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A pilot study to assess Vitamin D and serum ferritin levels among prospective female blood donors with low hemoglobin levels

 Department of Transfusion Medicine, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Thiruvananthapuram, Kerala, India

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Date of Submission19-Jan-2022
Date of Decision20-Sep-2022
Date of Acceptance02-Oct-2022
Date of Web Publication12-Dec-2022


INTRODUCTION: Anemia is one of the leading causes of deferral from blood donation. Classically, an individual's iron status is assessed with respect to serum ferritin (S. ferritin) levels. S. ferritin levels vary with phases of the menstrual cycle and may not reflect the actual iron stores. Currently, there are certain studies that explain the role of Vitamin D in erythropoiesis. The present study was conducted to assess the emerging association of Vitamin D with hemoglobin (Hb) levels among the female blood donor population.
MATERIALS AND METHODS: A hospital-based cross-sectional study was conducted on 111 female blood donors over 1 year. The association of Hb levels with Vitamin D and S. ferritin levels, dietary factors, frequency of blood donations, and the number of pregnancies was analyzed.
RESULTS: The prevalence of Vitamin D deficiency was 80.18% (n = 89) in this study. Among donors with low Hb levels, 24.32% (n = 27) had low ferritin and 51.34% (n = 57) had low Vitamin D levels. 18.01% (n = 20) of the donors had low Hb levels with coexistent low ferritin and low Vitamin D levels.
CONCLUSION: It is imperative to replenish the iron stores of blood donors who face deferral from blood donation due to low Hb levels. This is a preliminary study from India which looks into the Hb levels of female donors with regard to their Vitamin D status. Those females who were found to be deficient in Hb and Vitamin D levels were referred to treating clinicians for supplementation and dietary modifications. The major goal of this study was to bring those women who got deferred due to low Hb levels back to the voluntary blood donor pool once they replenished their iron stores.

Keywords: Blood donor, hemoglobin, postmenopausal women, premenopausal women, serum ferritin, Vitamin D

How to cite this URL:
Mani A, Nair AR, Gupta D. A pilot study to assess Vitamin D and serum ferritin levels among prospective female blood donors with low hemoglobin levels. Asian J Transfus Sci [Epub ahead of print] [cited 2023 Jan 28]. Available from:

   Introduction Top

Anemia is a common nutritional disorder, i.e., one of the major public health concerns faced by developing countries.[1] The frequency of iron deficiency anemia is high among blood donors (1.8%–8.4% in males and 4.5%–34.8% in females).[2],[3] The hemoglobin (Hb) level should be ≥12.5 mg/dl for blood donation as per the blood donor selection criteria laid down by the Government of India.[4] Iron deficiency is the primary underlying cause for low Hb levels among 70% of the low Hb-deferred donors.[5]

In our institute, blood donor deferrals due to low Hb levels constitute approximately 17% a year. Low Hb is the most common cause of blood donor deferral among female blood donors.[6] Pregnancy and menstruation with poor nutrition contribute to the high incidence of anemia among females.[5] Following deferral, 15% of the donors never return for another blood donation.[7] This causes a loss of critical pool of motivated individuals from voluntary blood donor pool.

Classically, a diagnosis of iron-deficiency anemia is made by the estimation of serum ferritin (S. ferritin) levels.[8] In recent years, evidence indicating potential roles for Vitamin D in iron homeostasis and erythropoiesis is emerging in several studies in healthy and diseased populations and in in vitro studies as well.[9],[10] Vitamin D influences iron metabolism and erythropoiesis by downregulating pro-inflammatory cytokines and hepcidin, thereby increasing iron availability.[11] Vitamin D directly stimulates erythroid precursors for enhanced erythropoiesis.[12] There is also evidence to suggest that Vitamin D may support erythropoiesis in inflammatory causes of anemia.[13],[14],[15]

Vitamin D deficiency is more likely among females. Possible explanations for this observation include increased subcutaneous adiposity and higher body mass index, resulting in increased 25(OH)-Vitamin D storage in adipose tissue, thus not readily available for conversion to calcitriol.[16]

Rationale of the study

In most blood centers, point-of-care test using finger-prick sample alone is used for Hb estimation. Since donors deferred for low Hb represents a significant proportion of the total donor pool, understanding the underlying cause is important. Iron deficiency is usually assumed as the cause for low Hb, and donors are counseled about improving their dietary intake of iron-rich foods. Many studies have found simultaneous occurrence of anemia and Vitamin D deficiency.[9],[11],[17] Vitamin D has got a major role in erythropoiesis and iron homeostasis. The present study was conducted to estimate S. ferritin and Vitamin D levels in prospective female blood donors with and without Hb ≥12.5 mg/dl. This will help in correctly identifying the underlying deficiencies and to advice regarding supplementation.

Objectives of the study

The study was conducted to estimate the S. ferritin and Vitamin D levels in prospective female blood donors with and without Hb ≥12.5 mg/dl. This would help find out the prevalence of S. ferritin and Vitamin D deficiency. Another objective was to correlate the levels of Hb with S. ferritin and Vitamin D levels.

   Materials and Methods Top

A hospital-based cross-sectional study was started after obtaining clearance from the Institutional Ethics Committee (SCT/IEC/1611/DECEMBER-2020). A total of 111 female blood donors in the age group of 18–65 years who registered for blood donation for 1 year were enrolled in the study. The blood donor selection criteria was in accordance with the guidelines for blood donor selection and deferral criteria laid down by the Government of India.[4]

Exclusion criteria

Donors on iron and/or Vitamin D supplements in the past 3 months, history of acute infections in the past 1 month or chronic cardiac, hepatic, or renal disorders, collagen vascular disorders, any known bleeding disorder or blood cell dyscrasias, history of recent blood loss including menstrual cycles, history of blood transfusions in the past 3 months. Participants with the above-mentioned conditions were excluded from the study due to the possibility of false values of S. ferritin as it is an acute-phase reactant.

Data collection

On obtaining informed written consent, in addition to the blood donor screening questionnaire, information regarding premenopausal or postmenopausal age group, parity, last menstrual period, frequency and number of previous blood donations, previous deferral due to low Hb, dietary intake of iron and Vitamin D-rich foods, hours of sun exposure, and any history of chronic diseases was elicited from the study participants by the investigator.

A volume of 2 ml of blood sample was collected from the study participants who were deferred from blood donation due to low Hb levels. In all accepted blood donors, pilot samples collected during blood donation was used for testing purpose. S. ferritin and Vitamin D estimation was performed using enzyme-linked fluorescent assay technique on Biomerieux-VIDAS semiautomated analyzer system. Collected samples were centrifuged at 4000 rpm for 10 min and serum was dispensed to Biomerieux serum S. ferritin (Lot No. 1008313880) and Vitamin D-25-Hydroxy calcitriol (Lot No. 1008163180) estimation kits. As per the manufacturer's instructions, the normal S. ferritin levels were 10–160 ng/ml for premenopausal women and 25–280 ng/ml for postmenopausal women. Normal Vitamin D levels were >30 ng/ml and those individuals with levels <30 ng/ml were considered Vitamin D deficient. A Hb level of ≤12.5 mg/dl was considered low Hb levels in our study as per the blood donor selection criteria laid down by the Government of India.[4] After the estimation of Vitamin D and S. ferritin levels, the donors were informed of their reports over phone, and necessary recommendations for dietary modifications were advised. Those donors with severe deficiencies of Vitamin D and serum S. ferritin were advised to visit the blood center and were referred to the peripheral health centers for further management.

Statistical analysis

Numerical data were expressed as mean ± standard deviation, and categorical data were expressed in frequency (n) and percentage (%). Association between categorical variables was analyzed using Pearson's Chi-square test. Pearson's correlation coefficient (r value) was used to measure the relationship between two continuous variables. Continuous variables were compared between two groups using Student's t-test and with more than two groups with the ANOVA test. All statistical tests were two sided. P < 0.05 was considered statistically significant, and data analysis was performed using SPSS (version 23.0) : SPSS inc. Chicago, USA.

   Results Top

The study was conducted on 111 female blood donors for 1 year. The median age was 40 years (interquartile range [IQR] = 22–59), and the median weight was 62 kg (IQR = 45–93). 77.5% (n = 86) of women were premenopausal and 22.5% (n = 25) were postmenopausal. Multiparous women constituted about 58.6% (n = 65), and 1.8% (n = 2) were nulliparous married women. We had a total of 72.1% (n = 80) of first-time donors and 27.9% (n = 31) were repeat donors. Of the total 111 female blood donors, 80.2% (n = 89) had a history of deferral from blood donation due to low Hb levels [Table 1].
Table 1: Type of blood donors and deferral history due to low hemoglobin levels (≤12.5 g/dl)

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Since the normal S. ferritin levels were different for premenopausal women (10–160 ng/ml) and postmenopausal women (25–280 ng/ml), participants were categorized into premenopausal and postmenopausal age groups for further assessment.

The median age of premenopausal women was 37 years (IQR = 22–52) and that of postmenopausal women was 54 years (IQR = 39–59). The median Hb was 11.9 g/dl (IQR = 8.60–14.20) for premenopausal and 12.3 g/dl (IQR = 11.10–13.70) for postmenopausal women. The median S. ferritin level was 17.33 ng/ml (IQR = 1.0–91.48) and median Vitamin D level was 15.15 ng/ml (IQR = 4–68.6) for premenopausal women. The median S. ferritin level was 55.5 ng/ml (IQR = 15.80–122.12) and median Vitamin D level was 19.20 ng/ml (IQR = 4–64) for postmenopausal women.

In the present study, the prevalence of low Hb and low S. ferritin was 64.85% (n = 72) and 24.32% (n = 27), respectively. The prevalence of Vitamin D deficiency was 80.18% (n = 89) [Table 2]. Among donors with low Hb, 24.32% (n = 27) had low S. ferritin levels and 51.34% (n = 57) had low Vitamin D levels. 18.01% (n = 20) of the donors had low Hb levels with coexistent low S. ferritin and low Vitamin D levels [Table 3]. The donors with low Hb, S. ferritin, and Vitamin D levels are represented as an area of clustering in [Figure 1].
Table 2: The prevalence of low hemoglobin, ferritin, and Vitamin D levels among the study participants

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Table 3: Serum ferritin and Vitamin D levels of the study participants with hemoglobin levels ≤12.5 g/dl and ≥12.5 g/dl*

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Figure 1: Scatter plot illustrating hemoglobin levels on X-axis, Vitamin D and S. ferritin levels on Y-axis. Clustering of Vitamin D values <30 ng/ml and hemoglobin <12.5 g/dl observed. Serum ferritin = S. ferritin

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The prevalence of chronic diseases among the study participants: 11.7% of the study participants were on medication for hypothyroidism and 0.9% on antihypertensive medications. Both groups had optimal control of their thyroid hormone levels and blood pressure. 94.6% (n = 105) of the females were nonvegetarians and 5.4% (n = 6) were pure vegetarians [Table 4]. Of 111 study participants, 83.7% (n = 93) were on iron-rich foods such as green leafy vegetables, citrus fruits, dates, and meat products.
Table 4: Percentage of donors with chronic disorders under control on medications and dietary types

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Since our blood center accepts only voluntary blood donations, the major mode of blood collection was through outreach blood donation camps either in educational institutions, corporate establishments or public and private sector offices. Hence, the female blood donors enrolled in our study were all working women who would stay indoors during daytime hours. Only five women commented sun exposure of 3 h/day, and this was the maximum hours of sun exposure we could observe in this study. A majority of women accounted for <2 h of sun exposure per day. We could not elicit an association between hours of sun exposure and Vitamin D levels (P = 0.172). Multiparous women constituted about 58.6% (n = 65), and 1.8% (n = 2) were nulliparous married women. We could not find any association between the number of pregnancies and low Hb (P = 0.215) and Vitamin D levels (P = 0.469).

In this study, we had 33 repeat female blood donors. There were only two women who donated more than two times among repeat donors, and the duration between the donation in these women were more than 1 year. Therefore, we could not find any association between the frequency of blood donations and low Hb or Vitamin D levels as well.

Our study could significantly correlate Hb with S. ferritin levels among the total study participants (r = 0.315) and premenopausal females (r = 0.316) [Table 5]. No statistical significance was observed between Hb levels and Vitamin D levels and between the number of pregnancies with Hb levels (P = 0.215) or Vitamin D levels (P = 0.469).
Table 5: Statistical correlation of hemoglobin with serum ferritin and Vitamin D levels (Pearson's correlation-two tailed, paired t-test r values and p values)

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   Discussion Top

Iron deficiency is common among women of childbearing age group, for whom pregnancy and menstruation are the dominant causes.[6] In this study, the prevalence of low Hb (<12.5 g/dl) was 69% (n = 59) among premenopausal women. In blood centers, finger-prick Hb level alone is used as a screening test for the eligibility to donate blood even though iron stores may be depleted in donors with Hb values above the arbitrarily cutoff for blood donation.[18]

S. ferritin is probably the most useful laboratory test to measure iron levels and is directly a fraction of iron stores.[19] The Canadian blood services used S. ferritin levels, in addition to the standard point-of-care Hb testing, to determine the extent of iron-deficiency anemia in blood donors and to update recommendations in accordance.[20] However, S. ferritin is an acute-phase reactant that is elevated in acute inflammatory conditions as well.

Vitamin D has an important role in iron metabolism and erythropoiesis, as evidenced by animal and human studies.[10] Many recent studies have found association between Vitamin D and anemia and suggested that maintenance of sufficient Vitamin D status may be important in preventing anemia.[9] The present study found suboptimal Vitamin D and serum S. ferritin levels in 23.43% of the study participants.

The prevalence of Vitamin D deficiency was more than 80% in the current study, which is comparable to the community-based studies on apparently healthy population with a prevalence ranging from 50% to 94%.[21],[22],[23] Less than 10% of the study participants alone had a normal Vitamin D and S. ferritin levels. We could observe that Vitamin D deficiency was more common than iron deficiency.

Our study could observe 33% (n = 37) of females with low Hb and low Vitamin D levels with normal S. ferritin values. S. ferritin levels are known to fluctuate during the menstrual cycle, with the lowest levels during menses and the highest in the luteal or late luteal phase. These cyclic variations are a potential source of error when S. ferritin assays are performed in premenopausal females.[24] Elevated S. ferritin levels are also seen in chronic inflammatory causes of anemia. Hence, even in the presence of elevated S. ferritin levels also, iron-deficiency anemia can be present. A study by Munasinghe et al. showed that in patients with low Hb and elevated S. ferritin levels, Vitamin D was found to be deficient.[25]

Recent studies have demonstrated that the frequency of iron deficiency is more dependent on the frequency of blood donations than on the accumulated number of blood donations.[18] There were only two repeat blood donors, and the maximum number of blood donations in the present study was ten times. However, both these women had normal Hb levels even though they were Vitamin D deficient. The interdonation interval of these repeat donors was more than 6 months.

The most common causes of Vitamin D deficiency are due to vegetarian diet, inadequate exposure to sunlight, use of sunscreens, cultural practices like wearing burqa/purdah, which can hamper the synthesis of Vitamin D in the skin by ultraviolet rays, and unspaced and/or unplanned pregnancies in women with preexisting dietary deficit of Vitamin D.[22]

In a meta-analysis of 14 randomized control trials, Vitamin D supplementation had positive effects on transferrin receptor saturation and iron status.[17] Even though our study could not demonstrate a clear-cut correlation between Hb levels and Vitamin D levels, Vitamin D could become a new modality to tackle low Hb levels, and this issue can be addressed with a major concern in future studies. It is important to secure an adequate iron reserves in the donor population to maintain an appropriate donation potential and to avoid side effects of iron-deficiency anemia.[26] Thereby, we can increase the number of eligible female blood donors in the voluntary blood donor pool.

Even though we have not included apheresis donors in our study, normal Vitamin D levels might play a vital role among them, as hypocalcemia is a very common adverse reaction during apheresis procedures.[27] Vitamin D is important in calcium absorption, and donors with deficient Vitamin D levels can have defective calcium absorption complicating apheresis donations.

   Conclusion Top

This study appears to be the first to look into Vitamin D levels among the female donor population from India. Vitamin D deficiency among women is most often underdiagnosed and undertreated compared to iron deficiency. This pilot study was carried out in light of few studies that state the role of Vitamin D for erythropoiesis. The current study could observe a very high prevalence of Vitamin D deficiency among female blood donors accounting for more than 80%. We need to create awareness among the general public and in motivated blood donors, in particular, about the importance of Vitamin D deficiency and the expected consequences. We could tackle iron and Vitamin D deficiency by advice on dietary modifications and supplementation for those with severe deficiency. The participants in our study include only women, and henceforth, more studies should come forward involving the general population to establish an association between Vitamin D deficiency and low Hb levels if they are related.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

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Correspondence Address:
Amita Radhakrishnan Nair,
Department of Transfusion Medicine, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Medical College P. O., Thiruvananthapuram - 695 011, Kerala
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/ajts.ajts_13_22


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  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5]


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