Sphingomyelin solubility

FIGURE 10.12 The mole ratio of carotenoid/phospholipid and carotenoid/total lipid (phospholipid + cholesterol) in raft domain (detergent-resistant membrane, DRM) and bulk domain (detergent-soluble membrane, DSM) isolated from membranes made of raft-forming mixture (equimolar ternary mixture of dioleoyl-PC (DOPC)/sphingomyelin/cholesterol) with 1 mol% lutein (LUT), zeaxanthin (ZEA), P-cryptoxanthin (P-CXT), or P-carotene (P-CAR). 

The experimental procedure below describes the uptake of ciprofloxacin into sphingomyelin (SPM)/Chol LUVs. Drug delivery vehicles prepared from SPM/Chol often exhibit greater efficacy than those prepared from DSPC/Chol (13). Included is a description of the Bligh-Dyer extraction procedure (78), which involves partitioning the lipid and water-soluble drug into organic solvent and aqueous layers, respectively. This is necessary because lipid interferes with the ciprofloxacin assay. 

Hormonal lipids. We have already considered a number of hormones that are not water-soluble but may have to be transported by carrier proteins to their sites of action. These include retinoic acid (Box 22-A), metabolites of vitamin D (Box 22-C), and the plateletactivating factor (Box 8-A). The last functions in the brain311 as well as in blood. Hormonal lipids also include the prostaglandins (Fig. 21-7), leukotrienes, and lipoxins (Fig. 21-8). These are products of the eicosenoid cascade or network, which is activated by receptors linked to phospholipase C (Fig. 11-9). Cera-mide formed by hydrolysis of sphingomyelin initiates... 

Some commercial preparations of purified sphingomyehnase from B. cereus may be contaminated with phospholipase C. This is irrelevant when the substrate consists of pure sphingomyelin, but when the LUV contain glycerophospholipids, e.g. phosphatidylcholine, contamination is serious because both enzymes will give off the water-soluble end-product phosphorylcholine. In such cases, the assays include o-phenanthroline, a specific inhibitor of phospholipase C that is innocuous for SMase [92-94]. 

Antonis (A3) has estimated phospholipids by a procedure for determining fatty acids. This technique requires a total serum extract and a phospholipid-free extract for the measurement of both total and free fatty acids, the difference between them being a measure of the phospholipid content. Free fatty acids (A4) are determined on a phospholipid-free extract by a procedure based on partitioning the fatty acids as copper soaps into chloroform, and subsequent photometric determination of the copper with diethyldithiocarbamate. Phospholipids, as well as the free fatty acids present in the total lipid-extract, are measured by the same method, since they also form a complex with copper that is soluble in chloroform. A criticism of this technique is that equal response is not given by dipalmitoyl lecithin, dipalmitoyl cephalin, or beef brain sphingomyelin. 

CjH,4N0, Mr 104.17 (ion) the hydroxide, CjHijNOj, Mr 121.18, is a highly hygroscopic syrup that crystallizes only with difficulty, is very soluble in water and alcohols but not soluble in ether. It has a strongly basic reaction and absorbs carbon dioxide. C. is weakly toxic, LD50 400 mg/kg (rat i.p.), 3400 mg/kg (rat, hydrochloride p. o.), 6640 mg/kg (cat p.o.). The chloride, C5H,4C1N0, Mr 139.63, forms hygroscopic crystals. C. occurs both in the free form and bound in lipids (e.g., sphingomyelines and lecithines) in bile, brain, egg yolk... 

Sphingomyelin is soluble in benzene, hot ethanol and ethylacetate insoluble in acetone and ether. An approved method of the isolation and purification by chromatographic separation on silicic acid has been reported (Hanahan 1961). 

Earlier methods employed for the resolution of the phospholipid fraction of tissue depend primarily on the different solubility characteristics of its components. Kirk (46) extracted the precipitate obtained from the alcohol-ether extract of tissues by treatment with acetone and magnesium chloride with moist ether which dissolves only lecithin and cephalin and leaves sphingomyelin as an insoluble residue. Determination of choline in the ether-soluble fraction after the hydrolysis was a measure of the lecithin present and cephalin was calculated by difference from the phosphorus determinations. 

The relative resistance of sphingomyelin to alkaline hydrolysis has been used to some extent as a method for estimating the sphingomyelin content of tissues (75) and of lecithins and cephalin (37,45,103). Lecithin and cephalin are quantitatively hydrolyzed into acid-soluble phosphorus compounds by normal KOH at 37 C. within 24 hours. Thus, the difference between the total phospholipid phosphorus and the saponifiable phosphorus represents the amount of sphingomyelin phosphorus. 

Hack (37) extended this method to the estimation of sphingomyelin, lecithin, and cephalin in blood and plasma of humans. Under the conditions described for saponification and subsequent acidification, choline was quantitatively liberated from lecithin. The determination of choline and acid-soluble phosphorus after saponification provides a basis for the... 

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