Certain impurities in the color additives drug and cosmetic (D&C) Red No. 21 (R21), D&C Red No. 22 (R22), and their lakes are limited to levels specified in the Code of Federal Regulations (CFR) and ...are quantified by the U.S. Food and Drug Administration in batches of these color additives submitted for certification. Currently, a lengthy and tedious method based on gravity flow elution column chromatography is used to quantify the following CFR-specified impurities: the intermediate, phthalic acid (PhthAc); the manufacturing by-products, 2-(3',5'-dibromo-2',4'-dihydroxybenzoyl)benzoic acid (Br2BBA); and brominated resorcinol. "Brominated resorcinol" implies the sum of all possible brominated resorcinols, but the current work focused on 2,4,6-tribromoresorcinol (Br3R) as the most probable side-reaction product.
An improved method was needed to quantify PhthAc, Br2BBA, and Br3R in R21, R22, and their lakes.
A rapid ultra-HPLC (UHPLC) method was developed to replace the gravity flow method for quantitative determination of PhthAc, Br2BBA, and Br3R.
PhthAc, Br2BBA, and Br3R were quantified by using five-point calibration curves with data point ranges of 0.11-1.55, 0.06-0.77, and 0.04-0.61% by weight, respectively. LODs for the analytes ranged from 0.01 to 0.03%. Recoveries of the analytes ranged from 90.6 to 99.9%.
The UHPLC method is accurate and significantly more rapid than the gravity -flow method, requiring approximately 7 min as compared with 6 h to detect PhthAc, Br2BBA, and Br3R in one sample.
A rapid UHPLC method was developed to determine CFR-specified impurities in the color additives D&C Red Nos. 21 and 22 and their lakes.
Background: Certain impurities in the color additives drug and cosmetic (D&C) Red No. 21 (R21), D&C Red No. 22 (R22), and their lakes are limited to levels specified in the Code of Federal ...Regulations (CFR) and are quantified by the U.S. Food and Drug Administration in batches of these color additives submitted for certification. Currently, a lengthy and tedious method based on gravity flow elution column chromatography is used to quantify the following CFR-specified impurities: the intermediate, phthalic acid (PhthAc); the manufacturing byproducts, 2-(3',5'-dibromo-2',4'-dihydroxybenzoyl) benzoic acid (Br2BBA); and brominated resorcinol. "Brominated resorcinol" implies the sum of all possible brominated resorcinols, but the current work focused on 2,4,6-tribromoresorcinol (Br3R) as the most probable side-reaction product. Objective: An improved method was needed to quantify PhthAc, Br2BBA, and Br3R in R21, R22, and their lakes. Methods: A rapid ultra-HPLC (UHPLC) method was developed to replace the gravity flow method for quantitative determination of PhthAc, Br2BBA, and Br3R. Results: PhthAc, Br2BBA, and Br3R were quantified by using five-point calibration curves with data point ranges of 0.11-1.55, 0.06-0.77, and 0.04-0.61% by weight, respectively. LODs for the analytes ranged from 0.01 to 0.03%. Recoveries of the analytes ranged from 90.6 to 99.9%. Conclusions: The UHPLC method is accurate and significantly more rapid than the gravity -flow method, requiring approximately 7 min as compared with 6 h to detect PhthAc, Br2BBA, and Br3R in one sample. Highlights: A rapid UHPLC method was developed to determine CFR-specified impurities in the color additives D&C Red Nos. 21 and 22 and their lakes.
Asher Mandelbaum (1934-2020) Weisz, Adrian; Markey, Sanford P
Journal of the American Society for Mass Spectrometry,
2021-Jan-06, 2021-01-06, Volume:
32, Issue:
1
Journal Article
The performance of three types of high-speed counter-current chromatography (HSCCC) instruments was assessed for their use in separating components in hydrophilic and hydrophobic dye mixtures. The ...HSCCC instruments compared were: (i) a J-type coil planet centrifuge (CPC) system with a conventional multilayer-coil column, (ii) a J-type CPC system with a spiral-tube assembly-coil column, and (iii) a cross-axis CPC system with a multilayer-coil column. The hydrophilic dye mixture consisted of a sample of FD&C Blue No. 2 that contained mainly two isomeric components, 5,5′- and 5,7′-disulfonated indigo, in the ratio of ∼7:1. The hydrophobic dye mixture consisted of a sample of D&C Red No. 17 (mainly Sudan III) and Sudan II in the ratio of ∼4:1. The two-phase solvent systems used for these separations were 1-butanol/1.3M HCl and hexane/acetonitrile. Each of the three instruments was used in two experiments for the hydrophilic dye mixture and two for the hydrophobic dye mixture, for a total of 12 experiments. In one set of experiments, the lower phase was used as the mobile phase, and in the second set of experiments, the upper phase was used as the mobile phase. The results suggest that: (a) use of a J-type instrument with either a multilayer-coil column or a spiral-tube assembly column, applying the lower phase as the mobile phase, is preferable for separating the hydrophilic components of FD&C Blue No. 2; and (b) use of a J-type instrument with multilayer-coil column, while applying either the upper phase or the lower phase as the mobile phase, is preferable for separating the hydrophobic dye mixture of D&C Red No. 17 and Sudan II.
The anthraquinone anionic dye Acid Violet 43 (AV43, Colour Index No. 60730) is certifiable in the United States as Ext. D&C Violet No. 2 (EV2) for use as a color additive in externally applied ...cosmetics. The dye is also permitted in cosmetics in the European Union, Japan, and other countries. To be certified in the U.S., the dye may not contain more than 1% subsidiary colors. The present work was aimed at identifying an impurity, presumed to be a subsidiary color, found in batches of AV43 submitted for certification as EV2. The concentration level of the impurity in a sample of EV2 was enriched from ∼5.5% to ∼33%, and the enriched sample, 260 mg, was subjected to high-speed countercurrent chromatography (HSCCC). The resulting 50.7 mg of the isolated impurity was sufficient for its identification by spectroscopic methods as the sulfonated phthaloyl-carbazole derivative 6-hydroxy-9-methyl-5H-naphtho2,3-acarbazole-5,13(12H)-dione-11-sulfonic acid (AV43C). That previously unreported compound was synthesized (0.5 g) for use as a reference material to quantify the impurity in batches of EV2. Test portions of 29 batches, produced by 11 manufacturers from six countries, were analyzed by HPLC and UHPLC. The levels of AV43C ranged from <0.05% to 5.66% across batches. Carbazolization through an intramolecular Scholl-type reaction in the sulfonation step of the manufacturing process is suggested as the likely source of AV43C formation. Control of the conditions that enable carbazolization is proposed as a way to reduce or eliminate contamination by AV43C.
•Anthraquinone dye Acid Violet 43 impurity isolated by countercurrent-chromatography.•Impurity identified as novel compound, a sulfonated phthaloyl-carbazole (AV43C).•AV43C synthesized for use as a reference material.•AV43C quantified in 29 batches of the cosmetic dye Ext. Violet No. 2.•Source of AV43C contamination during dye manufacture proposed.
•Counter-current chromatography for separating impurity in Sudan-type color additive.•A hydrophobic biphasic solvent system was used for the separation.•Impurity identified as ...1,3-bis(4-phenylazophenyl)triazene (PAPT) and synthesized.•Synthesized impurity used as reference to quantify PAPT in D&C Red No. 17/CI 26100
The present work describes the application of high-speed counter-current chromatography to the preparative separation of a previously unreported impurity in the color additive D&C Red No. 17 (R17, Colour Index No. 26100, Sudan III). Due to the hydrophobic nature of the impurity, a hydrophobic two-phase solvent system (hexane-ethanol-water, 5:4:1) was used for its separation. The separated impurity was chemically characterized by spectroscopic methods as a disazo triazene, 1,3-bis(4-phenylazophenyl)triazene (PAPT). This impurity was synthesized and used as a reference material to quantify it in 15 batches of the color additive produced by various domestic and foreign manufacturers and certified by the U.S. Food and Drug Administration (FDA). Analysis of test portions by high-performance liquid chromatography showed a range of PAPT levels, from “not detected” (<0.006%) to 0.70%, across batches. The variability suggests that contamination by PAPT can be decreased or eliminated through manufacturing modifications. A chemical pathway for PAPT formation and an associated adjustment to minimize it during the process of manufacturing R17 are proposed.
•Spiral HSCCC with intermittently pressed tubing for small-molecule separation•Spiral HSCCC of moderately hydrophobic impurity in thioindigoid color additive•Impurity identified as ...7-chloro-5-methyl-2H-1,4-benzothiazin-3(4H)-one (BTZ)•Separated impurity used to quantify BTZ in D&C Red No. 30/CI 73360 by HPLC
An impurity in the color additives D&C Red No. 30 (R30) and D&C Red No. 30 lakes (R30L) was newly identified and characterized as 7-chloro-5-methyl-2H-1,4-benzothiazin-3(4H)-one (BTZ), and its extent and level in certified batches of these color additives was determined. BTZ was extracted from the dye with ethanol, resulting in a crude extract enriched to a concentration of over 60%. BTZ was then separated from a portion of the enriched extract by high-speed counter-current chromatography using a spiral-tube assembly column with intermittently pressed tubing of 60 ml capacity. It was the first reported use of such a column to separate a small, moderately hydrophobic compound. The two-phase solvent system was also moderately hydrophobic, consisting of hexane-ethyl acetate-methanol-water (5:2:5:2), and the retention of the organic stationary phase measured after the separation was 83.3%. The separation yielded BTZ of two purity grades, the higher of which (~95.5%) was used as a standard to quantify the impurity in 37 batches of R30 and R30L using an HPLC method developed and validated for that purpose. Analyses revealed a wide range of BTZ levels across batches, <0.05 - 0.84%, and suggested that BTZ contamination could be reduced by appropriate adjustments in the manufacturing process. An explanation of the likely source of BTZ – as a side-reaction product in a particular step of the manufacturing process – was also presented.
► Minor component 1,3,6-pyrenetrisulfonate (P3S) of D&C Green No. 8 (G8) separated. ► pH-zone-refining CCC separation of 20.3
g of G8 using dodecylamine as ion-exchanger. ► Major component ...8-hydroxy-1,3,6-pyrenetrisulfonate (HP3S) also separated. ► Obtained 0.58
g P3S and 12.1
g HP3S of ∼99% purity. ► Identification and characterization by
1H NMR, HR-MS, UV–vis, and HPLC.
In developing analytical methods for batch certification of the color additive D&C Green No. 8 (G8), the U.S. Food and Drug Administration needed the trisodium salt of 1,3,6-pyrenetrisulfonic acid (P3S) for use as a reference material. Since P3S was not commercially available, preparative quantities of it were separated from portions of a sample of G8 that contained ∼3.5% P3S. The separations were performed by pH-zone-refining counter-current chromatography using dodecylamine (DA) as the hydrophobic counterion. The added DA enabled partitioning of the polysulfonated components into the organic stationary phase of the two-phase solvent system used, 1-butanol–water (1:1). Thus, a typical separation that involved 20.3
g of G8, using sulfuric acid as the retainer acid and 20% DA in the stationary phase and 0.1
M sodium hydroxide as the mobile phase, resulted in ∼0.58
g of P3S of greater than 99% purity. The identification and characterization of the separated P3S were performed by elemental analyses, proton nuclear magnetic resonance, high-resolution mass spectrometry, ultra-violet spectra, and high-performance liquid chromatography.
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