Carbohydrates ammonia, chloroform

Phosphorus, fatty acids, carbohydrates, glycerol, and amino acids were analyzed by the method described in our previous paper [8] and references cited therein. SDS-PAGE [8], TLC [9], HPLC [9], determination of phos-phomonoester [8], reducing sugar analysis [13], methylation analysis [14], and hexose analysis [15] were performed as described in the respective literature. Two dimensional TLC was performed on silica-gel plate (Merck Silicagel 60 F254 No. 5715) using the solvent systems, chloroform-methanol-acetic acid (65/10/1, v/v/v) for the first development and chloroform-methanol-25% ammonia solution (65/10/1) for the second. 

Fig. 150. Thin-layer chromatogram of phospholipids and other polar lipids of bovine milk [159 a]. 1 carbohydrate (lactose) and protein, 2 sphingomyelin, 3 phosphatidyl choline, 4 phosphatidyl serine, 5 phosphatidyl inositol, 6 phosphatidyl ethanolamine, 7 cerebroside dihexoside ( ), 8 cerebroside mono-hexoside ( ), P fatty acids, 10 neutral lipids. Adsorbent Silica gel HR. Solvents I, chloroform-methanol-water-28% aqu. ammonia (130 + 70+8 + 0.5) II, chloroform-acetone-methanol-acetic acid-water (50 + 20 + 10 + 10 + 5). Time 40 min in each direction. Indicator charring with chromic sulphuric acid solution...

Figure 7 Two-dimensional TLC separation of serum lipids. The TLC plate was developed in the first direction with chloroform-methanol-ammonia (65 25 5) and in the second direction with chloroform-acetonc-methanol-acetic acid-water (30 40 10 10 0.5). Abbreviations are as follows LPC, lysophosphatidylcholine PA, phospha-tidic acid PC, phosphatidylcholine PE, phosphatidylethanolamine PG, phosphatidylglycerol PI, phosphatidyl-inositol PS, phosphatidylserine DPG, diphosphatidylglyccrol (cardiolipin) Sph, sphingomyelin. Redrawn with permission of Supelco, Inc., Bellefonte, Pa., from Supelco Handbook of Lipids and Selected Carbohydrates, 7th ed 1981, p. 48.

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