Intestinal Mucosa Microsomes Research Articles

Sanguinarine metabolism and pharmacokinetics study in vitro and in vivo.

Sanguinarine (SA) is a benzo[c] phenanthridine alkaloid which has a variety of pharmacological properties. However, very little was known about the pharmacokinetics of SA and its metabolite dihydrosanguinarine (DHSA) in pigs. The purpose of this work was to study the intestinal metabolism of SA in vitro and in vivo. Reductive metabolite DHSA was detected during incubation of SA with intestinal mucosa microsomes, cytosol, and gut flora. After oral (p.o.) administration of SA, the result showed SA might be reduced to DHSA in pig intestine. After i.m. administration, SA and DHSA rapidly increased to reach their peak concentrations (Cmax , 30.16±5.85, 5.61±0.73ng/ml, respectively) at 0.25hr. Both compounds were completely eliminated from the plasma after 24hr. After single oral administration, SA and DHSA rapidly increased to reach their Cmax (3.41±0.36, 2.41±0.24ng/ml, respectively) at 2.75±0.27hr. The half-life (T1/2 ) values were 2.33±0.11hr and 2.20±0.12hr for SA and DHSA, respectively. After multiple oral administration, the average steady-state concentrations (Css ) of SA and DHSA were 3.03±0.39 and 1.42±0.20ng/ml. The accumulation indexes for SA and DHSA were 1.21 and 1.11. The work reported here provides important information on the metabolism sites and pharmacokinetic character of SA. It explains the reasons for low toxicity of SA, which is useful for the evaluation of its performance.

Metabolism of cyadox by the intestinal mucosa microsomes and gut flora of swine, and identification of metabolites by high‐performance liquid chromatography combined with ion trap/time‐of‐flight mass spectrometry

Cyadox (CYX), 2-formylquinoxaline-1,4-dioxide cyanoacetylhydrazone, is an antimicrobial and growth-promoting feed additive for food-producing animals. To reveal biotransformation of CYX in swine intestine, CYX was incubated with swine intestinal microsomes and mucosa in the presence of an NADPH-generating system and swine ileal flora and colonic flora, respectively. The metabolites of CYX were identified using high-performance liquid chromatography combined with ion trap/time-of-flight mass spectrometry (LC/MS-ITTOF). Structural elucidation of the metabolites was precisely performed by comparing their changes in molecular mass, full scan MS/MS spectra and accurate mass measurements with those of the parent drug. Finally, seven metabolites were identified as follows: three reduced metabolites (cyadox 1-monoxide (Cy1), cyadox 4-monoxide (Cy2) and bisdesoxycyadox (Cy4)); hydroxylation metabolite (3-hydroxylcyadox 1-monoxide (Cy3)); hydrolysis metabolite of the amide bond (N-decyanoacetyl cyadox (Cy5)); a hydrogenation metabolite (11,12-dihydro-bisdesoxycyadox (Cy6)) and a side-chain cleavage metabolite (2-hydromethylquinoxaline (Cy7)). Only one metabolite (Cy1) was found in intestinal microsomes. Cy1, Cy2 and Cy4 were detected in intestinal mucosa, ileal and colonic flora. In addition, Cy3 and Cy5 were only obtained from ileal flora, and Cy6 and Cy7 alone were observed in colonic bacteria. The results indicated that N→O group reduction was the main metabolic pathway of CYX metabolism in swine ileal flora, intestinal microsomes and mucosa. New metabolic profiles of hydrogenation and cleavage on the side chain were found in colonic bacteria. Among the identified metabolites, two new metabolites (Cy6, Cy7) were detected for the first time. These studies will contribute to clarify comprehensively the metabolism of CYX in animals, and provide evidence to explain the pharmacology and toxicology effects of CYX in animals.

Comparison of in vitro metabolic conversion of capecitabine to 5-FU in rats, mice, monkeys and humans - toxicological implications

Capecitabine is an oral anticancer prodrug which is converted to 5-fluorouracil (5-FU) via 3 enzymatic steps, these being 5'-deoxy-5-fluorocytidine (5'-DFCR), 5'-deoxy-5-fluorouridine (5'-DFUR), and finally 5-FU by carboxylesterase (CES), cytidine deaminase (CDA), and thymidine phosphorylase (TP), respectively. Because rats, mice and monkeys are used for preclinical safety studies, we investigated the in vitro conversion from capecitabine to 5-FU by hepatic and intestinal mucosal microsomes and cytosols, to compare their metabolic activity to that of humans. Capecitabine was hydrolyzed to 5'-DFCR in hepatic and intestinal mucosal microsomes in these animal species. In humans and monkeys, CL(int) (V(max)/K(m)) for the hydrolysis of capecitabine in intestine (expressed as µl/min/g tissue) was much lower than that in hepatic microsomes but, in rats and mice, CL(int) was higher in intestine than in liver. In humans and monkeys, similar K(m) values and inhibition patterns by tetrahydrouridine (THU) a CDA inhibitor, were observed in CDA activity of hepatic and intestinal cytosols. However, rats showed very low CDA activity and mice showed non-Michaelis-Menten kinetics and a different inhibition pattern by THU. K(m) values for TP activity were almost similar in rats, mice, monkeys and humans. In conclusion, it was confirmed that monkeys are a suitable animal model for the safety assessment of capecitabine in terms of metabolic enzymes and it was suggested that higher toxic incidences in mouse small intestine were related to high hydrolytic activity of capecitabine in the small intestine.

QUANTITATIVE CONTRIBUTION OF CYP2D6 AND CYP3A TO OXYCODONE METABOLISM IN HUMAN LIVER AND INTESTINAL MICROSOMES

Oxycodone undergoes N-demethylation to noroxycodone and O-demethylation to oxymorphone. The cytochrome P450 (P450) isoforms capable of mediating the oxidation of oxycodone to oxymorphone and noroxycodone were identified using a panel of recombinant human P450s. CYP3A4 and CYP3A5 displayed the highest activity for oxycodone N-demethylation; intrinsic clearance for CYP3A5 was slightly higher than that for CYP3A4. CYP2D6 had the highest activity for O-demethylation. Multienzyme, Michaelis-Menten kinetics were observed for both oxidative reactions in microsomes prepared from five human livers. Inhibition with ketoconazole showed that CYP3A is the high affinity enzyme for oxycodone N-demethylation; ketoconazole inhibited >90% of noroxycodone formation at low substrate concentrations. CYP3A-mediated noroxycodone formation exhibited a mean K(m) of 600 +/- 119 microM and a V(max) that ranged from 716 to 14523 pmol/mg/min. Contribution from the low affinity enzyme(s) did not exceed 8% of total intrinsic clearance for N-demethylation. Quinidine inhibition showed that CYP2D6 is the high affinity enzyme for O-demethylation with a mean K(m) of 130 +/- 33 microM and a V(max) that ranged from 89 to 356 pmol/mg/min. Activity of the low affinity enzyme(s) accounted for 10 to 26% of total intrinsic clearance for O-demethylation. On average, the total intrinsic clearance for noroxycodone formation was 8 times greater than that for oxymorphone formation across the five liver microsomal preparations (10.5 microl/min/mg versus 1.5 microl/min/mg). Experiments with human intestinal mucosal microsomes indicated lower N-demethylation activity (20-50%) compared with liver microsomes and negligible O-demethylation activity, which predict a minimal contribution of intestinal mucosa in the first-pass oxidative metabolism of oxycodone.

Secretion of insulin-like growth factors in milk and their effect on the neonate

Many hormones and growth factors appear in milk and colostrum. Interest and research concerning these factors has recently been revitalized to study their role in both mammary and neonatal development. Colostrum is rich in insulin-like growth factor-I (IGF-I), IGF-II and IGF binding proteins (IGFBPs). The appearance of IGFs in colostrum is specific and the pattern of appearance is similar to that of immunoglobulins. Blood concentrations of IGF-I are significantly altered by the cessation and initiation of lactation. Increasing blood IGF-I concentrations by somatotropin treatment results in increased mass appearance of IGF-I into colostrum during the prepartum period with only modest increases at parturition. Because local production of IGF-I is low and not epithelial in origin, and because blood flux has been demonstrated, it is likely that most colostrum IGF-I is of blood origin. Conversely, mammary tissue epithelial cells synthesize and secrete IGFBPs. Peak secretory IGF-I and IGFBP colostrum concentrations are 10–30-fold that of blood concentrations. Two possible mechanisms of appearance are described. Because of the occurrence of IGFs in colostrum, the dietary effects of IGF-I on the newborn were of interest. Both intestinal and systemic changes were hypothesized. Calves were fed dietary treatments of: (1) milk replacer (MR), (2) pooled colostrum for four feedings, followed by MR, and (3) MR supplemented with 750 ng/ml of recombinant human (rh) IGF-I for one week. The dietary treatments had no effect on circulating neonate IGFBP profiles, yet IGFBPs changed with neonate age. Blood insulin and prolactin concentrations, but not IGF-I were altered initially. A latent increase in blood IGF-I concentrations in IGF-I-fed animals appeared after 4 to 5 days of feeding. At slaughter, there was more growth in intestinal proximal regions in rhIGF-I-fedanimals than in the colostrum-fed controls. Thermodynamic analysis of the IGF type I and II receptors of intestinal mucosal microsomes indicated that IGF-I feeding up-regulated (increased binding capacity) the type I IGF receptor, whereas dietary colostrum down-regulated the type II IGF receptor. These data support the concept of a physiological effect of colostrum-borne IGFs on the neonate.

CL 277,082: a novel inhibitor of ACAT-catalyzed cholesterol esterification and cholesterol absorption.

CL 277,082 (I) was found to be a potent inhibitor of acyl CoA:cholesterol acyltransferase (ACAT, EC 2.3.1.26) in microsomes from a variety of tissues with IC50 values of 0.14 microM for intestinal mucosal microsomes, 0.74 microM for liver, and 1.18 microM for rat adrenal. I was also shown to inhibit ACAT in cultured smooth muscle cells (IC50 = 0.8 microM) and was found to be specific in inhibiting cholesterol esterification since it did not inhibit fatty acid incorporation into triglycerides or phospholipids. Also, other cholesterol esterifying enzymes such as lecithin:cholesterol acyltransferase (LCAT) and pancreatic cholesterol esterase were not inhibited by I, nor was esterification of retinol by acyl CoA:retinol acyltransferase (ARAT) from intestinal mucosal microsomes inhibited. I was a potent inhibitor of cholesterol absorption in cholesterol-fed rats by markedly inhibiting increases in liver and serum cholesterol concentration (ED50 = 5.2 mg/kg per day) while increasing the excretion of neutral 14C-labeled sterol in the feces.

Comparison of hepatic, renal and intestinal bilirubin UDP-glucuronyl transferase activities in rat microsomes

1. 1. Bilirubin UDP-glucuronyltransferase activity and its dependence on substrate concentrations in rat liver, renal cortex and intestinal mucosa microsomes were studied. 2. 2. Bilirubin monoglucuronide synthesis from unconjugated bilirubin was a higher capacity, lower affinity step in comparison with bilirubin diglucuronide formation in the three tissues tested. 3. 3. Bilirubin glucuronide formation in liver microsomes showed a higher capacity but a lower affinity than extrahepatic ones. Renal cortex and intestinal mucosa exhibited similar kinetics parameters. 4. 4. In vitro bilirubin glucuronidation in renal cortex and intestinal mucosa was quantitatively important as compared with the hepatic one.

Hydroxylation of dibutylnitrosamine in the human liver and intestinal microsomal fractions.

The metabolism of the bladder carcinogen N-nitroso-di-n-butylamine (NDBA) was studied in microsomal preparations of tissues of patients of both sexes, aged 59-69 years undergoing abdominal surgery. Samples of liver, ileum, and colon were of normal histological appearance. For comparison, samples of rat liver and small intestinal mucosa microsomes were included in the study. Using 1-14C-labeled NDBA, the biotransformation to hydroxylation products retaining the nitroso group, NDBA-2-OH, NDBA-3-OH, and NDBA-4-OH, respectively, was investigated by reversed phase HPLC. In order to separate these metabolites, pooled samples were analysed by normal phase HPLC. The rate of hydroxylation of NDBA was found to be 5.5 times higher in rat liver microsomes compared to those from human liver (2.86 +/- 0.29 vs 0.52 +/- 0.03 nM X min-1 X mg-1). NDBA-3-OH proved to be the major metabolite formed (greater than 80% of total metabolites). The metabolism of NDBA was low but detectable in seven out of nine specimens of human gut, 0.1-0.5 nM X mg-1 in 1 h of incubation, and of the same order of magnitude in rat intestinal tissue (0.4-0.6 nM X mg-1).

Separation of two constitutive forms of cytochrome P-450 active in aminopyrine N-demethylation from rabbit intestinal mucosa microsomes

Two constitutive forms of cytochrome P-450, designated P-450ib and P-450ic, were purified from intestinal mucosa microsomes of untreated rabbits. P-450ib and P-450ic have minimal molecular weights of 56 000 and 49 000, respectively, as determined by calibrated sodium dodecyl sulphate polyacylamide gel electrophoresis. The CO-reduced difference spectral maximum of cytochrome P-450ib is at 450 nm and P-450ic is at 451 nm. Both the cytochromes preferentially demethylate aminopyrine, benzphetamine and N, N-dimethylaniline in the presence of NADPH-cytochrome P-450 reductase. Cytochrome P-450ib has absorption maxima at 417, 535 and 573 nm in the oxidized form, indicating that this cytochrome is in a low-spin state. Ouchterlony double-diffusion studies show that cytochrome P-450ib does not cross-react with antisera against liver cytochrome P-450 LM2 purified from phenobarbital-treated rabbits, but P-450ic cross-reacts with spur formation. Unlike cytochrome P-450ib, P-450ic is very similar, if not identical, to liver cytochrome P-450 LM2 on the basis of its molecular weight, spectral properties, catalytic activities and immunochemical properties.

Effects of zinc deficiency and castration on fatty acid composition and desaturation in rats.

The effects of zinc deficiency and testosterone on fatty acid composition of plasma lipids and microsomes of liver, intestine and testes were studied. The activities of fatty acid desaturase (delta 6 and delta 5) in rat liver and testes were also measured. A significant decrease in the level of arachidonic acid was observed in plasma of normal rats fed the zinc-deficient diet. Castration significantly decreased arachidonic acid but increased 20:3 fatty acid, which is negligible in normal rats. Testosterone and zinc administration restored arachidonic acid to normal values. Zinc deficiency does not significantly change the fatty acid profile in liver, but castration decreased both arachidonic and 22:6 fatty acid. Intestinal mucosal microsomes showed that the predominant fatty acid in this tissue, palmitic acid, is independent of zinc status, whereas polyunsaturated fatty acids 18:2 and 20:4 were decreased by zinc-deficient diet or castration. Zinc deficiency sharply decreased 22:5 fatty acid and to some extent, other polyunsaturated fatty acids in testis microsomes. These changes in fatty acids are in agreement with increased delta 9 desaturation and decreased delta 5 desaturase activity. In testes, both delta 6 and delta 5 desaturase activities are decreased in zinc deficiency. It appears that zinc influences the conversion of linoleic to arachidonic acid, whereas testosterone influences delta 6 desaturase activity. The data suggest that zinc deficiency may be one of the important factors in the causation of polyunsaturated fatty acid deficiency, which, in turn, may induce serum hypertriglyceridemia.

Some properties of a hexadecane hydroxylation system in rabbit intestinal mucosa microsomes.

Among many different tissues of rabbit, hexadecane hydroxylation activity was found in only small intestine and liver microsomes. Both activities for hexadecane and decane hydroxylation in intestinal microsomes were significantly higher than those in liver microsomes. In contrast, the hydroxylation activities of p-xylene, benzo(a)pyrene, and decanol, and the demethylation activity of aminopyrine in the former were much lower than those in the latter. The intestinal microsomes converted [1-14C]hexadecane to cetyl alcohol, but not further to palmitic acid. Hexadecane hydroxylation activity was found to be markedly stimulated by non-ionic detergents such as Triton X-100, Nonident P-40, and Emulgen 913. Phosphatidylcholine and phosphatidylethanolamine stimulated the activity to a lesser extent. The hydroxylation was inhibited by various aliphatic hydrocarbons with carbon numbers larger than 10, but not by aromatic and polycyclic hydrocarbons. Hexadecane hydroxylation activity was solubilized from the intestinal microsomes and reconstituted with a partially purified cytochrome P-450 fraction, and intestinal NADPH-cytochrome c reductase, or spinach ferredoxin and ferredoxin-NADP reductase. The chromatography of the crude cytochrome P-450 preparation on hydroxylapatite separated at least two cytochrome P-450 fractions; one preferentially hydroxylating hexadecane, and the other preferentially hydroxylating myristic acid. The results suggest that rabbit intestinal mucosa microsomes had a cytochrome P-450 species specialized for hexadecane hydroxylation.

Studies on the incorporation of sn-[1,3- 14C]glycerol 3-phosphate into glycerolipids by intestinal mucosa

Intestinal mucosal microsomes from several animal species (non-fasted) were used to study the formation of glycerolipids from sn-[1,3- 14C]glycerol 3-phosphate and palmitoyl-CoA. Rates of glycerolipid biosynthesis were species dependent, since mouse and rat were quite low compared to hamster, guinea pig and man. Under the usual incubation conditions, guinea pig intestinal microsomes formed primarily phosphatidic acid (85–90%). However, when Mg 2+ was added to incubations containing intestinal microsomes prepared in the presence of 5 mM EDTA, there was a marked increase in neutral lipid biosynthesis (5–10-fold). These results suggest that intestinal microsomes contain Mg 2+-dependent phosphatidate phosphohydrolase (EC 3.1.3.4) which is involved in glycerolipid biosynthesis. All divalent cations studied increased sn-glycerol-3-phosphate acyltransferase (EC 2.3.1.15) activity; however, Ca 2+, Ba 2+, Zn 2+, Cd 2+, Ni 2+, and Mn 2+ antagonized the Mg 2+-dependent rise in neutral lipid formation. In all cellular preparations studied, the ratio of neutral lipid (diacylglycerol + triacylglycerol) to total lipid (phosphatic acid + neutral lipid) was low, suggesting that phosphatidate phosphohydrolase may be rate limiting in intestinal neutral glycerolipid biosynthesis.

Reconstruction of aryl hydrocarbon hydroxylase from rabbit intestinal mucosa microsomes.

Benzo(a)pyene hydroxylation activity was solubilized from rabbit intestinal mucosa microsomes and reconstituted with a cytochrome P-450 preparation obtained by fractionation with 6-amino-n-hexyl Sepharose 4B, hydroxylapatite and CM-Sephadex C-50, and partially purified NADPH-cytochrome c reductase. Phosphatidylserine was required for the maximal activity, while phosphatidylcholine had no stimulatory effect. The carbon monoxide difference spectrum of the cytochrome P-450 fraction showed a maximum peak at 450 nm. Although another cytochrome P-450 fraction was active for hexadecane hydroxylation, this fraction had little activity. The results indicate that more than one cytochrome P-450 exists in the intestinal mucosa microsomes.

Comparison of the drug metabolising ability of rat intestinal mucosal microsomes with that of liver

A study of the drug metabolizing capabilities of rat intestinal microsomes was undertaken and a direct comparison made with analogous activity in hepatic microsomes. The cofactor requirements and K m values of the cytochrome P-450 mediated 7-ethoxycoumarin O-de-ethylation in the epithelial cell microsomes was similar to that in hepatic microsomes, whereas the V max values were 5–6 fold lower in the intestinal preparation. Greater differences were seen in other cytochrome P-450 dependent reactions. The K m value for biphenyl-4-hydroxylation was an order of magnitude lower in the intestinal microsomes, whereas the 4-hydroxylation of aniline and acetanilide and the N-demethylation of ethylmorphine and aminopyrine, although very active in hepatic microsomes, were barely detectable in intestinal microsomes. The O-de-ethylation inhibition characteristics of metyrapone and 7,8-benzoflavone but not SKF 525 A, were similar in the liver and intestinal preparations. These findings and the alterations in drug oxidation activity after PB and 3MC pretreatment suggest that there are both quantitative and qualitative differences in the cytochrome P-450's between liver and intestine. This suggestion is further supported by the difference in the safrole adduct binding spectrum with NADPH or with cumene hydroperoxide between the two tissues from control, phenobarbitone and 3-MC pretreated rats. O-Deacetylation of 4-nitrophenyl acetate and indoxyl acetate in intestinal and liver preparations was very comparable while acetanilide N-deacetylation was three times more active in the liver. Glucuronidation of 1-naphthol by the intestinal microsomes showed a similar K m but a lower V max than the liver microsomes. No change was observed in any of these enzymes following pretreatment of rats with PB or with 3-MC.

Fatty Acid ω- and (ω-1)-Hydroxylation in Rabbit Intestinal Mucosa Microsomes

The microsomes from rabbit intestinal mucosa which had been washed quickly and thoroughly with phenylmethylsulfonyl fluoride were found to catalyze the hydroxylation of fatty acids in the presence of NADPH and molecular oxygen. Myristic and palmitic acids were converted to the corresponding omega-and (omega-1)-hydroxy fatty acids, whereas lauric acid was converted only to 12-hydroxylauric acid, and capric acid, to 9-and 10-hydroxycapric acids together with an unknown polar acid. Among these fatty acids, both myristic and lauric acids appeared to be the most efficient substrates. The inhibition of the hydroxylation by SKF 525-A and carbon monoxide suggested that the activity depended upon cytochrome P-450. The specific activity of the fatty acid hydroxylation was almost constant along the small intestine, while the aminopyrine N-demethylation activity and the cytochrome P-450 content were highest at the proximal end of the intestine and progressively declined toward the caudal end. The cytochrome P-450 was solubilized from the intestinal microsomes and purified by 6-amino-n-hexyl Sepharose 4B chromatography. The partially purified cytochrome P-450 was active in fatty acid hydroxylation in combination with intestinal NADPH-cytochrome c reductase and phosphatidylcholine.

Chick kidney microsomal cytochrome P-450 involvement in the metabolism of 25-hydroxyvitamin D 3

A new metabolic activity is described for liver, kidney, and intestinal mucosal microsomes of rachitic chicks that is associated with the metabolism of 25-hydroxyvitamin D 3 (25-OH-D 3). Cytochrome P-450 was shown to be an integral component of this metabolism, supporting the monooxygenase nature of the microsomal activities. The 25-OH-D 3 was the specific substrate inasmuch as the reactions could not be demonstrated to occur when vitamin D 3, 1,25-dihydroxyvitamin D 3, or cholesterol was the substrate. The microsomal cytochrome P-450 concentrations of rachitic chick liver, kidney, and intestinal mucosa were 0.108, 0.068, and 0.270 nmol/mg of protein, respectively. The cytochrome b 5 concentrations of the tissues in the same order were 0.198, 0.123, and 0.306 nmol/mg of protein. Normal chick tissue P-450 and b 5 concentrations were the same as those in the rachitic. The kidney enzyme has a pH optimum of 6.6 to 7.5 and an apparent K m of 2.5 μ m. At a substrate concentration of 0.417 μ m, the rate of 25-OH-D 3 metabolism was proportional to a microsomal protein concentration of up to 3 mg/ml. The activity was inhibited 100% by 10 μ m n-ethylmaleimide, 1 m m o-phenanthroline, 1 m m metyrapone, or by a carbon monoxide:oxygen mixture of 90:10 ( v v ). A 50% inhibition of activity was produced by 5 m m sodium cyanide, but 10 μ m antimycin A and rotenone were ineffective. Supporting evidence for the participation of a cytochrome P-450 species in the renal microsomal metabolism of 25-OH-D 3 included the type I substrate-induced spectral shift at 0.5 μ m 25-OH-D 3 with a modified positive absorption at 405 nm and a negative absorption at 422 nm. These findings suggest that a metabolically active cytochrome P-450-substrate complex is involved in the renal microsomal biotransformations of 25-OH-D 3 as a component of the regulatory processes of vitamin D 3 metabolism.

The metabolism in vitro of enantiomeric 1- O-alkyl glycerols and 1,2- and 1,3-alkyl acyl glycerols in the intestinal mucosa

1. 1. The utilization of a series of monoalkyl glycerols as substrates for triglyceride biosynthesis via the monoglyceride pathways has been investigated. 2. 2. Both the sn-1- and 2-alkyl glycerols are satisfactory substrates. However, the sn-3 derivatives are not acylated by intestinal mucosal microsomes. 3. 3. The sn-monoacyl glycerol is directly utilized for triglyceride biosynthesis. 4. 4. The intermediate 1,3-alkyl acyl glycerols are incorporated into triglycerides without isomerization to the corresponding 1,2 analogs. 5. 5. The substrate requirements for the mono- and diglyceride transacylases are discussed.

Biosynthesis of triglycerides from triose phosphates by microsomes of intestinal mucosa.

AbstractMicrosomes of hamster intestinal mucosa synthesize triglycerides from dihydroxyacetone phosphate or DL‐glyceraldehyde 3‐phosphate in the presence of palmitate, ATP, CoASH and NADPH or NADH but in the absence of soluble enzymes. An inhibitor of triose phosphate isomerase, 1‐hydroxy‐3‐chloro‐2‐propanone phosphate, completely inhibits glyceride synthesis from glyceraldehyde 3‐phosphate. This compound does not inhibit glyceride synthesis from dihydroxyacetone phosphate. These reactions confirm the conversion of glyceraldehyde 3‐phosphate to dihydroxyacetone phosphate prior to glyceride synthesis.