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Optimized acetyl esterase enzyme production conditions using Aspergillus niger ATCC 10864 in 14-L fermentation jars were determined to be 33 degrees C, 1.5 vvm aeration, and 300 rpm agitation without pH control. The acetyl esterase was purified by precipitation in 60-80% saturation in ammonium sulfate. The pellet was applied directly to a Pharmacia high-load Phenyl Sepharose column for hydrophobic interaction chromatography and purified to homogeneity in two steps. Stability and kinetic characteristics of the acetyl esterase were determined over a pH range of 4.0-7.5 and from 4 to 45 degrees C. At temperatures25 degrees C, stability was superior at pH values5.0. The temperature activity optimum was 35 degrees C, and the pH optimum was 7.0. The Vmax was determined to be 46,700 U/mg protein, and the Km was 0.023M p-nitrophenyl acetate at pH 6.5 in 0.2M phosphate buffer at 35 degrees C. The mol wt of the enzyme was 35,000 dalton by size-exclusion chromatography and SDS gel electrophoresis. The N-terminal amino acid sequence and the glycosylation composition were also determined.

The morphology and size expected for impact craters on small icy bodies are presented. Such bodies are for example minor satellites of the outer planets (some of which are ice covered) or comet nuclei. The differences between the impact craters that result on such bodies, compared to those on more traditional ice targets (effectively large, well consolidated, semi-infinite ice surfaces) is discussed with particular reference to the impact on a comet nucleus expected in the Deep Impact space mission. Finally extrapolation of laboratory scale experiments is carried out to try and quantify crater size and shape for impacts on small, porous bodies. It is found that given our present knowledge, simple scaling with impact energy produces a result compatible to scaling via more sophisticated methods. The handling of the influence of the detailed composition of the ice target (porosity, volatile content, silicate content etc.) is less certain.

The influence of a purified rat diet containing 20 or 33% of the saturated medium chain-length triglycerides (MCT) with and without linoleic acid supplements on growth, caloric requirements for weight maintenance and weight increase, fertility, lactation performance, and serum cholesterol levels was compared with that of similar diets containing lard, coconut oil, or no fat.

In view of the interest in the biological properties of products of fat oxidation, lauroyl and myristoyl peroxides were fed and their nutritional effects compared with those of autoxidized cottonseed oil, which had been analyzed for its composition. Purified diets containing no fat +2% of linoleic acid, 5% lauroyl or myristol peroxide, or 10% oxidized cottonseed oil were fed to weanling male albino rats for 73 to 98 days, after which they were killed and their organs weighed. Their sera, livers, and testicular fat bodies were used for lipid analysis. With peroxides, growth was significantly depressed but not as much as when oxidized cottonseed oil was fed. Analysis of organ weight data showed that peroxides and oxidized cottonseed oil differed in their effects. Animals fed the latter had significantly heavier livers, kidneys, and hearts. The rats fed peroxides were also different from those fed the fat-free diet and those kept on restricted food intake. Gas chromatographic analysis of the testicular fat bodies revealed a greater deposition of oleate in the animals fed oxidized cottonseed oil, which suggested that these animals were unable to use the oxidized oil for depot fat formation. In the anials fed lauroyl and myristoyl peroxides, appreciable amounts of laurate and myristate, respectively, were found. The composition of the liver neutral fat of the animals fed peroxides was similar to that of the animals fed the low-fat diet +2%, linoleic acid. Serum cholesterol levels of the rats fed peroxides were about 70 mg. %, and of those fed oxidized cottonseed oil, 53 mg. %. The groups fed peroxides also had significantly higher liver cholesterol levels, which suggests that peroxides and oxidized cottonseed oil differed in their effects on cholesterol formation and transport.

Lard and cottonseed oil which had been autoxidized at about 100°C. for 210 hrs. were fractionated by a technique involving molecular distillation, conversion to ethyl esters, urea-complex formation, and redistillation. The ethyl esters were then fed to rats for three weeks at a level of 8% in a purified diet. Growth, water intake, organ weights, total liver lipids, and serum and liver cholesterol levels were determined. Groups fed 8% lard served as controls. Growth was severely depressed by the residue fractions of the urea-complex- and noncomplex-forming portions of the original molecular distillates. Of the three fractions from the original molecular distillation residues, the dimeric and polymeric fractions were the most active. The relative liver and kidney weights were usually increased by feeding the growth-depressing fractions. However there were a number of exceptions indicating more specific effects from some of the fractions. Water intakes were lower with the fractions derived from the original molecular distillates than with those from the original molecular distillation residues. Testicular fat body weights suggested that feeding of autoxidized CSO fractions led to less neutral fat deposition than feeding of corresponding autoxidized lard fractions. Dry weight of the enlarged livers was higher, and the total lipid lower than of the control livers. Total liver cholesterol was higher in animals with smaller livers, but there was no difference in the cholesterol content of the total liver lipids. Serum cholesterol levels were lower in animals with large livers. Further study of those fractions having pharmacological properties is suggested.

1. Effects of medium-chain (C6–C12) saturated triglycerides (MCT) and long-chain (C14–C18) saturated triglycerides (LCT) with and without linoleic acid (LA) supplementation were studied on rats fed purified diets 2. With 2% linoleic acid rats fed MCT and LCT grew somewhat better than those on a low-fat diet with the same supplement. Without linoleic acid those fed MCT grew better, and those fed LCT grew worse than those on the corresponding low-fat diet. MCT seemed to decrease, and LCT to increase linoleic acid requirements. 3. In survival studies 14 out of 18 rats fed 20% MCT were alive after 2 years; of their controls fed 20% lard, 10 out of 19 survived. 4. Reproduction studies in females gave equally poor results on unsupplemented low-fat, MCT, and LCT diets regarding implantation, birth weight, and survival rate. The weaning weights of the young on MCT were however the highest. With 2% LA weaning weights were equally high with LCT and MCT but lower with low-fat diet. 5. In animals fed low-fat diets not supplemented with LA, low serum cholesterol was associated with high liver cholesterol. With MCT, serum values were higher and liver values were significantly lower. With unsupplemented LCT, serum and liver values were high. When the three diets were supplemented with 2% LA, there were no longer any differences in the serum levels and in the liver levels. Whether ar not the presence of some oleate in the MCT and LCT influenced the cholesterol results is not certain. 6. The differences in the effects of MCT and LCT are discussed.

Fresh and oxidized cotton seed oil (CO) olive oil (OO), chicken fat (CF) and beef fat (BF) were fed to male weanling rats for 33 to 108 weeks. Groups fed oxidized fats except OO showed a higher death rate than those fed the corresponding fresh fats. Groups fed oxidized CO and BF had the highest death rate. Histological studies of animals dying from natural causes showed more pronounced cardiac lesions in the animals fed oxidized CO. Serum, liver and brain cholesterol levels were not influenced by oxidized fats. Fatty acid composition of depot fats and of heart and liver lipids did not show significant differences between groups fed fresh and the corresponding oxidized fats.

The biological effects of MCT (medium-chain triglycerides) differed from those of conventional fats, and certain of these suggested examination of the fatty acid composition and triglyceride structure of the adipose tissues and other organs of rats which were fed medium- or long-chain triglyceride mixtures with two levels of linoleic acid. Fatty acid compositions and TG patterns varied with the tissue, with the dietary fat, and with the level of linoleate in the diet. Increasing the latter resulted in the deposition of more linoleate at the expense of oleate and in the deposition of more of the dietary fatty acids. The percentage of completely saturated triglyc-erides (S3), in adipose tissue with the higher linoleate supplement in the rats fed MCT, decreased in those fed LCT and was unchanged in the controls fed a fat-free diet. The increased deposition of the dietary fatty acids was more pronounced in the S3 band.

The absorption of saturated long chain triglycerides across the jejunal epithelium of the rat has been studied by electron microscope. The absence of lipid droplets in the microvilli, the terminal web and the absence of phagocytosis of lipid droplets at the cell surface is consistent with the concept that the majority of the triglyceride is hydrolyzed in the intestinal lumen, is transferred across the cell surface as fatty acids, glycerol, and monoglyceride. Visible lipid first appears in the channels of the endoplasmic reticulum, probably as a triglyceride resynthesized from the absorbed fatty acids. As with the saturated medium chain acids, the long chain acids also produced a negative image in the electron micrographs due to the absence of the double bonds necessary to reduce osmium. The chylomicrons developed by the intestinal epithelial cells are ellipsoidal or elongated in comparison to those formed with MCT and unsaturated fatty acids, which are spherical. These unstained chylomicrons were visible only because of the formation of a tiny “membrane” immediately investing each chylomicron. These “membranes” have been interpreted as lipoprotein or phospholipid acquired by the chylomicron in its transit through the epithelial cell. One of the major differences in the absorption of LCT in comparison with other lipids is the development of large cleft-like deposits of this lipid within the cytoplasm of the epithelial cells. This ultimately causes damage to the cell as evidenced by the destruction of mitochondria and the development of “myelin” forms within the cytoplasm of some of the cells.

Two matching groups of weanling male albino rats were fed a purified diet containing 5% dihydroxystearic acid. All animals survived until they were killed; the first group was sacrificed at age 150 days and the second at 305 days. Fecal fat analyses showed that at least 80% of the ingested dihydroxystearic acid was not exereted. Growth was depressed during the first four weeks; this was followed by a period of rapid weight inerease. The weight of the epididymal fat pads indicated depression of neutral fat deposition. Livers and adrenals were relatively heavy in the group killed earlier and essentially normal in those killed later. This and the eventual weight increase showed that the rats adapted themselves to the intake of the material. Gas liquid chromatography carried out on the methyl ethers of the epididymal fat did not reveal the presence of dihydroxystearic acid. The fatty acid composition of the lipids of serum, kidney, liver and epididymal fat was determined on the methyl esters of the lipid extracts.

RECENTLY it has been shown that a diet which is low in vitamin E and which contains 5 per cent of oxidized lipid is capable of producing the generalized Shwartzman reaction in pregnant rats1,2. This reaction is characterized by an episode of disseminated intravascular coagulation and, in the past, has only been produced experimentally by intravenous injection of bacterial endotoxin. The fact that the only variant in the diet which was responsible for its lethal effect was the oxidized lipid plus the fact that an increase in stainable lipid could be demonstrated in the kidney, liver, and placentas of some animals led to an investigation of the distribution of fatty acids in the blood and tissues of these animals. It seemed appropriate to compare the fatty acid compositions of tissue lipids of pregnant and non-pregnant female rats fed the oxidized lipid diet (20 per cent casein, 61 per cent corn starch, 10 per cent brewers' yeast, 4 per cent salt mixture, and 5 per cent of the ethyl esters of a molecularly distilled fraction of oxidized cod liver oil) with those of similar rats fed a pellet stock diet containing approximately 5 per cent fat.