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Estimation of Trace Amounts of Chromium (III) in Multi-Vitamin With ANALYTICAL SCIENCES SEPTEMBER 2004, VOL. 20 1321 2004 © The Japan Society for Analytical Chemistry Estimation of Trace Amounts of Chromium(III) in Multi-Vitamin with Multi-Mineral Formulations Jolly M. SANKALIA, Rajashree C. MASHRU,† Mayur G. SANKALIA, Priti P. PARIKH, and Vijay B. SUTARIYA Pharmacy Department, Faculty of Technology & Engineering, The Maharaja Sayajirao University of Baroda, Kalabhavan, Vadodara, Gujarat 390001, India Two new specific, selective, simple and inexpensive spectroscopic methods for estimating a trace amount of chromium (Cr3+) from a multi-vitamin with multi-mineral pharmaceutical formulations were developed. The proposed methods are based on the conversion of Cr3+ to Cr6+ either by oxidation with a nitric acid–perchloric acid mixture (method I) or by fusion with an excess amount of sodium carbonate (method II), followed by the complexation of Cr6+ with 1,5- diphenylcarbazide (DPC) in a mineral acidic solution of pH 1.0 ± 0.5. The pink-colored complex was estimated at 544 nm. Both methods were found to be linear in the range of 0.1 – 0.8 µg/ml with a limit of detection in the range of 0.0123 – 0.0157 µg/ml and a limit of quantitation in the range of 0.0419 – 0.0525 µg/ml. Method I was found to be suitable for estimating Cr3+ species in various formulations, like tablets, capsules and syrups, while method II was found to be suitable for tablets and capsules. Satisfactory recovery from spiked samples of standard Cr3+ suggests no interference of any excipients and diverse ions present in the formulations. The developed methods were compared with AAS by ANOVA, and no significant difference was observed. (Received February 4, 2004; Accepted May 28, 2004) tea and tea leaves, food, dietary products, lung tissues, standard Introduction alloys and biological samples; zeeman AAS from blood and serum; electro thermal AAS (ETAAS) from cocaine samples; Chromium (Cr) can exist in various chemical valence states AAS–flame emission spectrophotometry (AAS-FES) from ranging from Cr2+ to Cr6+, among which more stable chemical ancient Perivian hair; mass spectroscopy (MS) from metal forms are Cr3+ and Cr6+. Cr3+ is relatively non-toxic and is an allergenic substances on skin; inductively coupled plasma–mass essential nutrient in the human diet to maintain effective spectrometry (ICP-MS) from seawater; ICP-atomic emission glucose, lipid and protein metabolism.1 However, Cr6+ is spectroscopy (ICP-AES) from human gallstone; neutron 2– 4– primarily anthropogenic and, as CrO4 or HCrO , can diffuse activation analysis (NAA) from biological specimens, tissue through cell membranes and oxidize biological molecules with surroundings, blood and milk; electron spin resonance (ESR) toxic results.2 Cr6+ has been classified by the International from rat blood; scanning electron microscopy (SEM), Agency for Research on Cancer (IARC) as a Group I human transmission electron microscopy (TEM) and electron probe carcinogen3 and by the US Environmental Protection Agency microanalyses (EPMA) from metal-bearing contaminated soil (EPA) as a Group A inhalation carcinogen.4 and industrial waste; colorimetric methods6 from shrimp feed,7 Chromium is one of the essential trace elements in multi- air,8,9 urine8,10,11 and catgut suture,12 etc. vitamin with multi-mineral pharmaceutical formulations that Not even a single spectroscopic method has been reported for contain only Cr3+ either in the form of chromium chloride estimating Cr3+ from pharmaceutical formulations. Thus the (inorganic source) or chromium picolinate and chromium primary objective of the present work was to develop, optimize polynicotinate (organic source). The methods for estimating and validate sensitive, selective, precise, accurate, cost- Cr3+ from pharmaceutical formulations are atomic absorption effective, and simple spectroscopic methods. spectroscopy (AAS) and graphite furnace AAS (GF-AAS). The proposed methods are based on removing all organic Other methods reported in the literature for estimating Cr3+ are matter of a multi-vitamin with multi-mineral pharmaceutical chemiluminescence analysis, flame AAS, X-ray fluorescence formulations by heating in a muffle furnace13 at 550˚C – 600˚C spectroscopy (XRFS) and colorimetric from air.5 Various for 6 – 7 h. Cr3+ was converted to Cr6+ by oxidation with a nitric methods for estimating Cr6+ and total Cr include flow-injection acid–perchloric acid mixture13 (method I) and by fusion with flame AAS from industrial effluents; turbidimetry; ion excess of sodium carbonate14 (method II), followed by chromatography; anion-exchange chromatography; HPLC with estimating a pink-colored complex in a mineral acidic solution diode array detection (HPLC-DAD) from environmental of pH 1.0 ± 0.5 at 544 nm obtained by the reaction of Cr6+ with matrices; fast protein liquid chromatography (FPLC); thermal 1,5-diphenylcarbazide (DPC), which is considered to be one of lens spectrometry (TLS); X-ray fluorescence; AAS from black the most sensitive and selective reactions for Cr6+ determination.15–19 The developed methods were studied † To whom correspondence should be addressed. concerning the effect of the pH, dye volume, Cr6+:dye ratio, E-mail: [email protected] stability of the colored complex, etc. for methods I and II, while 1322 ANALYTICAL SCIENCES SEPTEMBER 2004, VOL. 20 Fig. 1 Visible spectra of Cr6+: dye complex at 0.1 µg/ml (a), 0.2 µg/ml (b), 0.3 µg/ml (c), 0.4 µg/ml (d), 0.5 µg/ml (e), 0.6 µg/ml (f), 0.7 µg/ml (g) and 0.8 µg/ml (h) concentration levels. Fig. 2 Optimization of the Na2CO3 composition using chromium the Na2CO3 composition, heating time were considered for method II. chloride as a standard. Experimental 0.5 and the volume was made to obtain 5 µg/ml of Cr6+. From this stock solution, suitable aliquots were taken and 0.2 ml of Apparatus DPC dye was added to each, allowed to stand for 5 min for full A Shimadzu UV-VIS spectrophotometer (Model UV-1601) color development and the volume was made up to 10 ml with with matched 1 cm silica cells was used for all spectral and distilled water to obtain the final concentration in the range of absorbance measurements. The glassware used in each 0.1 to 0.8 µg/ml; the absorbance was measured at 544 nm. procedure was soaked overnight in a chromic mixture (K2Cr2O7 + concentrated H2SO4), rinsed thoroughly with double-distilled Preparation of a sample solution (method I) water and dried in dust-free air. Whatman filter paper No. 42 After twenty tablets/capsules were weighed and triturated, a was used to filter solutions of different formulations to separate powder equivalent to 1.0 mg of Cr3+ was subjected to heating in them from the solvent immiscible formulation excipients. a muffle furnace as per the conditions stated under “Preparation of standard solution for calibration curve (method I)”. The ash Reagents was then digested with 30 ml of a nitric acid–water–perchloric Double-distilled water and analytical-reagent grade chemicals acid mixture (5:5:20 v/v/v) on a burner until white fumes of were used. Standard solutions of chromium were prepared by perchloric acid started to appear, became vigorous and then using chromium chloride hexahydrate (CrCl3·6H2O) or ceased. This solution was allowed to cool, and was diluted with 6+ chromium picolinate (C18H12N3O6Cr) salts. A Cr complexing distilled water; the pH was adjusted at 1 ± 0.5 and the volume reagent solution of 1,5-diphenylcarbazide (DPC) (S. D. Fine was made to obtain a final concentration of 5 µg/ml of Cr6+ to Chem Ltd., Mumbai, India) was prepared by dissolving 0.25 g proceed further, as mentioned above. of DPC in 50 ml of acetone, and stored in an amber-colored bottle. This solution was discarded when it became slightly Preparation of a sample solution (method II) colored, and should not be used after 48 h of its preparation; it is After twenty tablets/capsules were weighed and triturated, a always preferable to use it freshly prepared. powder equivalent to 1.0 mg of Cr3+ was mixed with sodium carbonate (Na2CO3) equivalent to 5-times the weight of the Preparation of standard solution for calibration curve (method I) sample taken, mixed/triturated well and then subjected to Chromium salts (chromium chloride hexahydrate or heating in a muffle furnace, as described under “Preparation of chromium picolinate) equivalent to 1.0 mg of Cr3+ were standard solution for calibration curve (method II)”. The ash subjected to heating in a muffle furnace in a silica crucible at was dissolved in about 100 ml of water and boiled on a water- about 550˚C for 6 – 7 h; the ash was then dissolved in 0.2 M bath for about 45 min, cooled and filtered. The pH of the sulfuric acid (pH 1 ± 0.5) in order to obtain a stock solution of 5 filtrate was adjusted at around 1 ± 0.5, and the volume was µg/ml of Cr6+. From this stock solution, suitable aliquots were made to obtain 5 µg/ml of Cr6+ to proceed further, as mentioned taken and 0.2 ml of DPC dye was added to each, allowed to above. stand for 5 min for full color development and volume was made up to 10 ml with distilled water to obtain the final concentration in the range of 0.1 to 0.8 µg/ml; the absorbance Results and Discussion was measured at 544 nm. Absorption spectra Preparation of standard solution for calibration curve (method II) The absorption spectra of a Cr6+-DPC dye complex are In method II, chromium salts (chromium chloride hexahydrate graphically presented in Fig. 1. As can be seen, the complex or chromium picolinate) equivalent to 1.0 mg of Cr3+ along with has its maximum absorbance at 544 nm. sodium carbonate (Na2CO3) equivalent to 5-times its weight were heated in a muffle furnace in a silica crucible at about Effect of the Na2CO3 composition (method II) 550˚C for 6 – 7 h, and then the ash was dissolved in about 100 The Na2CO3 composition in method II is a very critical ml of water, boiled on a water-bath for about 45 min, cooled parameter, which was investigated for optimum conditions.
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