Comments on the paper "the nephrotoxicity risk in rats subjected to heavy muscle activity" by gülsen oner and selma cirrik (2009) journal of sports science and medicine 8, 481-488" and response of the authors.

I have read with great interest the work reported by Gülsen Oner and Selma Cirrik (2009) entitled “The nephrotoxicity risk in rats subjected to heavy muscle activity”. The authors reported that LDL-Cholesterol results of the serum from the rats were calculated by subtracting the HDL-Cholesterol from the Total Cholesterol. I am, however, of the opinion that the formula: LDLCholesterol = Total Cholesterol HDL-Cholesterol should not have been applied by the authors, because the contribution of VLDL-Cholesterol to the Total Cholesterol has not been accounted for. Rats have a high HDLCholesterol level with relatively low VLDL-Cholesterol and very low levels of LDL-Cholesterol (Terpstra et al., 1982). The formula: Total Cholesterol = VLDLCholesterol + LDL-Cholesterol + HDL-Cholesterol, is generally accepted as a convenient approximation, because the LDL fraction is heterogenous and contains intermediate-density lipoprotein (IDL), with a density of 1.006-1.019 kg/L, the main LDL, with a density of 1.0191. 063 kg/L and Lipoprotein (a) (Nuack et al., 2002; Rifai and Warnick, 2006). LDL-Cholesterol can be estimated using both direct and indirect methods. In the indirect methods a number of lipid-related analytes are measured and the results of these analytes are then used in the calculation of the LDL-Cholesterol (Rifai and Warnick, 2006). The two indirect methods of LDL-Cholesterol estimation are betaquantification and the Friedewald equation. In the beta-quantification method, ultracentrifugetion is employed to prepare the LDLCholesterol plus HDLCholesterol fraction. An accurately measured aliquot of the specimen is centrifuged at d 1.006 kg/L for the equivalent of 18 hours at 105,000 x g. The VLDLCholesterol and, if present, Chylomicrons and β-VLDLCholesterol, will accumulate in a floating layer, with the infranant containing primarily LDL-Cholesterol and HDL-Cholesterol plus any IDL-Cholesterol and Lp(a) that may be present. The infranatant fraction can now be reconstituted to the original volume with 0.15 M NaCl after the supernatant VLDLand chylomicron-containing fractions have been quantitatively removed. The cholesterol level can then be estimated in the reconstituted infranate. Using a polyanion-divalent cation reagent that provides HDL-cholesterol measurements equivalent to those obtained with the heparin-manganese chloride method, the HDL-containing fraction should be prepared from the ultracentrifugal infranate. The LDL-cholesterol should then be calculated as follows: [LDL-Cholesterol] = [d >1.006 kg/L Cholesterol] – [HDL-Cholesterol] (Bachorik and Ross, 1995). In the Friedewald equation, Total Cholesterol, Triglyceride, and HDL-Cholesterol are measured and LDL-Cholesterol is calculated from these measurements using the equation: [LDL-Cholesterol] mg/dL = [Total-Cholesterol] mg/dL – [HDL-Cholesterol] mg/dL [Triglyceride] mg/dL /5 (Friedewald et al., 1972). In this formula, the VLDL-cholesterol concentration is estimated by the factor of triglycerides divided by 5. In conclusion, the formula applied by the authors for the calculation of LDL-Cholesterol would only be appropriate if the beta-quantification method of LDLCholesterol estimation was performed. In their study the authors clearly did not perform the beta-quantification method of LDL-Cholesterol estimation and therefore they cannot apply the formula as stated in their article. If the triglycerides were estimated by the authors as part of the rat lipid profile, the Friedewald equation may be applied to estimate LDL-Cholesterol. Alternatively, a direct method for the estimation of LDL-Cholesterol should have been used.