some synthetic additives. Although they can be more expensive, the fact that they are natural lends an aura of safety in the eyes of many consumers.

One of the most widely used natural colorants is crocin, sometimes called crocin yellow, which is a common food dye in the Far East. It is also an ingredient of traditional Chinese medicine, where it has been applied to the treatment of various ailments, including diabetes, painful urination and haematuria.

Crocin is responsible for the colour of saffron, which is prepared from flowers of the crocus, Crocus sativus L., and it is also extracted from the fruits of the gardenia, Gardenia jasminoides Ellis. As a solid it exists as a red powder but it imparts a yellow colour to foods.

For control purposes, it is important to be able to confirm the amounts of natural colorants in foods, just as it is with synthetic additives. However, a group of Chinese scientists has observed that there is a general lack of published methods for natural food additives.

So, Zhenxia Du and Wenhan Yang from the Beijing University of Chemical Technology and Jinhua Wang and Xiaolin Li from the Beijing Entry-Exit Inspection and Quarantine Bureau devised a novel method for the determination of crocin in a number of foods.

One of the major problems in food analysis is the complex and varied nature of the food matrix itself. The researchers had previously attempted to use a polyamide adsorbent for extracting natural colours but found it to be unsatisfactory for several reasons, including analyte decomposition when using alkaline ammonia, a common eluent for this type of material, and low elution rates with organic solvents.

They turned to macroporous resins based on poly(styrene-divinylbenzene), which are acquiring a good reputation for natural products purification. Three types of resin were tested and optimised and the best one gave recoveries of 82% following washes with acetic acid and water and elution with an aqueous ethanol solution.

Furthermore, resin regeneration without loss of performance was achieved for more than ten cycles, giving them an advantage over conventional SPE cartridges which tend to be used only once before disposal.


Fast HPLC for major crocin component

The extracts were analysed by ultrahigh pressure liquid chromatography (UPLC) which brought about elution within 5 minutes on a short column. It was prepared from commercial high-strength silica which was designed to separate polar compounds.

The principal component of crocin is a carotenoid diester with the disaccharide gentiobiose but there are also minor amounts of the gentiobiose monoester, as well as the glucosyl diester.

The best separation was achieved using a gradient of acetonitrile which brought about the separation of the three crocins for UV detection at 440 nm. The gentiobiose diester eluted first at 2.31 minutes, giving the largest peak which was consistent with it being the major crocin component.

The diglucose ester gave a far smaller peak at 2.75 minutes and the gentiobiose monoester eluted as two small isomeric peaks in just less than 4 minutes.

The researchers selected the gentiobiosyl diester for quantitation, being the largest crocin-based peak present, and it provided linear calibrations over 0.5-10 mg/L.

Crocin was spiked into sausages, sauces, juice and sodas which were extracted on the macroporous column and analysed by HPLC-UV. The recoveries ranged from 81.3-106.2% with low relative standard deviations below 10%, so the macroporous column performed well.

The quantitation limit for sodas and juice was 0.5 mg/kg and those for the sauces and sausages were 5 mg/kg. These limits are entirely adequate for use in China, where the maximum permitted levels of crocin in soft drinks, sausages and sauces are 0.3, 1.5 and 1.5 g/kg, respectively.

The novel extraction and analysis procedure provided a fast and simple method for the analysis of crocin in different types of food, giving good recoveries and quantitation limits. It could be used as the basis for the analysis of other polar natural food additives, with appropriate adjustments to the HPLC conditions for maximum sensitivity.



The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

 

The content of the natural pigment crocin in foods, drinks and sauces has been measured by a fast UPLC method following a novel SPE step using macroporous resin, leading to quantitation limits within national guidelines Gardenia jasminoides Ellis

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