Cholesterol is an essential lipid which is a precursor for many sterol-based compounds and is involved in the regulation of the precise pattering of embryonic structures [1] . Thus, a defeat in its availability during pregnancy has severe consequences to the fetus, interfering in its regular development [2] .

Cholesterol is produced from lanosterol through a complex biosynthetic pathway involving several multienzymatic reactions, including demethylations, isomerization, desaturation, and double-bond reductions. Several polimalformative disorders [3] -[6] have been identified, deriving from enzyme defects in cholesterol biosynthesis [7] [8] . In particular the Smith-Lemli-Opitz syndrome (SLOS), is an autosomal recessive disorder affecting 1:20000 individuals, characterized by a recognizable pattern of minor facial anomalies, limb abnormalities and multiple congenital anomalies including pseudohermaphroditism in males, neonatal hypotonia, mental retardation and failure to thrive [9] -[12] . This disease, caused by a 7-dehydrocholesterol reductase deficiency, is characterized by high blood levels of specific metabolites of postsqualene cholesterol biosynthesis [13] . A clinical suspicion of SLOS is usually confirmed by a marked increase of 7-dehydrocholesterol (7-DHC) in plasma or tissues, accompanied by decreased levels of cholesterol, which is consistent with a 7-dehydrocholesterol reductase deficiency [14] . The determination of serum cholesterol and 7-DHC by GC/MS is the method of choice for SLOS diagnosis [15] , even if other analytical approaches have been described [16] -[18] figure 1. This method was also applied to the prenatal diagnosis for the determination of sterols in amniotic fluid [15] [19] -[26] . It was demonstrated that abnormally elevated amniotic fluid 7-DHC concentrations are an accurate predictor of fetal SLOS [27] [28] . In this paper, we describe the results obtained for the determination of free cholesterol, total cholesterol and its derivatives in 39 amniotic fluids without genetic alterations, considered as control fluids and in 60 amniotic fluids withdrawn from pregnant women, showing a fetal growth retardation potentially related to SLOS, in order to confirm the correlation between this pathology and the sterol levels in amniotic fluid.

The fetal growth retardation were divided into three classes: an overt intra uterine growth retardation (IUGR), a smaller fetus respect to the gestational age (SGA) and cases in which further biomedical studies were necessary (ACC).

In the case of the amniotic fluids related to fetal growth retardations only the quantitative determination of total cholesterol was carried out, in fact it was not possible to determine its free portion because of the small amount of amniotic fluid available after the routine cytogenetic investigations.

In the screening phase the determinations were carried out by means of the GC/FID technique, which allows a fast and accurate quali-quantitative detection of the analytes of interest; the fluids which presented altered concentration values of sterols were further investigated by GC/MS, a technique able to detect compounds structurally related to cholesterol, belonging to its biosynthetic pathway.