A sensitive, reliable, and easily performed procedure is described for the prenatal and postnatal detection of inborn errors of propionate, methylmalonate, and cobalamin metabolism using cultured amniotic cells and skin fibroblasts. With this assay, control fibroblast lines incorporated a mean of 6.89 nanoatoms 14C/mg protein from [1-14C]propionate into trichloroacetic acid (TCA)-precipitable cell material in 10 h. Twenty-five mutant fibroblast lines from patients with propionicacidemia or one of the methylmalonicacidemias fixed 0.04 to 0.93 nanoatoms 14C/mg. Considerable variation was observed, both among and within discrete mutant classes, with respect to the residual amount of propionate pathway activity, possibly reflecting further molecular heterogeneity in these disorders. We applied this procedure to cultured amniotic cells from controls and 4 midtrimester pregnancies at risk for methylmalonicacidemia and diagnosed one fetus with a methylmalonyl CoA apomutase defect and 3 fetuses which were unaffected.

We measured the synthesis of 5'-deoxyadenosylcobalamin (AdoCbl) in fibroblast extracts from patients with inherited methylmalonicacidemia due to deficient activity of the cobalamin-dependent holoenzyme, methylmalonyl-CoA mutase (EC 5.4.99.2). Previous studies with intact fibroblasts from patients whose holoenzyme deficiency was secondary to abnormal cobalamin metabolism had defined two phenotypes, one in which whole cells failed to accumulate AdoCbl and a second in which they failed to accumulate both AdoCbl and the second cobalamin coenzyme, methylcobalamin. With a broken cell assay of AdoCbl synthesis, we have further subdivided the first phenotype into two classes.

and P e d i a t r i c s , New Haven. Inherited methylmalonicacidemia (MM-emia) due t o d e f i c i e n t a c t i v i t y of methylmalonyl-CoA mutase may be caused by an abnormal mutase apoenzyme or by reduced holoenzyme a c t i v i t y secondary t o impaired synthesis of t h e mutase-requiring coenzyme, 5'-deoxyadenosylcobalamin (Ado-Cbl).. We studied Ado-Cbl synthesis i n 5 f i b r o b l a s t l i n e s from unrelated p a t i e n t s who have MM-emia due t o defective cobalamin (vitamin B12) metabolism. Their whole c e l l s i n c u l t u r e f a i l t o synthesize Ado-Cbl but make normal amounts of t h e other cobalamin coenzyme, methy lcobalamin.After breaking t h e c e l l s t o define t h e s t e p s i n Ado-Cbl synthesis, we assayed t h e f i n a l s t e p which combines the reduct i o n of cob(1I)alamin t o cob(1)alamin and the adenosylation of cob(1)alamin. Subcellular f r a c t i o n a t i o n s t u d i e s localized t h i s a c t i v i t y t o mitochondria. I n crude broken c e l l e x t r a c t s , enzyme a c t i v i t y was d e f i c i e n t or absent i n 3 of t h e 5 mutant c e l l l i n e s (4,0,0 pg Ado-~bl/mg protein/30 min;control l i n e s , 23 + 8). The other two MM-emia l i n e s had normal a c t i v i t i e s (23 and 17). These r e s u l t s e s t a b l i s h deficiency of Ado-Cbl synthesizing enzyme a s one cause of abnormal cobalamin metabolism and MM-emia. The two MM-emia l i n e s t h a t had normal enzyme a c t i v i t y suggest t h e r e i s f u r t h e r heterogeneity within the disorders of cobalamin metabolism a s well a s i n the MM-emia phenotype.IS CYSTINE ESSENTIAL FOR PREMATURE INFANTS? John I . Malone, John S. Curran, and Lewis A. Barness. Univ. of S. Fla. and Tampa General Hospital, Dept. of P e d i a t r i c s , Tampa 33620.Growth i n premature i n f a n t s (weight 1000-2000 gms, 26-32wks gestation) fed a commercially a v a i l a b l e low c y s t i n e formulawas not s i g n i f i c a n t l y d i f f e r e n t from those fed a formulation cont a i n i n g twice t h e d i e t a r y cystine. The formulas were i d e n t i c a l with t h e exception of p r o t e i n qua1ity;the low cystine formula contained 13mg/100ml and t h e high cystine formula 24mg/100ml. There was no s i g n i f i c a n t difference i n growth r a t e of i n f a n t s , i n t h e serum concentration o r t h e t i s s u e l e v e l s (glutathione) of c y s t i n e i n t h e two groups. Cystathionase act i v i t y i n t h e l i v e r has been reported t o be absent i n premature i n f a n t s . These r e s u l t s i n d i c a t e t h a t t h e low c y s t i n e formula contains adequate c y s t i n e f o r normal growth of premature inf a n t s & provides normal serum & t i s s u e l e v e l s of t h i s amino acid. W e have postulated (APS-SPR, Apr. '72) t h a t s h o r t and/or medium chain f a t t y acid accumulation i s important i n t h e pathogenesis of RS, causing mitochondria1 swelling. c e r e b r a l edema, hypoglycemia and f a t t y viscera. This study r e p o r t s gas chromatographic q u a n t...