Abstract
Refractory hypothyroidism is associated with high morbidity and increased healthcare expenditure. In general, the use of the levothyroxine absorption test looks promising in evaluating refractory hypothyroidism but has shown significant variability in protocols in multiple settings. We intended to assess the usefulness of the levothyroxine absorption test in a low-resource setting and to assess the factors associated with refractory hypothyroidism. A cross-sectional study among age-matched 25 cases of refractory hypothyroidism and 24 treatment-responsive hypothyroid controls was conducted. A supervised levothyroxine absorption test was performed with levothyroxine 1000 μg tablets after a 10-h fast, and serum free tetraiodothyronine (FT4) levels were measured at 0, 1, 2, 3, 4, and 5 h. Descriptive statistics, chi-square test, Student’s t-test, and logistic regression were used in the analysis. Results showed no significant difference in age, body weight, etiology of hypothyroidism, interfering medications, thyroxine storage, and ingestion technique in cases and controls. Cases had a longer duration of hypothyroidism and males had a higher peak FT4 concentration. During pooled analysis, serum FT4 peaked at 3 h with an increment of 149.4% (128.4–170.5%) from baseline and plateaued thereafter. The absolute value of FT4 at 3 h was 41.59 (s.d. 14.14) pmol/L (3.23 ng/dL). We concluded that there was no significant difference in the pattern of levothyroxine absorption in both groups. The most common cause of refractory disease was pseudo-malabsorption. Rapid supervised levothyroxine absorption test with two blood samples for FT4 at baseline and at the peak of absorption (3 h) is simple, convenient, and cost-effective, particularly in low-resource settings.
Introduction
Refractory hypothyroidism is characterized by an inadequate response to the standard dose of levothyroxine, resulting in persistent biochemical or clinical hypothyroidism (1). The recommended average daily dose of levothyroxine for the treatment of hypothyroidism is 1.6 μg/kg (2).
Once ingested on an empty stomach, levothyroxine is disintegrated and dissolves in the stomach, and nearly 70–80% of the ingested levothyroxine is absorbed in the small intestine in a healthy person (3). Upon absorption, thyroxine is transported in the circulation bound to transport proteins, and subsequently, tetraiodothyronine (T4) undergoes deiodination in peripheral tissues to produce the active form of thyroxine, triiodothyronine (T3). Free triiodothyronine (FT3) binds to the nuclear receptors in the cells to exert its biological action.
Refractory hypothyroidism is defined as the requirement of higher doses of levothyroxine, typically 1.9–2.5 μg/kg/day, to maintain biochemical and/or clinical euthyroidism (1, 4). The exact prevalence of refractory hypothyroidism is unknown. However, a large retrospective cohort study done in the United Kingdom revealed that about 20–30% had TSH above 4 mIU/L after 5 years of levothyroxine treatment (5). This suggests the possibility of a considerable number of patients with undiagnosed refractory hypothyroidism. Patients with refractory hypothyroidism often require frequent clinic visits, close monitoring, and multiple laboratory investigations, which increases the financial burden on the healthcare system and individual patients (6). Poor control of hypothyroidism is associated with significantly increased mortality and morbidity (7). Therefore, optimizing the treatment of refractory hypothyroidism is invariably beneficial for individuals as well as for healthcare systems.
Refractory hypothyroidism can be multifactorial. The commonest cause of refractory hypothyroidism is poor levothyroxine compliance, which is also known as ‘pseudo-malabsorption’. Decreased levothyroxine bioavailability due to interference with drugs and food, and increased demand for levothyroxine secondary to weight gain and pregnancy can also cause refractory hypothyroidism. Malabsorption remains a rare but important cause of refractory hypothyroidism. Underlying autoimmune disorders, such as impaired metabolism, genetic factors, and nonthyroidal conditions that affect hormonal regulation, are a few uncommon causes (4).
A supervised levothyroxine absorption test is being used by many clinicians and researchers for the evaluation of refractory hypothyroidism before conducting extensive and invasive investigations for malabsorption. It is carried out by measuring the serum thyroxine levels at various intervals after ingestion of a specified dose of levothyroxine to assess the levothyroxine absorption. Protocols used for the evaluation of levothyroxine absorption vary greatly according to the duration of the test, levothyroxine formulation and dose being used, frequency and number of venesections required, metrics and analytes used for comparison, etc. (8).
There are no studies available in Sri Lanka to date that assess the characteristics and associations of refractory hypothyroidism. We carried out a cross-sectional study in patients with refractory hypothyroidism in a tertiary care thyroid clinic to evaluate the factors associated with refractory hypothyroidism and to assess the applicability of the thyroxine absorption test in a low-resource setting like Sri Lanka using a simple, convenient, and a cost-effective test protocol. Following the analysis of the results, we propose the use of two blood samples taken at baseline and at the peak of absorption (3 h) for FT4 assessment to assess the absolute FT4 level and calculation of the percentage rise of FT4 to interpret the levothyroxine absorption in a low-resource setting like Sri Lanka.
Materials and methods
Study design, setting, and participants
An age-matched, single-center, case-control study was conducted using attendees of a thyroid clinic at a tertiary care hospital in Sri Lanka. Since the exact prevalence of refractory hypothyroidism in Sri Lanka is not known, all the patients who had refractory hypothyroidism among those who presented for clinic follow-up over 1 year were selected to take part in the study.
Refractory hypothyroidism was defined as taking levothyroxine at more than 2.5 µg/kg/day for 6 months before presentation with normal or elevated thyroid-stimulating hormone (TSH) (TSH above 2 mIU/L). Age-matched controls were selected by systematic sampling (every other patient who met the eligibility criteria for the control group was selected during their routine clinic visit). Eligibility for the control group was defined as having normal TSH for at least 6 months before recruitment and taking levothyroxine equal to or less than 1.6 µg/kg/day. Patients who were either pregnant or planning pregnancy, had secondary hypothyroidism, had a history of thyroid malignancies, were on treatment for psychiatric illnesses, or had been either diagnosed with or had symptoms of cardiovascular disease or angina were excluded from the study. Patients were asked about compliance with levothyroxine, and those who volunteered poor compliance were not recruited for the study. The levothyroxine ingestion technique was assessed by asking patients whether they took levothyroxine on an empty stomach and waited at least 30–60 min before consuming food or drinks.
Twenty-five cases and 24 controls were enrolled for the study with informed written consent from the participants after a detailed verbal and written explanation of the procedure. The study was performed in accordance with the ethical principles of the Declaration of Helsinki, and ethical clearance for the study was obtained from the Ethical Review Committee, Teaching Hospital, Kandy.
Data collection
Basic socio-demographic and clinical data were collected and entered into a data sheet through an interviewer-administered questionnaire and by reviewing patients’ medical records. Anthropometric measurements were taken using well-calibrated equipment (the same stadiometer and floor scale) by the investigators to improve the reliability and accuracy of the data while minimizing measurement errors. Following the collection of demographic, clinical, and anthropometric data, the participants were given both verbal and written instructions and discussed the procedure and protocol of preparation for the levothyroxine absorption test. A date for a supervised levothyroxine absorption test was arranged within the next 6–8 weeks for all the patients recruited into the study.
Protocol of supervised rapid levothyroxine absorption test
A supervised levothyroxine absorption test was done following an overnight fast of 10 h. All the participants were advised not to take their usual dose of levothyroxine in the morning on the day of the test. Further, participants were advised to avoid any interfering medications that are known to interact with levothyroxine absorption on the day prior to and on the day of the absorption test.
Levothyroxine absorption test was carried out in the outpatient investigation unit of the Endocrine clinic, National Hospital, Kandy, Sri Lanka. An intravenous cannula (22–24 G) was inserted, and the first blood sample was drawn for baseline TSH and FT4 level assessment at ‘0’ h. Then 1000 µg of levothyroxine of the same brand (Thyronorm by Abbott Pharmaceuticals) was administered to all participants orally with 300 mL of water under the direct supervision of the study investigators. The participants were continuously monitored 2-hourly for blood pressure, heart rate, development of thyrotoxic symptoms, or any new symptoms starting from just before the administration of the levothyroxine dose until 1 h after drawing the last blood sample.
The participants were advised to remain fasting until the end of the test (till blood samples for thyroid functions were taken at 5 h after levothyroxine ingestion) and were allowed to drink water after 1 h of ingestion of levothyroxine.
Blood samples were collected in 3 mL syringes with no heparin from all the participants at 60, 120, 180, 240, and 300 min from ingestion of levothyroxine to check serum FT4 levels. The IV cannula was flushed with 2 mL of normal saline following each blood withdrawal to maintain patency. At the end of the test (300 min after levothyroxine ingestion), participants were allowed to take their meals. After monitoring for another 60 min after the last blood test, they were discharged from the hospital. All participants were advised not to take their routine levothyroxine dose for 1 week before restarting their regular dose. The participants were followed up at their routine clinics, and the results of the tests were discussed during their clinic review.
Hormone assays
Blood samples were immediately centrifuged and serum was separated for the analyses of TSH and FT4. Serum TSH and FT4 were estimated by chemiluminescence immunoassay technique using commercially available kits with a Vitros 3600 analyzer. Analytical sensitivity for the Vitros 3rd generation TSH assay was 0.015 mIU/L and for FT4 was 0.08 pmol/L (0.006 ng/dL). The laboratory reference range for TSH was 0.465–4.68 mIU/L, and for FT4 was 10–28.2 pmol/L (0.77–2.19 ng/dL).
Data analysis
R programming language (version 4.1.1) was used for data analysis. Continuous variables were statistically presented with measures of central tendency (means) and variation (s.d.) and categorical variables were presented with percentages. Student’s t-test and Pearson’s chi-square were used for continuous and categorical variables, respectively. The comparison of the serial FT4 values during the thyroid absorption test at specific intervals was conducted using logistic regression with a linear mixed-effects model incorporating random intercepts for each participant. Levothyroxine absorption test values were analyzed in individualized case and control groups and subsequently in a pooled manner among groups, and the predictions of expected rise in the free T4 levels were modulated using a linear mixed-effects model for pooled data. P < 0.01 was considered indicative of a statistically significant difference, while 0.01 ≤ P < 0.10 was interpreted as a possible difference.
Results
Baseline characteristics
The baseline characteristics of the participants are summarized in Table 1. There were no statistically significant differences in the mean age, gender, body weight, body mass index (BMI), etiology of hypothyroidism, interfering medications, levothyroxine storage technique, and levothyroxine ingestion technique between cases and controls. Cases had a longer mean duration of being detected with hypothyroidism than the controls while the most recent serum TSH levels were also significantly higher in the cases as expected. The mean dose of levothyroxine among the cases was 219 µg/day, which was significantly higher with a P-value of 0.002 (95% CI), while it was 100 µg/day among the patients in the control group. The mean daily dose of levothyroxine was 3.58 µg/kg in cases and 1.58 µg/kg in controls.
Baseline characteristics of study participants.
| Cases (n = 25) | Controls (n = 24) | P | ||
|---|---|---|---|---|
| Mean age in years (s.d.) | 38.97 (13.08) | 43.83 (13.12) | 0.2 | |
| Gender (%) | Females | 18 (72%) | 22 (91.7%) | 0.159 |
| Males | 7 (18%) | 2 (8.3%) | ||
| Mean duration of hypothyroidism in years (s.d.) | 9.64 (8.03) | 5.83 (3.87) | 0.044 | |
| Aetiology of hypothyroidism | Autoimmune thyroiditis | 20 | 21 | 0.659 |
| Post-surgical | 2 | 2 | ||
| Radioiodine treatment | 1 | 1 | ||
| Congenital hypothyroidism | 2 | 0 | ||
| Mean levothyroxine dose in micrograms (s.d.) | 219.10 (71.0) | 100.00 (29.95) | 0.002 | |
| Mean body weight in kg (s.d.) | 61.16 (16.21) | 63.2 (10.64) | 0.605 | |
| Mean BMI, kg m-2 (s.d.) | 25.52 (5.69) | 26.89 (3.97) | 0.336 | |
| Mean Last TSH, mIU/L (s.d.) | 12.80 (8.50) | 2.37 (0.95) | 0.008 | |
Patients in both groups were on antihypertensives, diuretics, aspirin, statins, vitamin D, 1-alpha cholecalciferol, vitamin C, ferrous sulfate, multivitamins, folic acid, proton pump inhibitors, oral hypoglycemics, inhalers, antacids, calcium, coffee, prednisolone, and mesalamine. However, there was no statistically significant difference in the medications that the patients were taking between the groups.
Supervised levothyroxine absorption test among the cases and controls
The mean serum TSH level at baseline (0 min) was 17.46 (7.36) mIU/L in the cases, while it was 2.49 (0.58) mIU/L in the control group (P-value < 0.01) at the start of the test. Measured serum values of FT4 levels following the Levothyroxine absorption test at 0, 60, 120, 180, 240, and 300 min following administration of levothyroxine are depicted below in scatter plot diagrams.
Except for one participant in the cases (patient no. 15, who had a flat levothyroxine absorption curve), the rest of the participants in both groups had a gradual rise in FT4 levels during the test (Fig. 1). Most patients reached a plateau in their FT4 levels after 180 min. There was no statistically significant difference between the mean FT4 values between cases and controls at all time intervals (0, 60, 120, 180, 240, and 300 min) as shown in plot (B) of Fig. 2 and Table 2. No adverse events or symptoms of hyperthyroidism were reported, and heart rate and blood pressure recordings did not show significant changes during the monitoring period (from ingestion until 6 h after ingestion of levothyroxine). In patient no. 15, who exhibited a flat levothyroxine absorption curve, celiac screening was negative, upper and lower gastrointestinal endoscopy results were normal, and abdominal imaging was unremarkable, so no cause was identified for her malabsorption.
A line graph with the FT4 distribution pattern in all the patients (cases and controls pooled together).
Citation: Endocrine Connections 13, 10; 10.1530/EC-24-0277
Scatter plots showing the distribution of FT4 values at 0, 60, 120, 180, 240, and 300 min in individual patients during a supervised thyroxine absorption test in cases and controls (A),and when cases and controls are plotted in the same graph (B). Solid lines indicate the smoothed conditional means from local polynomial regression fitting for FT4 values for cases and controls. Shaded area indicates the confidence interval around the smooth for cases and controls.
Citation: Endocrine Connections 13, 10; 10.1530/EC-24-0277
Comparison of serial serum FT4 levels during levothyroxine absorption test.
| 0 min | 60 min (1 h) | 120 min (2 h) | 180 min (3 h) | 240 min (4 h) | 300 min (5 h) | |
|---|---|---|---|---|---|---|
| FT4 level in cases (s.d.) pmol/L | 17.02 (8.17) | 24.28 (11.47) | 31.95 (14.32) | 40.88 (16.64) | 42.07 (16.29) | 41.24 (15.15) |
| FT4 level in controls (s.d.) pmol/L | 17.67 (4.77) | 27.32 (7.60) | 37.21 (10.60) | 42.32 (11.27) | 42.79 (10.43) | 41.48 (10.47) |
| P | <0.735 | <0.282 | <0.152 | <0.725 | <0.855 | <0.953 |
Pooled analysis of the cases and controls
We pooled the cases and controls together for further analysis of the thyroxine absorption test, as there was no statistically significant difference between the cases and the controls. The pattern of absorption of individual participants is depicted in Fig. 1.
During the subgroup analysis, there was no statistically significant difference in the pattern of levothyroxine absorption according to the category of BMI (BMI category of 18.6–24.9 vs 25–30), and age (20–29 years vs 30–50 years). Males had a higher rate of levothyroxine absorption compared to females, and the mean peak FT4 concentration was higher for males than females (Fig. 3).
Scatter plots indicating the distribution of FT4 values at 0, 60, 120, 180, 240, and 300 min in individual patients during a supervised thyroxine absorption test during subgroup analysis. (A) According to BMI category (Blue: BMI 18.6–24.9 kg/m-2, Red: BMI 25–30 kg/m-2); (B) According to the age category (Red: 20–29 years, Blue: 30–50 years); (C) According to the gender (Red – Females, Blue – Males). Solid lines indicate the smoothed conditional means from local polynomial regression fitting for FT4 values for cases and controls. Shaded area indicates the CI around the smooth for cases and controls. Age_cat, Age category; BMI_cat, BMI category.
Citation: Endocrine Connections 13, 10; 10.1530/EC-24-0277
Development of a formula to predict the expected FT4 value during the supervised levothyroxine absorption test
After pooling the data of all the participants in the study (excluding patient no. 15, who had poor absorption), we did a logistic regression analysis with a linear mixed-effects model incorporating random intercepts for each participant to predict the expected FT4 value during the levothyroxine absorption test. It can be used to calculate the expected FT4 level at a given time during the absorption test (60, 120, 180, 240, and 300 min). Parameters required for calculation were gender, baseline TSH level, and the time of FT4 after levothyroxine ingestion (in minutes).
Formula
Expected FT4 (pmol/L) = 12.70 (a constant) + 9.50 if gender is male (0 if female) + 0.13 × time of the blood test in minutes – 0.13 × (time – 180) if time is more than 180 minutes − 0.21 × baseline TSH level before the test in mIU/L.
Percentage increase of FT4
There was nearly a 1.5 times increase in the mean FT4 level from the baseline at 180 min (3 h) and the line plateaued afterward (Fig. 4). Table 3 shows the mean percentage FT4 rise with 95% CI and absolute FT4 level during the test at a given hour.
A line graph showing the percentage rise of mean FT4 value compared to the baseline at 1, 2, 3, 4, and 5 h in all the patients during the supervised thyroxine absorption test. Blue lines indicate 95% CI.
Citation: Endocrine Connections 13, 10; 10.1530/EC-24-0277
Absolute FT4 level and percentage rise of FT4 level compared to the baseline over 1, 2, 3, 4, and 5 h during the thyroxine absorption test.
| Time of the FT4 assessment | Percentage rise of FT4 level from baseline (95% CI) | Absolute FT4 level (s.d.) pmol/L |
|---|---|---|
| 0 h | – | 17.34 (6.66) |
| 1 h | 52.1 (40.2–64.0) | 25.77 (9.79) |
| 2 h | 104.3 (88.0–121) | 34.53 (12.78) |
| 3 h | 149.4 (128.4–170.5) | 41.59 (14.14) |
| 4 h | 157.2 (136.7–177.7) | 42.42 (13.60) |
| 5 h | 156.9 (133.8–180.0) | 41.1 (12.91) |
Discussion
A multitude of variations is observed among the protocols in the measurement of serum levothyroxine levels worldwide. Therefore, when selecting the appropriate test protocol, we selected a short-supervised levothyroxine absorption protocol since it is more convenient for patients as well as for clinicians. In our protocol, the test was carried out in an outpatient setting and the maximum time of in-hospital stay was less than 7 h. A long-supervised levothyroxine absorption test, which was the preferred protocol in the initial studies, involved prescribing a weight-adjusted weekly dose of levothyroxine given to patients under direct supervision every week for 4–6 weeks and monitoring TSH at the end of the test and FT4 at weekly intervals (9). It is time-consuming, requires a significant number of resources and commitment, and potentially has a higher patient dropout rate. Alhasan et al. compared the rapid test with the long levothyroxine absorption test in a cross-sectional analysis and showed that the rapid 2-h absorption test has 88.8% sensitivity and 91.6% negative predictive value for diagnosing levothyroxine malabsorption, justifying the use of the rapid absorption test in clinical practice (10).
In the current study, we carried out hourly FT4 assessments for 5 h. In the described protocols in the literature, the frequency of blood draws varied between every 15 min (11), 30 min (12), and hourly intervals (13). The serum FT4 concentration is known to peak within 3–4 h of ingestion of levothyroxine in pharmacokinetic studies (3). We believe our protocol of hourly sampling for 5 h was more comfortable for the patients, and it allowed adequate time to observe the pattern of absorption in our population.
We have used 1000 µg of levothyroxine for all the patients, and none of the 49 patients reported any symptoms of hyperthyroidism or adverse events during the study. Even though we used a fixed dose of levothyroxine to minimize errors and due to less variability in anthropometric measurements in our study population, several studies suggest the alteration of dosage. A higher dose of levothyroxine (1500 µg) was suggested if BMI was above 40 kg/m-2 (14), and a lower dose (600 µg) was suggested if the age was above 65 years and if the patient had either low or normal BMI (13). We believe using a standard dose of 1000 µg is more convenient and minimizes errors, and a recent review by Caron et al. justified the use of 1000 µg (8).
In this study, except for a single patient, all the participants in the case group were observed to have a gradual rise in serum FT4 levels during the absorption test, indicating that the possible etiology for refractory disease is not malabsorption but ‘pseudo-malabsorption’. Although true absorption problems are commonly sought for the etiology of refractory hypothyroidism, most of the time refractory hypothyroidism is due to pseudo-malabsorption. The rate of pseudo-malabsorption in patients with refractory hypothyroidism may be as high as 84.5%, according to one observational study done in Brazil (15). In a population study done in the USA to evaluate compliance with levothyroxine, 51.9% of people were nonadherent to levothyroxine treatment after 12 months of initiating treatment (16). In our study, we inquired from our participants about the compliance with levothyroxine medication, as anyone would generally do in the routine clinical setup when recruiting for our study. Even though we excluded the patients who admitted to missing levothyroxine tablets in the exclusion criteria, this study indicates that patients with pseudo-malabsorption may still be missed, particularly in busy clinical settings. Asking targeted questions regarding compliance in a non-judgmental manner, such as ‘How many tablets have been missed per week/month?’ and ‘How many tablets remain on the next clinic date?’ can be very helpful in assessing adherence. Establishing a strong therapeutic alliance between a clinician and a patient through continuity of care can enhance the detection of medication noncompliance, potentially avoiding the need for absorption tests.
When comparing the absorption pattern and the baseline characteristics, there were no statistically significant differences in age, weight, BMI, etiology of hypothyroidism, storage and ingestion techniques, or interfering medications among the cases and controls. This study showed an increased rate of absorption of FT4 in males compared to females, which has not been previously reported. The exact reason for this difference is not clear, and due to the small number of males in our study, we suggest further evaluation with a larger sample of males and females. During the literature search regarding available studies reporting gender differences in the characteristics of levothyroxine treatment, one study indicated that males required a lower dose of levothyroxine replacement, which they discussed as related to their lower body weight compared to the females in this particular study (17). We observed that cases had a longer duration of hypothyroidism compared to controls. One possible explanation for this difference is that patients with long-standing disease may become less compliant with levothyroxine therapy over time.
In the pooled analysis, there was a gradual rise in the serum FT4 concentration, which had a mean peak FT4 level at 3 h followed by a plateau. According to pre-existing studies, the rate of levothyroxine absorption may demonstrate individual variation (8) and is reported to be about 2 h in euthyroid individuals (9), and about 4–6 h in hypothyroid individuals (18). Following a review of the studies available so far, Caron et al. suggested the use of a 4-h levothyroxine absorption test for evaluating levothyroxine absorption (8). In the current study, we found to have a mean peak serum FT4 concentration at 3 h coinciding with the results of the majority of reported studies in the literature (8).
In this study, serum FT4 concentration increased by 52.1% (40.2–64.0) at 1 h, 104.3% (88.0–121) at 2 h, 149.4% (128.4–170.5) at 3 h and 157.2% (136.7–177.7) at 4 h. In the literature, the percentage rise of FT4 concentration at the peak of absorption was variable and ranged from more than 60% at 2 h (13) to 160 ± 58% s.d. at 4 h (19). Even though the levothyroxine absorption test is widely used for evaluating refractory hypothyroidism, there is still debate regarding which metrics and cutoff values should be used in deciding adequate levothyroxine absorption. Some authors have calculated the peak TT4 after ingestion of a weight-based dose of levothyroxine (13), a calculation that may be complex in routine clinical settings. Further, it is an oversimplified method of calculating the volume of distribution using the same fraction of BMI, which may not be applicable to all patients. This method is influenced not only by the rate of drug absorption but also the rates of drug distribution and clearance. Some researchers have used the area under the curve (AUC) of FT4 (20). In both metrics, AUC and percentage TT4 rise, one should consider that there is a steady state of elimination of the drug.
Regarding the use of serum FT4 as the metric of choice instead of TT4, Sun et al. evaluated this in a retrospective analysis of levothyroxine absorption tests and showed that serum FT4 and TT4 were correlated (R = 0.8) (12). Using this evidence, and since TT4 measurement is not readily available in our setup, we measured serum FT4 concentration to assess the absolute rise and calculated the percentage rise in FT4 in our cohort. Furthermore, there is a variation in assay cut-offs and units of measurement of FT4 concentration in different laboratories. Hence, we measured the absolute FT4 rise and calculated the percentage rise in FT4, as it can be applicable to many settings.
Following a logistic regression analysis with a linear mixed-effects model incorporating random intercept, we introduced a novel formula to calculate the expected FT4 at a given time during the absorption test for a given patient according to the baseline TSH level, gender, and the point of FT4 testing during the absorption test. When using this formula, only one blood sample taken at the peak of absorption (3 or 4 h) is required and it can be compared with the expected FT4 level according to the formula to decide if the absorption of levothyroxine is satisfactory. Although the clinical utility of this formula can be limited given the small sample size, this study could serve as a pilot study for planning further large-scale studies for validation.
There are a few limitations to our study. If we had included subgroups of healthy individuals as well as individuals with confirmed malabsorption, we could have made broader comparisons and defined a cutoff value for confirmed malabsorption in our population. Regarding the sample size in our study, we had to limit it to 25 cases as the blood testing kits were limited and the study had to be completed within the specified time. Our study’s single-center, cross-sectional design may limit the generalizability of the findings due to racial and ethnic differences in levothyroxine absorption. Furthermore, due to the small sample size, it is difficult to draw a conclusion about the effects of different medications on levothyroxine absorption. Given our clinical setting and available infrastructure, we believe the best possible sampling was achieved. In comparison to similar studies conducted worldwide, the pooled sample of 48 patients is considered a high level of sample recruitment given the prevalence and the availability of the patients in a given setting.
Conclusion
There was no statistically significant difference in the supervised levothyroxine absorption test between the cases and controls in our study, indicating that pseudo-malabsorption was the cause of refractory hypothyroidism. During pooled analysis, we found that the mean peak serum FT4 occurred at 3 h and FT4 increased by 149.4% (128.4%–170.5%) and the mean absolute FT4 increased to 41.59 (s.d. 14.14) pmol/L at 3 h. These values can be used as the cutoff level for normal thyroxine absorption in our population. When performing the levothyroxine absorption test, two blood samples taken at baseline and at the peak (3 h according to this study) for serum FT4 after 1000 μg of levothyroxine ingestion would be a simple, convenient, and cost-effective way of evaluating refractory hypothyroidism before undergoing extensive investigations for malabsorption in a low-resource setting.
Declaration of interest
The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the study reported.
Funding
This study received funding support for blood investigations with test kits for FT4 and TSH supported by the research fund of the diabetes and endocrine unit of National Hospital Kandy.
Acknowledgements
We would like to acknowledge the patients for participating in the study and the staff of the diabetic and endocrine unit of the National Hospital Kandy for their support during the levothyroxine absorption test.
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