Human Reproduction

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Human Reproduction, Vol. 15, No. 1, 224-226, January 2000
© 2000 European Society of Human Reproduction and Embryology

Evaluation of low serum vitamin B₁₂ in the non-anaemic pregnant patient

J. Pardo^1,3, Y. Peled¹, J. Bar¹, M. Hod¹, B.A. Sela², Z.Ben Rafael¹ and R. Orvieto¹

¹ Department of Obstetrics and Gynecology, Rabin Medical Center, Beilinson Campus, Petah Tiqva, and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, and ² Institute of Chemical Pathology, Sheba Medical Center, Tel Hashomer, Israel

	Abstract

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Low serum vitamin B₁₂ concentrations in pregnancy may not indicate true megaloblastic anaemia. In the present study we compared biochemical indices of vitamin B₁₂ deficiency (serum homocysteine and urine methylmalonic acid) in non-anaemic pregnant women with and without low serum vitamin B₁₂ concentrations. The groups were matched for age, parity and gestational age. No differences were found, and all values were within normal range. These results suggest that the measurement of low serum B₁₂ concentrations in pregnant women should be followed by analysis at the biochemical level before vitamin B₁₂ injections are started.

Key words: homocysteine/methylmalonic acid/pregnancy/vitamin B₁₂

	Introduction

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Anaemia is a common complication of pregnancy, with an incidence of 20–80%, depending largely on socio-economic status (Puolakka et al., 1980). The vast majority of affected women have iron deficiency; more rarely, anaemia during pregnancy is due to a deficiency in vitamin B₁₂. Suspected anaemia, especially of the macrocytic type, is confirmed by measurement of serum folate and vitamin B₁₂ concentrations. However, several studies have shown that up to 20% of normal pregnancies are associated with lower than normal serum vitamin B₁₂ concentrations (Ball and Giles, 1964), which increase to normal within hours after delivery (Mollin et al., 1976).

Vitamin B₁₂ and folic acid catalyse the remethylation of homocysteine (Hcy) to methionine. Hyperhomocysteinaemia may be caused by a deficiency of cobalamin or folate metabolites. Nutrition also plays a significant role: a diet rich in fresh vegetables and fruits as well as vitamin supplementation has been associated with lower Hcy concentrations (Miner et al., 1996). Furthermore, studies have shown that methylmalonyl-CoA, a metabolite of various amino acids, accumulates when its mutase is blocked by deficiencies of its vitamin B₁₂ co-factors or in the presence of congenital abnormalities of the mutase (Cox and White, 1962).

The measurement of serum concentrations of methylmalonic acid (MMA) and Hcy metabolites related to vitamin B₁₂ deficiency was introduced in 1990 to establish vitamin B₁₂ deficiency at the biochemical level (Allen et al., 1990). Using a combination of serum metabolite concentrations, serum vitamin concentrations, clinical characteristics, and the response to therapy, it has been demonstrated that every vitamin B₁₂-deficient patient who will respond to therapy has an elevation of MMA and/or Hcy, whereas findings of normal concentrations of both metabolites rule out clinical vitamin B₁₂ deficiency (Stabler et al., 1996). In line with these findings, Metz et al. suggested that the drop in vitamin B₁₂ serum concentration in pregnancy does not reflect B₁₂ deficiency at the biochemical concentration, and that to establish true B₁₂ deficiency in pregnancy, the concentration of serum Hcy (but not MMA) should be used (Metz et al., 1995).

The aim of the present study was to assess vitamin B₁₂ concentrations in normal pregnant women and to examine whether low vitamin B₁₂ concentrations are associated with changes in the biochemical indices of vitamin B₁₂ deficiency, erythrocyte count or erythrocyte morphological indices, as reflected by erythrocyte mean corpuscular volume (MCV).

	Materials and methods

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The study population included 147 patients attending the ambulatory High Risk Pregnancy Clinic of Rabin Medical Center, Israel who met the following criteria: (i) pregnancy duration 20–30 weeks; (ii) normal diet; (iii) singleton pregnancy; (iv) no history of anaemia before the present pregnancy; (v) no evidence of malabsorption. Patients with diseases known to cause anaemia (renal disease, thalassaemia, etc.) or maintaining a vegetarian diet were excluded.

All study participants underwent a complete blood count and serum B₁₂ determination. Those with low B₁₂ concentrations (100 pmol/l) were designated the study group and compared for plasma Hcy and urine MMA concentrations with an equal number of subjects with normal serum B₁₂ concentrations (>100 pmol/l) matched for age, parity and gestational age (control group).

Hcy was measured in plasma samples that were separated from erythrocytes shortly after blood was drawn. Plasma was reduced by sodium borohydride, followed by treatment with the fluorescent probe monobromobimane. Separation of Hcy by HPLC column and fluorescent detection was performed as previously described (Ueland et al., 1993). Urine analysis for MMA was performed by thin-layer chromatography on silica gel G plate, followed by chromatography in amyl acetate–glacial acetic acid H₂O and Fast Blue B reagent. Further assessment of MMA concentrations was achieved by capillary gas chromatography-mass spectrometry (Stabler et al., 1986).

Normal values for plasma Hcy and urine MMA concentrations are 4–11 µmol/l and <9 mg/g creatinine respectively. Values are presented as mean ±SD. The study groups were compared with Student's t-test; P-values less than 0.05 were considered significant.

	Results

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A below-normal value of serum B₁₂ was found in 32 of the 147 patients tested (study group). The results for the study and control groups are shown in Table I. No significant between-group difference was noted for mean haemoglobin and mean corpuscular volume (MCV) on complete blood count. There were no cases of MCV >100 fl in either group. Serum Hcy and urine MMA were also comparable between the two groups, and were within normal range.

View this table: [in this window] [in a new window]   Table I. Patient characteristics and laboratory results

 

	Discussion

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Decreased haemoglobin concentration is a frequent finding in pregnancy, but it seldom mandates a full laboratory evaluation, as iron and folic acid supplementation are usually sufficient to avoid anaemia. Furthermore, the diagnosis of specific anaemic deficiencies by laboratory methods is more complicated during pregnancy, because pregnancy may alter `normal' values. Accordingly, the decrease in serum B₁₂ concentrations noted by several studies during pregnancy does not reflect a true tissue depletion of B₁₂ (megaloblastic anaemia) (Ball and Giles, 1964). In a recent comparison of 50 pregnant patients with low serum B₁₂ concentrations and 25 pregnant patients with normal concentrations (Metz et al., 1995), no significant difference in either serum MMA or total Hcy was reported between the groups. These findings are supported by the present study. Of our 147 randomly selected pregnant patients, 21.7% had low serum B₁₂ concentrations, a rate comparable to the 20% reported (Ball and Giles, 1964), but none of them had biochemical indices of B₁₂ deficiency.

Vitamin B₁₂ deficiency may be associated with various pregnancy complications. A derangement of methionine-Hcy metabolism has been noted in ~20% of cases of neural tube defects and recurrent spontaneous abortion (Eskes et al., 1996). However, in a later study (Sutterlin et al., 1997), no alteration could be demonstrated in serum folic acid and vitamin B₁₂ concentrations in women with unexplained recurrent spontaneous abortions. Pre-eclampsia may also be related to high plasma Hcy concentrations (Laivuori et al., 1999), with a direct relationship between the increase in Hcy and the severity of the pre-eclampsia. In the same study, though vitamin B₁₂ concentrations were found to be reduced in pre-eclampsia, there was no correlation between plasma Hcy and vitamin B₁₂. Finally, vitamin B₁₂ apparently plays an important role in the metabolism of the nervous system (Healton et al., 1995). Using cranial magnetic resonance imaging, a retardation of myelination in a child's nervous system due to maternal dietary vitamin B₁₂ deficiency has been demonstrated (Lovblad et al., 1997).

The usual means for delineating vitamin B₁₂ malabsorption secondary to intrinsic factor deficiency versus other malabsorption factors (Schilling test) is contraindicated in pregnancy because of the risk of radiation exposure. Therefore, therapy is usually instituted empirically in pregnant patients with low serum vitamin B₁₂, and diagnostic testing is delayed until the postpartum period (Campbell, 1995). This practice, however, may lead to overtreatment of pregnant patients without true vitamin B₁₂ deficiency. Our results, together with others (Metz et al., 1995), suggest that serum Hcy and urine MMA be measured in every pregnant patient with low serum vitamin B₁₂ before treatment with vitamin B₁₂ injections is instituted.

In conclusion, routine serum B₁₂ testing is unnecessary in pregnancy. However, if low serum vitamin B₁₂ is demonstrated, for example, in cases of anaemia refractory to iron and folic acid treatment, it should be followed by MMA and Hcy evaluation.

	Acknowledgments

 
The authors wish to thank Mrs Gloria Ginzach and Mrs Marian Propp for their editorial and secretarial assistance.

	Notes

 
³ To whom correspondence should be addressed at: Department of Obstetrics and Gynecology, Rabin Medical Center (Beilinson Campus), Petah Tiqva 49100, Israel

	References

Top Abstract Introduction Materials and methods Results Discussion References

 
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Ball, E.W. and Giles, C. (1964) Folic acid and vitamin B₁₂ levels in pregnancy and their relation to megaloblastic anemia. J. Clin. Pathol., 17, 165–174.

Campbell, B.A. (1995) Megaloblastic anemia in pregnancy. Clin. Obstet. Gynecol., 38, 455–462.[ISI][Medline]

Cox, E.M. and White, A.M. (1962) Methylmalonic acid excretion: an index of vitamin B₁₂ deficiency. Lancet, ii, 853–856.

Eskes, T.K.A.B., Nelen W.K.D.M., van der Molen, E.F. et al. (1996) Reproductive failure and hyperhomocysteinaemia – the role of folic acid. J. Fertil. Res., 10, 14–17.

Healton, E.B., Savage, D.G., Brust, J.L.M. et al. (1995) Neurologic aspects of cobalamin deficiency. Medicine, 70, 229–245, Baltimore.

Laivuori, H., Kaaja, R., Turpeinen, U. et al. (1999) Plasma homocysteine levels elevated and inversely related to insulin sensitivity in preeclampsia. Obstet. Gynecol., 93, 489–493.[Abstract/Free Full Text]

Lovblad, K., Ramelli, G., Remonda, L. et al. (1997) Retardation of myelination due to dietary vitamin B₁₂ deficiency: cranial MRI findings. Pediatr. Radiol., 27, 155–158.[ISI][Medline]

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Puolakka, J., Janne, O., Pakarinen, A. et al. (1980) Serum ferritin in the diagnosis of anemia during pregnancy. Acta Obstet. Gynecol. Scand. (Suppl), 95, 57–65.

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Submitted on June 15, 1999; accepted on September 17, 1999.

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This Article Abstract FREE Full Text (PDF ) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Add to My Personal Archive Download to citation manager Search for citing articles in: ISI Web of Science (17) Request Permissions Google Scholar Articles by Pardo, J. Articles by Orvieto, R. Search for Related Content PubMed PubMed Citation Articles by Pardo, J. Articles by Orvieto, R. Social Bookmarking          What's this?

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Copyright © 2008 European Society of Human Reproduction and Embryology

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