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Cholesterol/7-DHC Analysis in Smith-Lemli-Opitz Syndrome
by Bob Wiedemer and Dr. Cary Thrall
Abstract
Baseline separation of underivatized cholesterol and 7-dehydrocholesterol, the marker compounds for Smith-Lemli-Opitz syndrome, is achieved in less than 4 minutes by capillary gas chromatography with an Alltech Heliflex® AT-1ms, (Part No. 15880) 15 m × 0.25 mm × 0.25 μm.
Introduction
Cholesterol is an essential component for human life, involved in many important biochemical mechanisms such as proper cell membrane and nerve function as well as hormone and bile acid synthesis. 7-dehydrocholesterol is a precursor to cholesterol in vivo and builds up into a disease known as Smith-Lemli-Opitz syndrome (SLOS), a hereditary disorder in which a gene encoding 7-dehydrocholesterol reductase is defective, causing the accumulation of noncholesterol sterols such as 7-dehydrocholesterol.1 Smith-Lemli-Opitz syndrome is defined by a constellation of severe birth defects affecting most organ systems. Abnormalities frequently include profound mental retardation, severe failure to thrive, and a high infant-mortality rate.2 SLOS is the second most common autosomal recessive disorder in the North American white population, after cystic fibrosis.
Results
click the image to enlarge
Figure 1: Chromatogram 2677 shows fast analysis of underivatized cholesterol and 7-dehydrolesterol, the marker compounds for Smith-Lemli-Opitz Syndrome.
Cholesterol/7-DHC Analysis (33 ppm each)
1. 5a-Cholestane
2. Cholesterol
3. 7-Dehydrocholesterol
Instrument: HP 6890GC
Column: Heliflex AT-1ms, (Part No. 15880) 15 m × 0.25 mm × 0.25 μm
Temperature: 305 C isothermal
Carrier: Helium at 0.6mL/min (27cm/sec)
Detector: FID 325 C
Inlet: Split at 300 C, split ratio 75:1, split flow 44.9 mL/min
Sample: 1.0 μL injection volume in chloroform (underivitized). |
Figure 1 demonstrates baseline separation of underivatized cholesterol and 7-dehydrocholesterol (peaks 2 and 3 respectively). Peak number 1 is 5a-cholestane, which is commonly used as an internal standard in cholesterol analysis.
Conclusions
The combination of abnormally low plasma cholesterol levels and a high concentration of the cholesterol precursor 7-dehydrocholesterol is an indication of a major block in cholesterol biosynthesis at the step in which the C-7(8) double bond of 7-dehydrocholesterol is reduced, forming cholesterol. The marked elevation of 7-dehydrocholesterol places the defect at a known step in the pathway of cholesterol biosynthesis and serves as a biochemical marker for SLOS.2 This method provides a reliable and rapid method for determining levels of cholesterol and 7-dehydrocholesterol in plasma.
References
1. B. Ruan, W. K. Wilson, J. Pang, N. Gerst, F. D. Pinkerton, J. Tsai, R. I. Kelley, F. G. Whitby, D. M Milewicz, J. Garbern, and G. J. Schroepfer Jr., Journal of Lipid Research 42 (2001): 799-812.
2. G. S. Tint, M. Irons, E. R. Elias, A. K. Batta, R. Frieden, T. S. Chen, and G. Salen, The New England Journal of Medicine 330 (1994): 107-13.
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