Table 1.
Brand name, type of formulation, salt form and amount claimed on the label of glucosamine supplements available in Australia and India.
Fig 1.
Influence of various compositions of acetonitrile on separation of glucosamine hydrochloride.
Experimental conditions: ZIC-HILIC column (150, 4.6 mm), 200 Å, 5 μm; mobile phase containing various compositions of acetonitrile (as specified in chromatograms) and 100 mM ammonium acetate; column temperature was 25°C; detection with Corona-CAD; flow rate-0.5 mL/min; injection volume 5 μL.
Fig 2.
Influence of flow rate and temperature on backpressure and retention time of glucosamine hydrochloride.
A) Retention time of glucosamine and backpressure of the column (Inset-1) when flow rate was 0.5 mL/min and column temperature was 25°C; B) Retention time of glucosamine when flow rate was 0.4 mL/min and column temperature was 25°C; C) Retention time of glucosamine and backpressure of the column (Inset-2) when flow rate was 0.3 mL/min and column temperature was 25°C; D) Retention time of glucosamine and backpressure of the column (Inset-3) when flow rate was 0.3 mL/min and column temperature was 40°C; Experimental conditions: ZIC-HILIC column (150, 4.6 mm), 200 Å, 5 μm; mobile phase containing 65% acetonitrile and 100 mM ammonium acetate; column temperature (as specified above); detection with Corona-CAD; flow rate- (as specified above); injection volume 5 μL.
Fig 3.
Influence of various composition of acetonitrile and ammonium acetate on separation of glucosamine.
A) Separation of glucosamine sulphate when mobile phase contain 65% acetonitrile and 100 mM ammonium acetate; B) Separation of sodium sulphate when mobile phase contain 65% acetonitrile and 100 mM ammonium; C) Separation of glucosamine sulphate when mobile phase contain 60% acetonitrile and 100 mM ammonium acetate; Inset-1: Separation of glucosamine sulphate when mobile phase contain 50% acetonitrile and 100 mM ammonium acetate; D) Separation of glucosamine sulphate when mobile phase contained 50% acetonitrile and 100mM ammonium acetate; E) Separation of glucosamine sulphate when mobile phase contain 60% acetonitrile and 85 mM ammonium acetate; F) Separation of glucosamine sulphate when mobile phase contain 60% acetonitrile and 80 mM ammonium acetate; G) Separation of glucosamine hydrochloride when mobile phase contain 60% acetonitrile and 85 mM ammonium acetate; Experimental conditions: ZIC-HILIC column (150, 4.6 mm), 200 Å, 5 μm; mobile phase containing various composition of acetonitrile (as specified above) and ammonium acetate (as specified above); column temperature 40°C; detection with Corona-CAD; flow rate 0.3 mL/minute; injection volume 5 μL.
Fig 4.
Calibration curve with linear regression equations and correlation coefficients (r2) generated by plotting peak areas versus glucosamine concentrations (10–200 μg/mL) of A) Glucosamine hydrochloride reference standard; B) Glucosamine sulphate reference standard.
Table 2.
Mean inter- and intra-day accuracy, precision, reproducibility and linearity for glucosamine hydrochloride and glucosamine sulphate.
Table 3.
Results of system suitability.
Table 4.
Mean percentage recovery of glucosamine from spiked sample solutions after filtration.
Table 5.
Stability of glucosamine stored at 4°C and room temperature up to 48 hours.
Table 6.
Labelled and average amount of glucosamine observed in commercial preparations.
Table 7.
Comparison of currently available methods for quantification of glucosamine with newly developed method.
Fig 5.
Application of newly-developed method for separation of glucosamine present in supplements.
A) Blank (water); B) Glucosamine hydrochloride reference standard; C) Glucosamine sulphate reference standard; D) Chondroitin sulphate reference standard; E) Product-b containing glucosamine 1500 mg of glucosamine hydrochloride with chondroitin (100 mg) F) Product-d containing 750 mg of glucosamine sulphate with chondroitin (421 mg); G) Product-f containing glucosamine sulphate-sodium chloride complex (1.88 mg) equivalent to 1500 mg of glucosamine sulphate with 500 mg Zingiber officinale or ginger equivalent to 2.5 mg gingerols; H) Product-h containing 750 mg of glucosamine sulphate-potassium chloride complex, equivalent to 446 mg of glucosamine with 250 mg of methylsulphonylmethane and 50 mg of diacerin; Experimental conditions: ZIC-HILIC column (150, 4.6 mm), 200 Å, 5 μm; mobile phase containing 60% acetonitrile and 85 mM ammonium acetate; column temperature 40°C; detection with Corona-CAD; flow rate 0.3 mL/min; injection volume 5 μL.