Absolute Quantification of Carminic Acid in Cochineal Extract by Quantitative NMR

Abstract

A quantitative NMR (qNMR) method was applied for the determination of carminic acid. Carminic acid is the main component in cochineal dye that is widely used as a natural food colorant. Since several manufacturers only provide reagent-grade carminic acid, there is no reference material of established purity. To improve the reliability of analytical data, we are developing quantitative nuclear magnetic resonance (qNMR), based on the fact that the intensity of a given NMR resonance is directly proportional to the molar amount of that nucleus in the sample. The purities and contents of carminic acid were calculated from the ratio of the signal intensities of an aromatic proton on carminic acid to nine protons of three methyl groups on DSS-d 6 used as the internal standard. The concentration of DSS-d 6 itself was corrected using potassium hydrogen phthalate, which is a certified reference material (CRM). The purities of the reagents and the contents of carminic acid in cochineal dye products were determined with SI-traceability as 25.3ῌ92.9 and 4.6ῌ30.5 based on the crystalline formula, carminic acid potassium salt trihydrate, which has been confirmed by X-ray analysis. The qNMR method does not require a reference compound, and is rapid and simple, with an overall analysis time of only 10 min. Our approach thus represents an absolute quantitation method with SI-traceability that should be readily applicable to analysis and quality control of any natural product.

Karminsäure

“…Die NMR-Spektren wurden in D 2 O nach Zugabe eines Tropfens CF 3 COOH bei +50 °C aufgenommen, um die Tautomeren-Gleichgewichte zu beschleunigen, da diese vor allem die 13 Säurekatalyse erhöht die Geschwindigkeit der Einstellung des Tautomeren-Gleichgewichtes. Dies führt zum schnellen Austausch und auf der 100 MHz 13 C-NMR-Zeitskala werden Signale mit schmaler Linienbreite beobachtet.…”

“…Ein Beleg dafür ist die jüngste NMR-Arbeit, die eine interne Kalibrierung nutztdie aber eben nicht auf Karminsäure beruht [13]. Ein Beleg dafür ist die jüngste NMR-Arbeit, die eine interne Kalibrierung nutztdie aber eben nicht auf Karminsäure beruht [13].…”

Application of 1H-NMR spectroscopy to validation of berberine alkaloid reagents and to chemical evaluation of Coptidis Rhizoma

“…Determination of HMD concentration in qHNMR reference stock solution Preparation of qHNMR reference stock solution was essentially performed as described by Tahara et al [11] and Sugimoto et al [12]. In brief, HMD (10 mg) was dissolved in methanol-d 4 (50 ml) to make qHNMR reference stock solution, and the accurate concentration of HMD was determined by measuring 1 H-NMR spectrum with PHP as an internal standard.…”

“…Proton-specific quantitative NMR (qHNMR) has an enormous potential for quantitative analysis of natural products [7,8]. Two of the unique features of the qHNMR method are: (1) it leads to absolute quantification of the organic compounds by comparing the signal integral of the particular compound with that of an appropriate certified reference material (CRM) [9], and (2) the obtained values are theoretically traceable to the International System of Units (SI) by using CRMs or other compounds whose concentrations are SI-traceably determined as a qHNMR reference material [10][11][12]. In addition, the advantage of qHNMR over HPLC is that it is rapid and non-invasive, and in most cases does not require laborious sample pre-clean steps.…”

Development of a Novel Method for Quantifying Quassin and Neoquassin in Jamaica Quassia Extracts Using the Molar Absorption Coefficient Ratio

“…Each integral value is the mean SD of six independent determinations. Solvent, methanol-d 4 . The molar values of 4HBA in each test solution were calculated from the 7.86 derived from H-2 and H-6 and 6.80 ppm signals derived from H-3 and H-5 and were set at 1.00.…”