Serum levels of folic acid, vitamin B12, and iron in anemia of pregnancy

Serum levels of folic acid, vitamin

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and

iron in anemia of pregnancy FRED BENJAMI!',", M.D. FRANK A. BASSEN, M.D.

LEO M. MEYER. M.D. jamaica. New l'ork

[\toup of pn·gnant patients m a nearbY ,-oluntary hospital, 1 he Long lsland je\\JS!I Hospital, was also studied. In Yiew of previous reports, it was c·xpected that iron deficiE'ncy would be found to be the commonest cause and that folic acid deficiency would be responsible for mnst of the remainins{ small minority. This CO!ll-

I T H As been widely accepted that the commonest cause of anemia during pregnancy in the United States'; and Britain' is iron deficiency. In recent years more detailed studies of anemias during pregnancy have demonstrated that megaloblastic anemia is more common in the Occident than had hitherto been appreciated. Such reports have etnanated fron1 Britain~H-n, ''n Ireland/ 7 and Canada. 19 We l1ave bee-n unable to find any study of the incidence of megaloblastic anemia during pregnancy m the United States. The present investigation was carried out in order to determine the relative role played by deficiencies of folic acid, vitamin B 12 , and iron in the causation of anemia of pregnancy in a population served by a large City of ~ew York hospital. A study was made of all the anemias occurring during pregnancy in a four month period at the Queens Hospital Center, Jamaica, New York. Patients were invE'stigated, inter alia, by sc>nnn iron, folic acid, and vitamin Bl2 dE'tcrminations. A

inunication is presented because ;_ 1 : an ttrlcxpected high incidence of red!lccd sc·nuu

folic acid and vitamin B,~ kn'ls were fn11tld in this study (2) the incidence' of prcg·nann anemia associated with folic acid and ,·itamin B, c deficiency in this countrv is not really known, and (3) simultaneous studie' of serum folic acid, vitamin B 13 and iron in anemias of pregnancy have not, to om knowledge, been reported, and CPrtainlv nol in the !.\eographic area of our patient popldation.

Materials and methods All patit~nts who registered at the Antepartum Clinic of the Queens Hospital C:mtt~r during th<' 4 months, April through .Juh. I 965, wt~n· studied. Hemoe;lobin and hematocr·it dc'ierrninations and crythrocvte coun h were carried out on each patient. A group of unsekcted patients attending th(' Antt-partum Clinic of The Lon!.',· hland Jewish Hospital, :";ew Hvde Park. New York, wen· similarly investigated. Patients were divided into three groups: 11 ·, normal ---hemoglobin a bon· 12 Gm. pet <'l'llt. hematocrit above 15 per cent. or c>ryth-

From the Departments of Obstetrics and Gynecology of the State Unh•ersity of New York, Downstate Medical Center, Brookl)'n, The Queens Hospital Ctmter, ]am;;.ica, New York, The Lung !,land fewzsh Ho.rpital, Neu' Hyde Park, and the Divisior; of Hematology, The Long Island ]ewuh Hospital-Queens Hospital Center Affiliation, jamaica, New York. Supported by Grant CA 8279-02 from The National Cancer Institute, Department of Health, Education and Welfare, Bethesda, Jv/aryland.

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rocytes above 3,750,000 per cubic milliliter: ( 2 I mild anemia~hemoglobin between 10 and 12 Gm. per cent, hematocrit between :lO and 35 per cent, or erythrocytes between :),500,000 and (3,750,000 per cubic milliliter: and ( 3) moderate and severe anemia~ hemoglobin below 10 Gm. per cent, hematocrit below 30 per cent, or erythrocytes below 3,500,000. In all cases where anemia was found, the hemoglobin, hematocrit, and erythrocyte count were repeated. Iron and vitamin preparations and all other hematinirs were withheld until these determinations and the rest of the hematological work-up was completed. The reason for cla.-;sifying as mildly anemic patients with a hemoglobin between 10 and 12 Gm., a hematocrit between 30 and 35 per cent, or an erythrocyte count between 3,750,000 and 3,500.000 was based on the work of HollyY· 1.-•• 1 n He showed that despite the tremendous increase in plasma volume, hemoglobin, hematocrit, and erythrocyte counts remain normal during pregnancy, provided adequate iron and other blood-forming factors are present and bone marrow function is normal. In as many cases as possible from the Queens Hospital Center group, where hemoglobin was below 10 Gm. per cent, hematocrit below 30 per cent, or erythrocyte count below 3.500,000 per cubic milliliter, serum iron, serum folic acid, and serum B,e levels were determined. A total of 130 cases was submitted to all 3 of these serum determinations, and the study of these 130 patients comprises the main subject of this report. Serum iron determinations were made according to a modification of the method described by Peters and associates,"' and serum iron-binding capacity by Ressler and Zak's technique."~ Normal values for serum iron and total iron-binding capacity by these techniques in our laboratories arc 50 to 180 micrograms and 300 to 350 micrograms per 100 mi. serum, respectively. Folic acid assays were done by the microbiologic method described by Baker and associates,·1 based on Lactobacillus casci. Vitamin B," assays were carried out with Euglena gracilis by the method described by Ross." 3 Using these as-

311

says, the normal serum levels of folic acid in the nonpregnant individual are 6 to 24 millimicrograms per milliliter and of vitamin B," 200 to 900 rnicromicrograms per milliliter (values below 100 micrornicrograrm being regarded as indicating deficiency, and 100 to 200 micrornicrograms as indeterminate). However. it was considered necessary to obtain control values in a group of nonanemic pregnant women drawn from thP same population as the study group and who had not preyiously taken iron and/or folic acid in the current pregnancy. Accordingly, serum folic acid. vitamin B,~ and iron determinations were made on 30 randomlv selected pregnant patients whose red cell counts, hemoglobins, and hcmatocrits wen· normal (i.e., above) 3,7 50,000 per milliliteL 12 Gm., and 35 per cent, respectively). Thl' range of folic acid levels was 3.3 to 13 millimicrograms per milliliter, with a mean of 7.5, but in only 2 cases was the reading bl·low 4. In the r..ase of vitamin B,", the rang1• was 110 to 616 micromicrograms per milliliter, the mean was 203, and in no case was the level below 100. The range for serum iron was 40 to 220 micrograms per 100 mi. with a mean of 135 and in only 2 cases was the level below 50. Accordingly, for the purposes of this study, a deficiency was considered to exist when the serum leYels of iron, folic acid, and vitamin B1 c were below r,o micrograms per 100 rnl., 4 millimicrograms per milliliter, and 100 micromicrograms 1wr milliliter, respecti\·ely. These knJs of serum folic acid and vitamin B 1 " were the same as those found by Lowenstein and associates 18 as indicating deficiency in pregnancy. Results

During the course of the study, 1,052 antepartum patients were registered at the Queens Hospital Center. L'sing the aforementioned standards, 71.9 per cent of the patients in this group were found to be anemic, 49.1 per cent (516 cases) being mildly anemic, and 22.8 per cent (239 cases I having moderate or severe anemia. Of the latter 239 patients, 130 (randomly selected) were investigated by serum determinations of iron,

312

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Benjamin, Bassen, and Meyer \111.

folic acid, and vitamin B12· Four of these were in the first, 51 in the second, and 75 in the third trimester of pregnancy. The numbers of patients exhibiting low levels of serum iron, folic acid, and vitamin R, 2 , or com binat ions of d ecreased levels of th ese factors arc sh own in Fig . 1. The commonest deficiency was a combination of iron and folic acid, 51 cases (39.2 per cent) falling into this category. The next most common categories were a low serum iron value only, or a combination of low values of all three factors, 20 patients ( 15.4 per cent)

J. Ohst.

&

falling into each of these two groups. 'J'hirteen patients (10 per n·nt· had low \alw·s of iron and vitarnin .B 1 :.:- 1 1 B.5 per cent~ folic acid only. 5 (3.8 per l'f'nt · vitamin H" only, and in 2 patients ! I ."1 per cent foli< acid a nd vita min B, 2 levels wen· below no rm a l. with nor ma l iron va lw ·s. In 8 pa ti en t-:. serum \ alues of iron. folil' ar id and \·itamin B,:.: were all well within the nurmal rangt· two of these had sickle cell anemia ( SS! . in thn·e, the only rccognizab!t· ca11SL' fotmd fo1 the anemia was an infection pyclorwphritis and one had chronic glomcru lonephri ti:;. l r1

39.2% 50

40

NUMBER OF CASES

30

20

.,

15.'1%

15.'1%

IRON

IRON ONLY

10

IRON

+

FOLIC ACID

+

FOLIC ACID

LYT!l'•

IRON

+

VIT. Bl2

FOLIC ACID ONLY

VIT. Bl2 ONLY

FOLIC ACID

ALL NORMAL

+

VIT. Bl2

+

VIT. Bl2

Fig. I. Distribution of low serum levels of iron, folic acid, and vitamin B, in 130 palif'nts with anemia in pregnancy. 120

100

80 NUMBER OF 60 PATIENTS 40

20

FOLIC ACID

VIT. 812

Fig. 2. Total number of anemic patients with low levels of serum iron folic acid. and vitamin B, in 130 patients studied. ' ·

Vohune 9ti .\umlwr:l

the remaining three cases, no apparent cause was found. There was no response to parenteral iron, folic acid, and vitamin Bu, and there was a spontaneous remission after deliYery. Thesr casPs may have fallen into the category of "hypoplastic (refractory) anemia of pregnancy" described by Holly. 14 The total number of cases with low serum iron values (alone or in combination with other deficiencies\ was 104 (80 per cent '1; and the total numbers for low levels of folic acid and vitamin B 1 " were 8+ (64.6 per cent) and 40 1:~0.8 per cent), respectively (Fig. 2). The response of these patients to specific medication, and the incidence of obstetric complications in the Yarious groups compared with the nonanemic (control) patients is currently being observed and followed, and will be the subject of a future report. Among 150 patients studied at The Long Island Jewish Hospital, 68 per cent were found to he mildly anemic and 5 per cent exhibited moderate to severe anemia using the same criteria applied to the City Hospital patients. Comment

The incidence of anemia in the group of 1.052 consecutive unselected pregnant patients in a City Hospital and 150 in a voluntary hospital is similar to that generally reported in the United States. However, the frequency of subnormal levels of folic acid and of vitamin B 1 ~ found in this study was surprisinp;. Although the high incidence of megaloblastic anemia in pregnancy in the Orient is well-known, it is only in recent vears it has been appreciated that such anentia is not uncommon in the Occident. The incidence of megaloblastic anemia in pregnancy in the United States has not been reported but recent figures indicate that it ocems in 2 to 4 per cent of pregnant women in Hritain,\•-n. ~u Ireland," and Canada. 19 The present study showed that moderate and sewre anemias in pregnancy in a region of NPw York City are associated with subnorma! serum folic acid levels in as many as 64.6 per cent of cases. Based on the total

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number of pregnant women among whom the cases occurred, 8+ out of 1,052 exhibited folic acid deficiency, an incidence of 7.g per cent Since folic acid levels were not clone in the cases of mild anemia and in somt· of the patients with severe anemia, this incidence is a conservative one. Apart from any geographic or other factors that 111ay account for it, this finding of such a hi~h frequency could be due to the fact that in other reports the diagnosis was made by peripheral blood and marrow smears. or by FIGLU excretion studies 11 whereas in the present series, serum folic acid determinations were made, which is a more direct and sensitive index of deficiency. A striking finding in the present study was the high incidence of severdy depres:wd serum vitamin B 12 le\·els in a group of l :lO unselected cases of anemia of pregnann'. Even though the normal range of St'n1m vitamin B1 ~ is 200 to 600 millimicrograms per milliliter, .'l9.8 pet· cent (J.O of the BO cases) had vitamin B ~~ levels below 100 millimicrograms per milliliter. This strict criterion for subnormal vitamin B 1 " levt>ls was used because of the findings in onr control studies and because of reports that tlw values decrease towards the end of normal pregnancy. Baker and associates." in a stud\' of 51 pregnant patirnts found a m(•dian serum vitamin B 12 level of 193 micromicrograms per milliliter. The role of \·itamin B,c deficiency as a cause of anemia in pregnane\· has been regarded as insignificant because: ( 1) in most of the reported cases. megaloblastic anemia does not respond to dtatuin B 1 " therapy, but responds well to folic acic\. 1 s, 1\l. ~· ( 2) the liver normally rontaim a resen'c of \-itamiu B, ~ sufficient for se\ eral years and the average daily requirement j,; only about 1 meg.: whereas the body stores of folic acid and its clcrivath·es are less extensive or durable than those of \·itantin B,~. and symptoms of deficiency may comntence within se\·eral months of dietary deficiency; and ( 3) the demand for folic acid by the rapidly developing fetus is overwhelming. Howe,·er, the question as to whether low serum vitamin B 1" is etiologically related to

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Benjamin, Bassen, and Meyer

anemia of pregnancy has yet to be clarified. Since in most cases the vitamin B1~ deficiency is associated with iron and folic acid deficiency, and therapy with iron and folic acid alone will result in at least partial remission, the contribution of vitamin Bt~ lack may have been missed in many series. A low serum Yitamin B 1 " w:1s associated with megaloblastic erythropoiesis in a high 1)!'0portion of cases reportrd hy Lowenstein and associatcs 1 ' , and they felt that this probably indicated true vitamin B 1 ~ deficiency. The response of our cases to specific Yitamin B1:: therapy. cmrently Hnder re\iew. may help to clarify its etiologic role. It is clear, howen~r, from the present study, that in only 27 per cent of the casl's of anemia of prrgnancy was the deficiency due to one factor only (iron 15.4 per cent, folic acid 8.5 per cent. and Yitamin H1 " 3.H percent). In 66.1 per cent there was mor(' than one deficiency i the commonest being iron and folic acid. 39.2 per cent 1. The patients were tn·atcd specifically according to the specific defi.ciency found. This was done to determine whether the hrmoglobin deficit could be corrected by supplying, in therapeutic doses. the hematinic which was lowered according to ::;ennn determinations. At the same timt>, an asse~sment is being mad<~ as to whether the incidence can be lowered of ob:stctric and pediatric complications attributed to folic acid deficiency (e.g., abruptio placentae,\. '·· 11 · ~'"· 1 ~ fetal malformations,u and abortions"!. This concurrent study will be reported later. It may help to determine whether there is justification for placing· pregnant women not only on prophylactic iron n1('dication, but also on folic acid and vitamin B," in doses higher than those giwn in multivitamin capsul~s usually prescribed for expectant mothers. It has been shown by Young- and associates"'· that the administration of 25 micrograms of Yitamin nl~ oraiiy, when combinc·d with a daily dose of 5 mg. folic acid, prevented the drop in serum H 1 " that usuallv occurred in preg·nancy, (whereas 100 micrograms vitamin B 1 ~ alone did not result in the maintenance of

\111.

such satisfactory serum lPYPb

.r

i

li

,,,

1l

1t

argument al;·ainst sucb rout inc )·I' :.nipl :· ,, ' oi fo\i( acid. is, of course. t\tat i: q,:l\. 11 inl'ipient cast's of true Acldisunia1t ,wtlllc · ., anemia. and may indeed precipit:. ical manifestations of this dist\bl'. l low1 \' r. the high incidence of folic ;1cid
Summary

1. The incidence of various grades ot ant·in prqrnancy in thl' populal ion sen t·d hv a largt• citv hospital and a voluntarv ho~­ pital in ~cw York was studied. Sr·nmt lc\els of irnn. folic acid, and vitamin B '" wen~ dl'tt·nnined in 1:~0 rases prior tn, and followint:·. specific tlwrapy. Similar dert'rminations wen· llladc in nonanemic cont wl subjects. 2. Eightv per cent of the :mcmic patir.·nb had lr>\\ serum iron \·a lues ! alon<' or in comhinatiDn with other deficiencirs · ·. tln~ lTSJW( tiw inridt·nn·s of folic acid and ,·it.alllin B,c ddicicnn were 64·.6 per cent and :W.8 per cent. lu only ?.7 per cent ()\ the n Ed· per cent, folic acid :l) pt·r crnt, and \itarnin B 1 :U! per cent). In 6f).l per cent there '.LIS ntow than one dPlicif'ncv. :\. Tlw r.onnnonest ane111ia w~ts tltrtt found in as~ocia tion with low lt·\·1'1' nf both iron and fo!il' :tcid (39.2 per tt'llt . :"Jt•xt 111m;t frequl'llt was a low serum iron \ a!tw only l.'i.-J. per cent·, or a cotnhination of lm\ \'f iron. folic acid. :111d Yitamin B, 115.+ per crntJ. +. The significance of these lindings i-: di.~­ cussed irom the point of \ icw of the etiolog~ of ancnria in prcgnann· in tltr: population studied. and its pre\·ention in the cour~e of antepartum care. t uia

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REFERENCES

1. Editorial: Brit. M. J. 2: 1422, 1963. Editorial: Obst. & Gynec. Surv. 15: 22, 1960. :\. Baker, H., Erdberg, R., Pasher. I., and Sobotka, H.: Proc. Soc. Exper. Biol. & Med. 94: 513. 1957. +. B.tkf-r, H., Herbert, V., Frank, 0 .. Pasher. I., Hutncr. S. H., Wasserman L. R. and Sobotka, H.: Clin. Chern. 5: 275, 1959. 5. Coyle, C., and Geoghegan, F.: Proc. Roy. Sue. Med. 55: 764. 1962. 6. Eastman, N. J., and Hellman. L. M., editors: Williams Obstetrics, ed. 12. New York, 1961. Appldon-Century-Crofts, Inc., p. 82+. 7. Eastman, N. J., and Hellman, L. M .. editors: Williams Obstetrics, ed. 12, N cw York, 1961, Appleton-C~ntury-Crofts. Inc .. p. 825. 8. Fisher . .M .. and Biggs, R.: Brit. 1\L J. 1: 385, ')

1955. 9. Forshaw, J. W. B., Thelwali-Jones, A., Chisholm, W. H., and McGinley, W. K.: J. Obst. & Gynaec. Brit. Emp. 64: 255, 1957. 10. Giles, C .. and Shuttclw"rth, E. M.: Lancet 2: 13+1, 1958. II. Hibbard. B. M.: J. Ohst. & Gynaec. Brit. C:ornrn. 71: 5~9, 19M.

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12. Hibbard, B. M., and Hibbard, E. D.: Brit. M. J. 2: 1430, 1963. 13. Holly, R. G.: Bull. Univ. Minnesota and Minnesota .M. Found. 25: 13+. 1953. 1+. Holly, R. G.: Obst. & Gynec. 1: 535, 1953 15. Holly, R. G: Obst. & Gynec. 2: 119, 1953. 16. Holly, R. G.: Obst. & Gynec. 4: 562, 1955. 17. Hourihane, R., Doyle, C. V., and Drury. M 1.: J. Irish M.A. 47:1,1960. 18. Lowenstein, L., Hsieh, Y. S., Brunton, L .. D<· Lceuw, N. K. M., and Cooper, B. A.: In Proceedings of 8th Congress of European Society of Hematology, Vienna, 1961, Part II, p. 337. 19. Lowenstein, L., Pick, C., and Philpet, N. W.: AM. J. 0BST, & GY!'EC. 70: 1309, 1955. ~0. MacKenzie, A., and Abbott. J.: Brit. M . .J. 2: 1114, 1960. ::1. Peters, T., Giovanniello, T. J., Apt, L., and Ross, J. E.: J. Lab. & Clin. Med. 48: 27-l. 1956. 2:!. Ressler, M., and Zak: B.: Am. J. Clin. Path. 30: 87' 1958. 2:{. Ross, G. I. N.: J. C:lin. Path. 5: 250, 1952. 2+. Thompson, R. B.: Lancet 1: 1171, 1957. 25. Young, J. E., Barrows, C., Okuda, K., and Chow. B. F.: Obst. & Gynec. 14: H:9. 1959.