A P P L I E D N U T R I T I O N A L INVESTIGATION Nutrition Vol. 13, No. 4, 1997

Home Parenteral Nutrition and Vitamin B12 Status

DANIEL LAMBERT, PHD,* BERNARD MESSING, MD,'~ SAMIRA BENHAYOUN, PHD,* FRANCOIS THUILLIER, MD,'~ CHARLES ADJALLA, PHD,* MARTINE BELIAH, RN,'~

MARIE-ANDRt~E GELOT, PHD, MD,:~ JEAN PIERRE NICOLAS, MD,* AND JEAN-LOUIS GUI~ANT, MD, PHD*

From *LN.S.E.R.M. U 308, Nancy, I'LN.S.E.R.M. U 290 H6pital Saint-Lazare, Paris, and SLaboratoire de Biochimie B, HOpital Central, CHRU Nancy, France

Date accepted: 6 July 1996

ABSTRACT

The vitamin B~2 status of 20 subjects who were on home parenteral nutrition after surgical or functional small bowel resection and were given 1000 #g cyanocobalamin every 3 mo was studied by comparing their plasma vitamin Bl2, homocysteine (HS), and methylmalonic acid (MMA) concentrations. The plasma vitamin B12 concentration (median 145 pmol/L, 95% confidence interval: 123-217) was subnormal in four cases and borderline in four others. In the "low B~2" group, the concentrations of the markers of vitamin B~2 deficiency were in the normal range: HS 10.7 #mol/L (8.0-12.3); and MMA, 0.15 #mol/L (0.09-0.19). References values were HS, 10.0 #mol/L (9.4-12.6); and MMA, 0.16 #mol/L (0.10-0.19). Thus, there were no metabolic signs of vitamin B12 deficiency in these subjects on parenteral nutrition, despite the fact that their plasma vitamin B12 levels were tow. Analysis of individual data showed that the four patients with low circulating Bx2 had markers of intracellular vitamin B12 deficiency in the normal range. Nutrition 1997;13:313-318. ©Elsevier Science Inc. 1997

Key words: parenteral nutrition, vitamin B12, analogs, malabsorption, transcobalamins

INTRODUCTION

All patients requiring home parenteral nutrition (HPN) fol- lowing surgical resection of the intestine are routinely given a vitamin B12 supplement. The few published studies on vitamin B12 and PN are somewhat contradictory. Shenkin and Fell ~ detected no lack of vitamin B12 in patients on artificial nutrition, while others 2 have reported that the lack of vitamin B~2 appears rapidly once patients begin PN and 12-16% of the subjects may suffer from low circulating B12.

There may be several reasons why patients suffering from gastrointestinal disorders that require PN may have an altered vitamin B~2 status, including malabsorption due to a damaged ileum or ileal excision, or a bacterial overgrowth (blind loop syndrome) that consumes cyanocobalamin and produces excess analogs such as cobinamides and cobamides. 3

The way in which the vitamin B~2 supplement is provided to subjects on HPN may vary considerably. 4-7 The amounts required and the frequency of doses via PN may not be suffi- cient for all subjects because of variations in individual needs.

None of the actual studies in PN has evaluated a true B12 deficiency by measuring the circulating concentration of cobal- amin-dependent metabolites such as homocysteine (HS) and methylmalonic acid (MMA). In fact, most patients having a low plasma B12 show no clear clinical sign of vitamin defi- ciency, as there is no megaloblastic anemia. 2 Several parameters provide a better picture of the vitamin B12 status than simply measuring the plasma vitamin B~2 concentration. One is the concentration of saturated transcobalamin II (TCII; holo-trans- cobalamin II), as TCII-B12 is the form of vitamin B12 that is exchanged with the tissues. The other, perhaps more important, parameters are the plasma concentrations of the metabolic markers of vitamin B12 deficiency, MMA and HS. Lindenbanm et al. 8 showed the sensitivity of these markers for detecting a lack of vitamin, even when there is no megaloblastic anemia or low plasma vitamin B12. However, HS is not a specific parameter for the diagnosis of vitamin B12 deficiency and checking the normalization of HS and MMA blood levels after B12 supplementation is recommended?

Correspondence to: D. Lambert, PhD, I.N.S.E.R.M. U 308, Facult6 de M6decine, Av. de la For~t de Haye, B.P. 184, 54505 Vandoeuvre C6dex, France.

Nutrition 13:313-318, 1997 ©Elsevier Science Inc. 1997 Printed in the USA. All rights reserved. ELSEVIER

0899-9007/97/$17.00 PII S0899-9007(97)00081-6

314 PARENTERAL NUTRITION AND VITAMIN B l2

TABLE I.

DESCRIPTION OF PATIENTS ON TOTAL PARENTERAL NUTRITION

Patient Age (y) Sex Diagnosis Stc D J I ICV RC TC LC S R

1 78 F Postradiation enteritis + + 85 0 0 0 0 + + + 2 27 M Intestinal ischemic syndrome + + 10 0 0 + + + + + 3 30 M Colon polyposis + desmoid tumor + + 60 0 0 0 0 0 0 (+) 4 65 F Postradiation enteritis + + + + + + + + 0 0 5 74 F Celiac disease + + + + + + + + + + 6 77 M Scleroderma + + + + + + + + + + 7 67 F Postradiafion enteritis (small intest. + colon) + + + + + + + + 0 0 8 69 M Intestinal ischemic syndrome + + 15 0 0 0 + + + + 9 60 M Crohn's disease + + 80 0 0 + + + + 0

10 57 M Intestinal ischemic syndrome + + 30 0 0 + + + + + 11 26 F Colon ileiomyosarcoma + + 15 0 0 0 + + + + 12 39 M Intestinal ischemic syndrome + cirrhosis + + 30 10 + + (+) (+) (+) (+) 13 44 M Crohn's disease + + 80 0 0 0 . + + + + 14 48 F Colon polyposis + desmoid tumor + + 0 0 0 0 0 0 0 0 15 43 M Intestinal ischemic syndrome + cirrhosis + 0 0 0 0 0 0 + + + 16 46 F Hypo IgG, IgA, IgM + intest, villi atrophy + + + + + + + + + + 17 41 F Pseudo obstruction + + + + + + 0 0 0 + 18 38 M Crohn's disease + + 45 0 0 0 + + + + 19 28 M Crohn's disease + + 90 0 0 0 0 0 0 0 20 41 M Crohn's disease + + 40 0 0 0 0 0 0 (+)

Stc, stomach; D, duodenum; J, jejunum; I, ileum; ICV, ileocecal valvula; RC, TC, LC, right, transverse, and left colon, respectively; S, sigmoid colon; R, rectum; +, present; 0, absent; (+), brackets corresponds to "present" but not in continuity. Numbers represent remnant length in cm.

This study is the first one that documents the vi tamin B12 status of patients on HPN by compar ing the plasma concen- trations of vi tamin B~2, saturated and unsaturated TCs, HS, and MMA.

MATERIALS AND METHODS

Patients

A total of 20 patients, 8 women and 12 men, ages 2 6 - 7 8 y (median 49.9 y) , requiring HPN for more than 1 y were studied. All suffered from severe disorders of gastrointestinal function (Table I) . The normal values for the parameters re- lated to vitamin Blz and its analogs were taken as those mea- stared in 30 healthy subjects ages 3 2 - 6 8 y (median 52.9 y) who had never been given vitamin BIe supplements.

Nutrition

All patients were on overnight cyclic PN, as described pre- viously) ° Details of the regime are given in Table II, except for minerals. Patients were given magnesium, potassium, sodium, and phosphorus according to their specific needs. Vitamin B 12 supple- ments (cyanocobalamine) were given as intramuscular injections of 1000 /.zg every 3 mo. Folate supplementation (Lederfoline, Lederlr, Oullins, France) was intramuscular 50 mg once a month. The last vitamin B12 and folate supplements had been given 3 mo (B12) and 1 mo (folate) previously. No patient was given any other treatment containing vitamin BI2.

Blood Samples

Samples (5 mL) of blood for measuring vitamin B~2, its analogs, and HS were taken in the postabsorptive state (over- night) via an antecubital vein and collected over 1 mg/rnL EDTA. Blood was collected without additive for MMA. All samples were centrifuged at 2,700 g for 20 rain and the p l a s m a or serum was stored at -20°C.

Analysis

Cobalamin and Its Analogs The plasma concentrations of cobalamin and its analogs

were determined by the method of Kolhouse et al.l~ as modified by our group] 2-14

TABLE II.

DESCRIPTION OF TPN SOLUTIONS

Nutrient Quantity/d

Protein (Vamine)* 65 __+ 15 g Dextrose 1040 _+ 340 kcal Fat (Intralipide)* 180 _+ 168 kcal Vitamins

Hydrosol polyvitamin** 6.0 mL a-tocopherol 7.7 _+ 0.2 mg Ergocalciferol 286 IU Vitamin C 1150 mg

Trace elements Nonan*** 12 _+ 4 mL Selenium 75 _+ 28 #g Zinc 10 mg

* Pharmacia, Saint Quentin Yvelines, France. ** Hydrosol polyvitamin (Roche Pharmaceuticals, Neuilly, France) contains retinol: 15 000 US; thiamin HCI: 6 mg; riboflavin: 4.5 rag; d-penthothenic acid: 12 mg; pyridoxine HCI: 6 mg; nicotinamide HCI: 30 mg; vitamin C: 150 rag; a-tocopherol: 6 rag. *** Nonan (40 mL) (Aguettant, Lyon, France) contains iron: 1 mg; copper: 0.48 rag; manganese 1 rag; zinc: 4 mg; fluorine 1.45 mg; cobalt: 1.47 #g; iodine: 1.52/~g; selenium: 40 #g; molybdenum: 25 #g; purified water q.s.p, for 40 mL.

PARENTERAL NUTRITION AND VITAMIN B12 315

Unsaturated TC The two proteins that bind corrinoids are TCII and haptocor-

rin (Hc or TC I+II I ) . The "apo" forms are the free carriers. The values discussed in RESULTS are the capacities of the card- ers to bind vitamin B12. Apo-TCII and apo-Hc concentrations are expressed in pmol vitamin Blz/L.

The unsaturated binding capacities of Hc and TCII were determined in duplicate by the method of Gottlieb et al. 15 TCII was separated from Hc by precipitation with microfine QUSO G32 (Philadelphia Quartz, Valley Forge, PA, USA). 16

Saturated TC The "holo" form is the carder bound to the vitamin BI2.

Concentrations of holo-TCII and holo-Hc are given in pmol vita- min B1z bound/L. In the protocol used, 17 saturated Hc is equiva- lent to the total corrinoid (vitamin B12 + analogs) content of serum treated with heparin Sepharose (HS) (Pharmacia, Saint Quentin Yvelines, France).

Other Tests The plasma concentration of HS was measured according

to Brattstr6m et al. 18 and that of serum MMA by capillary gas chromatography-mass spectrometry as tert.-butyldimethylsilyl derivatives, according to Straczek et al.19 Serum glutamate- oxaloacetic transaminase (GOT), glutamate-pyruvate transam- inase (GPT), alkaline phosphatase (AP), total bilirubin, and gamma glutamyl transferase (gamma-GT) were measured in a Kern-O-Mat Phase II (Coultronics, Margency, France) auto- analyzer using reagents from Biotrol, Paris, France. The specific proteins (albumin [ ALB ], transthyretin [TTR], retinol-binding protein [RBP], and C-reactive protein [CRP]) were assayed by nephelometry on an Array autoanalyser (Beckman, Gagny,

France). Folates were measured by immunoradiometric assay (IRMA kit, Ciba Coming, Eragny, France). Microbial over- growth was determined with a breath test based on the hydroly- sis of 50 g glucose given orally. 2°

The presence or absence of the stomach, duodenum, ileoce- cal valve, a segment of the colon (right, transverse, left, or distal segments) and rectum, and the presence of villus atrophy were all recorded.

Statistical Analysis

All data are given as medians and 95% confidence intervals (CI). As the distributions of vitamin B12 and corrinoid concen- trations were non-Gaussian, a nonparametric Mann-Whitney unpaired test was used to compare groups and the Spearman ranking test was used to determine correlation coefficients.

RESULTS

General

The residual vitamin B12 concentrations were within the range of values found by others for subjects on PN for each of the disorders examined (Table III). Under our analytical condition, a plasma vitamin B12 concentration below 85 pM was considered to be deficient, and values of 85-120 pmol/L were considered borderline. 21 The mean plasma vitamin B12 concentration in these patients was 145 pmol/L (95% CI: 126- 217). The mean values were not significantly different from the control group values for vitamin Bl2 parameters (Table IV). However, the plasma vitamin B12 concentrations were below 85 pM in 20% of cases (n = 4) and borderline in 20% of cases (n = 4). High vitamin B12 analog concentrations were observed in only two cases.

TABLE III.

INDIVIDUAL VALUES OF PLASMA VITAMIN B12 PARAMETERS (VITAMIN BI2, VITAMIN B12 ANALOGS, VITAMIN B12 BOUND TO HAPTOCORRIN [B12-HOLO-HAPTOCORRIN], VITAMIN B12 BOUND TO TRANSCOBALAMIN II [B12-HOLO-TRANSCOBALAMIN II], PLASMA HOMOCYSTEINE, SERUM METHYLMALONIC ACID (MMA),

AND SERUM FOLATES FOR PATIENTS ON TOTAL PARENTERAL NUTRITION

B12 Analogs B12-holo-hapotocorrin B12-hoto-trans-cobatamin II Homocysteine MMA Serum folates Patient B12 Value pmol/L pmol/L pmol/L pmol/L #mol/L #mol/L nmol/L

1 normal 295 249 272 22 18.3 0.23 30.5 2 normal 133 8 125 5 3.5 0.21 9.8 3 normal 248 116 222 25 10.7 0.15 18.9 4 normal 148 36 107 38 8.7 0.12 29.8 5 borderline 116 82 85 25 11.2 0.2 6.8 6 normal 186 6 176 10 9.3 0.12 7.7 7 normal 144 6 112 31 8.8 0.07 10.9 8 borderline 105 132 65 40 7.6 0.09 5.7 9 normal 171 63 134 36 16.8 0.22 16.4

10 low 66 75 54 7 11.2 0.15 28.2 11 low 40 151 33 6 8.2 0.17 7.3 12 borderline 111 94 68 39 8.3 0.15 6.1 13 low 83 73 71 11 10.6 0.1 4.5 14 normal 480 -0 450 48 15.6 0.15 4.5 15 borderline 117 186 62 54 6.8 0.13 3.4 16 normal 207 80 179 27 7.6 0.17 15.7 17 low 83 109 68 10 10.8 0.15 11.1 18 normal 224 7 209 14 11 0.23 10.0 19 normal 199 158 152 46 8 0.09 10.7 20 normal 263 167 229 39 9.2 0.1 7.3

% of anormal test: abnormal 20 10 35 35 0 0 20 borderline 15 45 - - - - 15 0 10

316 PARENTERAL NUTRITION AND VITAMIN B12

TABLE IV.

PLASMA VITAMIN B12 PARAMETERS (VITAMIN BI> TOTAL CORRINOIDS, VITAMIN B12 BOUND TO HAPTOCORRIN [B12- HOLO-HAPTOCORRIN], VITAMIN BI: BOUND TO TRANSCOBALAMIN II [B~2-HOLO-TRANSCOBALAMIN II] AND VITAMIN

BI2 ANALOGS), MARKERS OF VITAMIN B12 DEFICIENCY (PLASMA HOMOCYSTEINE AND SERUM METHYLMALONIC ACID, [MMA]), AND SERUM FOLATES FOR PATIENTS ON TOTAL PARENTERAL NUTRITION AND CONTROLS

B12-holo- B12 Corrinoids h a p t o c o r r i n Corrinoids holo-haptocorrin Analogs Analogs

Subject pmol/L pmol/L pmol/L pmol/L pmol/L (%)

Patient Whole group 145 (123-217) 217 (205-315) 118 (97-190) 152 (142-243) 81 (57-122) 40 (23-45) Normal B~2 178 (145-247)** 235 (219-348) 143 (112-218)* 163 (143-270) 81 (46-127) 29 (17-39)** Low B~2 74 (35-100) 173 (129-210) 61 (29-83) 146 (77-211) 92 (44-160) 55 (37-81) Reference value 141 (120-201) 219 (189-308) 170 (151-230) 251 (205-317) 88 (67-111) 35 (32-38)

B12-holo-transcobalamin II Corrinoids-holo-transcobalamin II Serum Folates Homocysteine MMA pM pM nM #M #M

Patient Whole group 26 (19-33) 48 (33-79) 9.9 (8.3-16.2) 9.2 (8.5-11.7) 0.15 (0.13-0.17) Normal B12 34 (24-37)*** 55 (34-90) 9.9 (7.7-16.5) 9.0 (8.0-12.1) 0.15 (0.12-0.18) Low B~2 8 (4-12) 38 (0-77) 9.2 (0.0-29.6) 10.7 (8.0-12.3) 0.15 (0.09-0.19) Reference value 54 (37-76) 78 (45-127) 8.9 (5.8-14.8) 10.0 (9.4-12.6) 0.16 (0.10-0.19)

Representation of data is median + confidence interval: 95% inferior, 95% superior. "Low Bt2" patients (n = 4); "Normal B~2" patients (n = 16). A nonparametric Mann-Whitney unpaired test was used between the two groups of patients. No significant difference was obtained with the control population: * P < 0.05; ** P < 0.02; *** P < 0.001.

The plasma folate concentration (9.9 nmol/L [ 8.3-16.2 ]) was in the range of that of controls (8.9 nmol/L [ 5.8-14.8 ] ). Twenty percent of patients had low values and 10% were borderline (6 - 7 nmol/L). There was no significant increase in the concentra- tions of either HS (9.2 #mol/L [ 8.5 - 11.7 ] ) or MMA (0.15/zmol/ L [0.13-0.17]) in the tested subjects. (Control values: HS, 10.0 #mol/L ([9.4-12.6]; and MMA, 0.16 /.tmol/L [0.10-0.19]) (Table IV). However, the HS concentration was borderline in 15% of cases (n = 3: 18.3, 16.8, and 15.6 #mol/L). These three patients had normal vitamin Blz values (295, 171, and 480 pmol/ L). One had a low folate value.

Regarding the relationship between vitamin B12 and markers of vitamin Bj2 deficiency, the main point is that the four patients with low B12 values also had markers of vitamin BI2 deficiency that lay within the 95% CI (vitamin B12 Values in Table III mad CIs in Table IV).

The binding capacities of the vitamin BI2 carriers, apo-TCII (598 pmol B12/L [542-707]), and apo-Hc (121 pmol B12/L [97-190]) in the test subjects were not significantly statistically different from those of the controls (apo-TCII, 590 pmol B12] L [536-652]; and apo-HC, 169 pmol B12/L [125-272]). The standard laboratory screening tests showed a decreased plasma ALB (34 g/L [33-37]; normal range 45-50, P < 0.001) and significant increases in AP (117 IU/L [93-194]; normal range < 90 IU/L) and gamma-GT (44 IU/L [48-186]; normal < 40 IU/L). The values for RBP (0.045 g/L [0.041-0.064]), TTR (0.32 g/L [0.26-0.35]), CRP (3.0 mg/L [3.6-12.5]), GOT (24 IU/L [21-40]), GPT (29 1U/L [22-43]), and bilirubin (7.0 #mol/L [5.9-10.5]) were not significantly statistically different from the reference values.

Patients With Low Plasma Vitamin B~2 Concentrations

The patients with low vitamin B12 concentrations (85 pmol/ L) also had lower amounts of holo-TCII (8 pmol B~z/L [4 - 12], P < 0.005) and holo-Hc (61 pmol B~2/L [29-83]) , P

< 0.05) than did patients with normal plasma vitamin B12 (holo-TCII, 34 pmol B J L [24-37]; holo-Hc, 143 pmol B12/ L [112-218]). The analog concentrations in the two groups were not significantly statistically different. Only the B12 values differed, which led to an increased percentage of analogs in the "low B12" group. The vitamin B12 analog values were not elevated in these patients with low B12 levels.

The plasma folate concentrations in patients with a low plasma vitamin B 12 concentration (9.2 nmol/L [ 0.0 - 29.6 ] ) and patients with normal B12 values (9.9 nmol/L [7.7-16.5]) were not significantly statistically different. One of the four patients with low B12 had a low serum folate value (below 6 nmol/L).

Among biological parameters, only the TTR was lower (0.21 g/L [0.17-0.32]) in a group of patients with "low B12" or borderline B12 values (85-95 pM B12) than in the normal B12 patients (0.37 g/L [0.28-0.38], P < 0.05). As there were only four cases in the "low Biz" group (n = 4), none of the other laboratory parameters for this group was significantly statistically different from those for the other TPN patients. The laboratory parameters were therefore analyzed to look for any correlation with vitamin B 12 individual values for the whole population. There was no correlation with the vitamin B 12 level, except for TTR (next section).

Vitamin B12 Status and Clinical Situation

None of the patients had anemia or macrocytosis or showed clinical evidence of B~2 deficiency. The vitamin B 12 and analog status was analyzed as a function of the type of colon resection. The presence or absence of the ileocecal valve did not influence the B12 parameters. There was no difference between BI2 values for the groups defined by a positive (B~2 = 143 pmol/L [79- 257]) or a negative (B~2 = 124 pmol/L [90-232]) breath test for microbial overgrowth. The analog concentrations also did not differ.

The patients with cholestasis (n = 8) had elevated levels

PARENTERAL NUTRITION AND VITAMIN B12 317

of GOT (39.5 IU/L [26.4-66.2], P < 0.0025), GPT (35.5 IU/L [21.9-69.0], (P < 0.030), AP (178 IU/L [120-335], (P < 0.001), plasma gamma-GT (206 IU/L [118-381], P < 0.0005), and total bilirubin ( 10.5 #mol /L [ 6.8-16.6 ], P < 0.015) compared with 18 IU GOT/L (15.2-23.9) , 21.5 IU GPT/L (16.6-30.9) , 85 IU AP/L (66-109) , 16.5 IU gamma- GT (10.0-48.2) , and 5.5 #tool total bilirubin (4 .4-7.2) in the other patients.

The patients with cholestasis also had increased plasma con- centrations of total corrinoids (vitamin B12 + analogs) (329 pmol/L [213-443] , P < 0.04) and apo-TCII (519 pmol B12/ L [452-591], P < 0.02), compared with 192 pmol/L (165- 265) for total corrinoids and 411 pmol B J L (329-510) apo- TCII in the other patients. There were no differences in the other vitamin B12 parameters, including B12 analogs.

The plasma TTR concentration was positively correlated with the total B12 concentration (P = 0.024; rho = 0.53), and with plasma folate (P = 0.004; rho = 0.73). The plasma vita- min B12, analogs, and folate concentrations were not correlated with the HS or MMA concentrations.

DISCUSSION

This'study evaluates the vitamin BI2 status of patients on TPN given four intramuscular injections of 1000 /~g vitamin BIz/y. The optimal method of delivering the supplement (intra- venous perfusion or intramuscular injection) and the dose rate are not clearly defined by published studies. 4-7 Some authors have used multivitamin solutions containing vitamin B12 plus ascorbic acid and trace minerals. But Herbert et al.22,23 sug- gested that vitamin B12 may be altered and its bioavallability decreased under these circumstances, although this has not been confirmed by others. 24 About 12-16% of patients on TPN gen- erally have low plasma vitamin B12 concentrations, despite in- travenous supplementation. 2 We therefore gave our patients vitamin B12 alone by intramuscular injection to minimize the chance of the vitamin being altered. The capacity of the liver to store the vitamin is great enough to make more frequent injections unnecessary. The frequency of low plasma levels was the same (20%) in patients given supplements intramuscu- larly as after intravenous supplementation. This suggests that the low plasma B12 of patients on TPN does not depend on the route by which it is given.

This study also shows that low plasma B~2 is not a true vitamin B12 deficiency, as none of the patients had elevated circulating MMA or HS. The levels of MMA and HS are con- sidered to be more sensitive indicators of B12 deficiency than the plasma B~2 level. Neither anemia nor macrocytosis was detected in any of these patients, and there was no clinical evidence of B12 deficiency. This discrepancy between plasma B12 concentration and metabolic parameters requires an expla- nation. The distribution of the plasma B12 concentration in a population of human subjects is known to be non-Ganssian and asymmetrical. It is thus possible that plasma B~2 concentrations of 85-95 pM may occur without there being any cellular deft- ciency~ Adenosyl-cobalamin and methyl-cobalamin are the two physiologically active forms of vitamin B12. Adenosyl-cobala- rain is required as cofactor for methylmalonyl-CoA mutase, which is necessary for the isomerization of methylmalonyl- CoA to succinyl CoA, and methyl-cobalamin is required as cofactor for methionine synthetase, which catalyzes the recy-

cling of HS methionineY The transformation of vitamin B12 into the active forms and the enzymatic reaction occur in the mitochondria for methylmalonyl-CoA mutase, and in the cyto- plasm for methionine synthetase. The intracellular pool of the enzymatic cofactors depends on the circulating vitamin B 12, but the intracellular storage level may be different. The circulating level of B12 may be subnormal while the tissue stores are normal in some TPN patients, since HS and MMA, depending on the intracellular vitamin B12 level, are in the normal range in these patients.

The transport of vitamin B12 and its analogs in the plasma was also studied by determining the amounts of saturated and unsaturated TCs, total corrinoids, and B i2 analogs. TPN patients suffering from cholestasis had elevated corrinoids, but this en- hanced circulating concentration was not due to cytolysis. It may be a result of the decreased enterohepatic circulation of corrinoids, as we have shown that B12 analogs are excreted mainly in the bile in humans) 2'26

The patients with subnormal plasma vitamin B12 levels had normal analog concentrations, and there was an inverse correla- tion between plasma BI2 and the percentage of analogs. The analog concentration was not obviously linked to the clinical picture known to lead to bacterial overgrowth. The concentra- tion of analogs was, however, increased in certain patients whose clinical condition did not imply bacterial overgrowth; for example, half of the patients having a jejunostomy also had elevated plasma analog levels.

The circulating TC concentration was not disturbed in our patients, except when the plasma B12 was 10w. The saturated TCs were generally low in these low B12 patients, while the total TCs (apo + holo) were normal. The low vitamin B12 level cannot be explained by a defect in circulating holo-TC or holo- Hc, as none of the individual values were subnormal in the "low B12" group.

The plasma protein status of the patients was normal, except that the TTR concentration was considerably decreased in pa- tients with low plasma B12 levels. This is unlikely to be due to a metabolic effect of Baz on protein synthesis, as the plasma HS level was normal in these patients.

Thus four of the 20 patients on TPN given 1000 #g Blz intramuscularly every 3 mo had low plasma B12 concentrations. One important point is that none of the patients with low vita- min B12 values had abnormal HS or MMA values. The 15% of patients that were borderline for HS had normal vitamin B12 values. This low vitamin B12 level did not correspond to a true vitamin B12 deficiency, as neither the metabolic nor the hematological parameters of these patients were abnormal, and this could predict a developing deficiency.

SUMMARY

The vitamin B12, HS, and MMA status of subjects on HPN were studied. When plasma vitamin B12 concentration was sub- normal, the values for the markers of vitamin B,2 deficiency (HS and MMA) were normal. As neither the metabolic nor the hematological parameters of these patients were abnormal, this could predict a developing deficiency.

ACKNOWLEDGEMENT

We wish to thank Mr. Philippe Grrard for technical assis- tance.

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