Effect of Oral Administration of Fe 2 +-Fumarate on Erythrocyte Profile and Growth Rate of Suckling Piglets

Svoboda M., J . Drábek: Effect of Oral Administration of Fe2+-Fumarate on Erythrocyte Profile and Growth Rate of Suckling Piglets. Acta Vet. Brno 2002, 71: 217–222. The aim of this study was to investigate efficiency of oral administration of Fe2+-fumarate in form of paste in preventing iron deficiency anaemia of suckling piglets. Piglets in group 1 (n = 20) were given 200 mg Fe2+-fumarate on days 6 and 11 after birth. Piglets in group 2 (n = 20) were given 200 mg Fe3+-dextran i.m. on day 3. A group of piglets (group 3, n = 10) treated with Fe3+dextran on day 21 was included in the study. All piglets had free access to weaning pellets (124 mg Fe/kg). Haemoglobin concentration (Hb), packed cell volume (PCV), erythrocyte count (RBC), MCV, MCH and MCHC, body weight (BW) and daily weight gain (DWG) were determined. On day 6, haemoglobin concentration and packed cell volume were significantly lower in group 1 compared to group 2 (P < 0.05). On days 14, 21, 28, and 35 no significant differences in examined indices were found between group 1 and group 2 and all indices were comparable to physiological values. Haematological indices of group 3 were characteristic for hypochromic anaemia. Anaemia in group 3 had detrimental effect on growth rate of piglets. Supplementation of Fe to piglets with Fe2+-fumarate under conditions of this trial was efficient in preventing anaemia and was comparable to i.m. administration of Fe3+-dextran. Anaemia, iron, haemoglobin, Fe3+-dextran, Fe2+-fumarate The piglet is born with an iron reserve of approximately 50 mg (Venn et al. 1947). During the first week, the piglet doubles its body mass from 1.5 to 3 kg. At the same time the plasma volume expands by 30 % (Jain 1986). In order to synthetise haemoglobin sufficient for the piglet not to develop anaemia, 7–10 mg Fe per day is required (Venn 1947). The sow ́s milk, however, provides the piglet with approximately 1 mg Fe per day only (Kleinbeck 1999). Suckling piglets with no extra iron supplementation develop iron deficiency after 14 days post partum (Zimmermann 1995). An early supply of iron is therefore essential for them. Parenteral or oral preparations for iron supply are available. Bivalent iron Fe2+ is up to 16 times better absorbed than trivalent iron Fe3+. Only preparations with bivalent iron can be recommended for oral iron therapy (Dietzfelbinger 1987). According to Thoren-Toll ing and Jonsson (1977) the distribution of stainable iron in lymph nodes, liver and spleen seven days after intramuscular injection of iron dextran in newborn piglets was comparable to that for oral administration (iron dextran) at that stage of the experiment. Oral Fe is available for haemoglobin synthesis sooner after administration than injected Fe3+–dextran (Framstad et al. 1997). Oral administration of amino acid–chelated iron to pregnant sows in preventing piglet anaemia has not been successful (Egel i et al. 1998a). In herds where water-based iron is used, there is a large proportion of anaemic piglets. It seems that as long as the piglets get enough milk, they are less interested in drinking the iron solution (Jorgensen 2000). Individual administration of oral Fe preparations is thus recommended. Fe2+-fumarate-based products ACTA VET. BRNO 2002, 71: 217–222 Address for correspondence: MVDr. Martin Svoboda, Ph.D. University of Veterinary and Pharmaceutical Sciences Palackého 1-3, 612 42 Brno Czech Republic Phone: + 420 5 4156 2433 Fax: +420 5 748841 E-mail: svobodama@vfu.cz http://www.vfu.cz/acta-vet/actavet.htm for oral administration have been used successfully in human medicine (Davidsson et al. 2000; Lares-Assef et al. 1999). Effect of oral Fe2+-fumarate administration on somatic growth and selected haematological indices in piglets under experimental conditions were evaluated by Kotrbáãek (2001). The aim of this study was to investigate the effect of oral administration of Fe2+-fumarate in the form of paste on erythrocyte profile and growth intensity of suckling piglets as compared with intramuscular administration of Fe3+–dextran under field conditions. Materials and Methods Experimental design The study took place in a commercial herd. The litters were kept with the sows in farrowing crates on plastic slatted floor until weaning. The piglets were given no medical treatment, and no death loss was recorded. All piglets were individually tattooed a number in the ear. Altogether 50 piglets (6 litters) were divided randomly into 3 groups (split litters). Group 1 had 20 piglets, group 2 had 20 piglets and group 3 had 10 piglets. The feeding frequency and nutrient concentration of the milk are the major factors underlying the rate of gain during the suckling period (Holub 1982). The design involving split litters minimised the effect of the sow. On day 6 of life, the piglets in group 1 were given each 200 mg of Fe2+-fumarate orally. The same dose was repeated on day 11. Fe2+-fumarate was administered in the form of paste. The paste contained Fe2+-fumarate 13.2 g in 40 g, Enteroccocus faecium 80. 109 CFU, Retinoli acetas 80 000 IU, Colecalciferolum 8000 IU, Tocoferoli alfa acetas 400 mg. The piglets in group 2 were injected with 200 mg Fe3+–dextran (i.m.) on day 3 of life. Piglets in group 3 were given 200 mg Fe3+–dextran (i.m.) on day 21 of life. Group 3 served as negative control. Weaning pellets (Fe 124 mg·kg-1) were offered to all litters ad libitum (from day 7-35). Piglets in all litters were weaned on day 28. Sampling Blood (1.5 ml) was collected from the piglets on days 6, 14, 21, 28 and 35 by puncture of the cranial vena cava, using ethylenediaminetetraacetic acid (EDTA ) as anticoagulant. Haematological examinat ion On the day of sampling, blood was analysed for the following indices: haemoglobin concentration (Hb), packed cell volume (PVC), erythrocyte count (RBC), mean corpuscular volume (MCV), mean corpuscular haemoglobin concentration (MCHC), mean corpuscular haemoglobin (MCH). Haemoglobin concentration was measured spectrophotometrically using cyanomethaemoglobin method. Packed cell volume was measured using standard capillary tubes and centrifugation in a microhematocrit Janetzki TH12 centrifuge. Erythrocytes were counted in Bürker ́s chamber using Haym ́s solution. MCV, MCH and MCHC were calculated. Body weight and weight gain The piglets were weighed (BW) at birth (day 1) and on days 6, 14, 21, 28, and 35. Mean daily weight gain (DWG) of piglets in each group after one week was calculated. Stat is t ical evaluat ion The results were statistically evaluated by analysis of variance (ANOVA). All results are presented as mean values and standard deviations of each index. Results Haemoglobin concentrat ion (Hb, Fig. 1) Hb in group 2 on day 6 was higher (P < 0.05) compared to groups 1 and 3. Between days 6 and 14, Hb in groups 1 a 2 increased (P < 0.01). Hb in group 3 decreased between days 6 and 21 (P < 0.05). Hb in group 3 on days 14 and 21 was lower (P < 0.01) than in groups 1 and 2. After administration of Fe-dextran in group 3 on day 21, an increase was observed on day 28 (P < 0.01) reaching values of groups 1 and 2. In group 1, Hb decreased between days 21 and 28 (P < 0.05). Differences in Hb between groups 1 and 2 on days 14, 21, 28 and 35 were not significant. Packed cel l volume (PCV, Fig. 2) On day 6, PCV in group 2 was higher (P < 0.05) compared to groups 1 and 3. Between days 6 and 14, PCV in groups 1 and 2 increased (P < 0.01). PCV in groups 1 and 2 on days 14 and 21 was higher than in group 3 (P < 0.01). PCV in group 1 decreased between days 218

The piglet is born with an iron reserve of approximately 50 mg (Venn et al. 1947).During the first week, the piglet doubles its body mass from 1.5 to 3 kg.At the same time the plasma volume expands by 30 % (Jain 1986).In order to synthetise haemoglobin sufficient for the piglet not to develop anaemia, 7-10 mg Fe per day is required (Venn 1947).The sow´s milk, however, provides the piglet with approximately 1 mg Fe per day only (Kleinbeck 1999).Suckling piglets with no extra iron supplementation develop iron deficiency after 14 days post partum (Zimmermann 1995).An early supply of iron is therefore essential for them.Parenteral or oral preparations for iron supply are available.Bivalent iron Fe 2+ is up to 16 times better absorbed than trivalent iron Fe 3+ .Only preparations with bivalent iron can be recommended for oral iron therapy (Dietzfelbinger 1987).According to Thoren-Tolling and Jonsson (1977) the distribution of stainable iron in lymph nodes, liver and spleen seven days after intramuscular injection of iron dextran in newborn piglets was comparable to that for oral administration (iron dextran) at that stage of the experiment.Oral Fe is available for haemoglobin synthesis sooner after administration than injected Fe 3+ -dextran (Framstad et al. 1997).Oral administration of amino acid-chelated iron to pregnant sows in preventing piglet anaemia has not been successful (Egeli et al. 1998a).In herds where water-based iron is used, there is a large proportion of anaemic piglets.It seems that as long as the piglets get enough milk, they are less interested in drinking the iron solution (Jorgensen 2000).Individual administration of oral Fe preparations is thus recommended.Fe 2+ -fumarate-based products for oral administration have been used successfully in human medicine (Davidsson et al. 2000;Lares-Assef et al. 1999).Effect of oral Fe 2+ -fumarate administration on somatic growth and selected haematological indices in piglets under experimental conditions were evaluated by Kotrbáãek (2001).
The aim of this study was to investigate the effect of oral administration of Fe 2+ -fumarate in the form of paste on erythrocyte profile and growth intensity of suckling piglets as compared with intramuscular administration of Fe 3+ -dextran under field conditions.

Experimental design
The study took place in a commercial herd.The litters were kept with the sows in farrowing crates on plastic slatted floor until weaning.The piglets were given no medical treatment, and no death loss was recorded.All piglets were individually tattooed a number in the ear.Altogether 50 piglets (6 litters) were divided randomly into 3 groups (split litters).Group 1 had 20 piglets, group 2 had 20 piglets and group 3 had 10 piglets.The feeding frequency and nutrient concentration of the milk are the major factors underlying the rate of gain during the suckling period (Holub 1982).The design involving split litters minimised the effect of the sow.On day 6 of life, the piglets in group 1 were given each 200 mg of Fe 2+ -fumarate orally.The same dose was repeated on day 11.Fe 2+ -fumarate was administered in the form of paste.The paste contained Fe 2+ -fumarate 13.2 g in 40 g, Enteroccocus faecium 80. 10 9 CFU, Retinoli acetas 80 000 IU, Colecalciferolum 8000 IU, Tocoferoli alfa acetas 400 mg.The piglets in group 2 were injected with 200 mg Fe 3+ -dextran (i.m.) on day 3 of life.Piglets in group 3 were given 200 mg Fe 3+ -dextran (i.m.) on day 21 of life.Group 3 served as negative control.Weaning pellets (Fe 124 mg•kg -1 ) were offered to all litters ad libitum (from day 7-35).Piglets in all litters were weaned on day 28.

Sampling
Blood (1.5 ml) was collected from the piglets on days 6, 14, 21, 28 and 35 by puncture of the cranial vena cava, using ethylenediaminetetraacetic acid (EDTA ) as anticoagulant.

Haematological examination
On the day of sampling, blood was analysed for the following indices: haemoglobin concentration (Hb), packed cell volume (PVC), erythrocyte count (RBC), mean corpuscular volume (MCV), mean corpuscular haemoglobin concentration (MCHC), mean corpuscular haemoglobin (MCH).Haemoglobin concentration was measured spectrophotometrically using cyanomethaemoglobin method.Packed cell volume was measured using standard capillary tubes and centrifugation in a microhematocrit Janetzki TH12 centrifuge.Erythrocytes were counted in Bürker´s chamber using Haym´s solution.MCV, MCH and MCHC were calculated.

Body weight and weight gain
The piglets were weighed (BW) at birth (day 1) and on days 6, 14, 21, 28, and 35.Mean daily weight gain (DWG) of piglets in each group after one week was calculated.

Statistical evaluation
The results were statistically evaluated by analysis of variance (ANOVA).All results are presented as mean values and standard deviations of each index.

Results
Haemoglobin concentration (Hb, Fig. 1) Hb in group 2 on day 6 was higher (P < 0.05) compared to groups 1 and 3. Between days 6 and 14, Hb in groups 1 a 2 increased (P < 0.01).Hb in group 3 decreased between days 6 and 21 (P < 0.05).Hb in group 3 on days 14 and 21 was lower (P < 0.01) than in groups 1 and 2. After administration of Fe-dextran in group 3 on day 21, an increase was observed on day 28 (P < 0.01) reaching values of groups 1 and 2. In group 1, Hb decreased between days 21 and 28 (P < 0.05).Differences in Hb between groups 1 and 2 on days 14, 21, 28 and 35 were not significant.
Packed cell volume (PCV, Fig. 2) On day 6, PCV in group 2 was higher (P < 0.05) compared to groups 1 and 3. Between days 6 and 14, PCV in groups 1 and 2 increased (P < 0.01).PCV in groups 1 and 2 on days 14 and 21 was higher than in group 3 (P < 0.01).PCV in group 1 decreased between days 21 and 28 (P < 0.05).PCV in group 3 increased between days 21 and 28 (P < 0.01) and on days 28 and 35 was higher (P < 0.01) than in groups 1 and 2. Differences in PCV between groups 1 and 2 on days 14, 21, 28 and 35 were not significant.
Red blood cell count (RBC, Fig. 3) RBC in group 3 on day 21 was lower compared to groups 1 and 2 (P > 0.01).RBC increased between days 21 and 28 in group 3 (P > 0.01).Differences in RBC between groups 1 and 2 were not significant.
Daily weight gain (DWG, Fig. 4) Between days 28 and 35, DWG in groups 1 (P < 0.01) and 2 (P < 0.01) increased.DWG in group 3 on days 14, 21 and 28 was lower compared to groups 1 and 2 (all, P > 0.01).DWG in group 3 increased between days 21 and 35 (P > 0.01), but body weight remained to be lower than in groups 1 and 2 (P > 0.01).No significant differences in DWG between groups 1 and 2 were found.
Body weight (BW, Fig. 5) With exception of days 1 and 6, BW in group 3 was lower than in groups 1 and 2 in all periods of the trial (all, P > 0.01).No significant differences in BW between groups 1 and 2 were found.

Discussion
Lower values of Hb and PCV in group 1 (Fe 2+ -fumarate) on day 6 did not result in a lower body weight and weight gain compared to group 2 (Fe 3+ -dextran).Although there was a significant decrease of Hb, PCV, MCV and MCH in group 1 from day 21-28, those indices remained comparable with group 2. Neither in group 1, nor in group 2 any piglet become anaemic (i.e.no piglet had Hb less than 80g/l).After weaning at 35 days of age no further decrease of Hb, PCV, MCV and MCH in group 1 was found.We suggest that this was due to the fact that piglets started to feed on weaning pellets (Fe 3+ 124 mg/kg) intensively.Reference ranges in haematology for healthy piglets were investigated by Egeli et al. (1998b).Reference ranges for 35-day-old piglets are as follows: Hb -101 ± 10; PCV -0.32 ± 0.03; RBC -5.79 ± 0.68; MCV-56.5 ± 6.6; MCH -17.6 ± 2.1; MCHC -311 ± 10.In our trial, haematological indices of piglets obtained on day 35 in groups 1 and 2 were found to be within normal physiological range.Since there was a significant decrease of Hb, PCV, MCV and MCH in group 1 on day 28, we consider the double dose of iron as high as 200 mg to be appropriate and we recommend to let piglets free access to feed with Fe to prevent further decline.In study conducted by Kotrbáãek (2001) under experimental conditions, Fe 2+ -fumarate paste was administered to day-old piglets that had been let to suck colostrum several times.Haematological values were measured till 21 days of age.The effect of oral administration of 100 mg fumarate-bound iron on haematological indexes in examined period was equal to that of parenteral administration of 200 mg Fe 3+ and even stronger on some indexes in the 2 nd week of life.In our study, when Fe 2+ -fumarate was administered later, repeated and higher dose of Fe 2+ -fumarate was necessary to achieve comparable haematological values to those obtained by i.m. administration of Fe 3+ -dextran.This difference could be explained by lower intestinal absorption of iron in 6-and 11-day-old piglets.Lower intestinal absorption of iron in 10-day-old piglets is described by Iben (1998).

Days of age DWG (g)
The differences between anaemic (group 3) and healthy piglets (groups 1 and 2) were as expected.Iron deficiency anaemia is generally accepted to be hypochromic (Egeli 1998b).This was also confirmed in our study.The lower Hb, PCV, RBC, MCV, MCH and MCHC showed to have detrimental effect on body weight and weight gain.The anaemic limit, i.e. the point when the anaemia begins to exert a detrimental effect on weight gain or gives rise to clinical symptoms of anaemia is set by most authors at a haemoglobin concentration of 80 g/l (Furugouri 1975;van Kempen 1987).One week after administration of Fe 3+ -dextran, Hb, PCV, RBC, MCV and MCH increased significantly and reached values of groups 1 and 2. MCV and MCH in group 3 on day 28 were found to be significantly higher compared to groups 1 and 2. In study conducted by Holter et al. (1991), injection with iron on day 13 led to a rapid increase in the indices, with statistically significant increase for Hb, PCV, and MCV four days after treatment.A rise in MCV indicates production of new erythrocytes with greater number of large immature cells released into the circulation ( H olter et al. 1991).This seemed also to be the case in this study.Although weight gain increased significantly after administration of Fe 3+ -dextran in the anaemic group, the body weight remained to be significantly lower compared to groups 1 and 2.
We conclude that oral administration of Fe 2+ -fumarate to suckling piglets was efficient in preventing anaemia under conditions of this trial and it resulted in good growth rate of the piglets, which was comparable to piglets that were given Fe 3+ -dextran.