Glycerol Potentiates the Effects of Glucose in Promoting Glucose Recovery During Hypoglycemia: From Basic to Clinical Investigations and Their Therapeutic Application

Abstract

We compared the effect of oral glucose versus oral glucose combined with glycerol (glucose + glycerol) in promoting glucose recovery during hypoglycemia. These studies were carried out in two series of experiments. In the first series of experiments, 16 overnight fasted rats received an intraperitoneal injection of lispro insulin (1 IU/kg), and 25 min later, they received oral water (control), glucose (0.25 g/kg), glycerol (2.5 g/kg), or glucose (0.25 g/kg) + glycerol (2.5 g/kg). In the second series of experiments on 164 eligible type 1 diabetic (T1D) patients, 30 individuals with a history of hypoglycemia were recruited. Five volunteers did not meet the inclusion criteria and two subjects were excluded after starting the clinical investigation; 23 patients concluded the study. All patients with symptoms of hypoglycemia ingested oral glucose (15 g) or glucose (15 g) + glycerol (9.45 g). To treat hypoglycemia in T1D patients, preparations containing glucose alone or glucose + glycerol were used alternately (2 weeks/2 weeks) in a double-blind crossover scheme. Throughout the clinical research (4 weeks), glucose concentrations were assessed with a continuous glucose monitoring device and the results after the use of glucose alone or glucose + glycerol preparations were compared. Oral glucose combined with glycerol was more effective in promoting glucose recovery in comparison with glucose alone, not only in rats but also in T1D patients. Taken together, our experimental and clinical investigations reported the best performance of oral administration of glucose + glycerol in comparison with isolated glucose.

Introduction

The absence of rigorous control of glycemia predisposes the type 1 diabetic (T1D) patients to a series of chronic complications that include nephropathy, retinopathy, neuropathy, cerebrovascular accident, and cardiovascular disease.^1,2 Therefore, the main objective in the treatment of T1D patients is to reach glycemic values that are close to nondiabetic subjects.³

Two important, classical multicenter studies, the Diabetes Control and Complications Trial (DCCT) and the United Kingdom Prospective Diabetes Study (UKPDS), demonstrated the benefits gained with rigorous glycemic control in the treatment of diabetes. The DCCT⁴ demonstrated that maintenance of glycemia close to normal levels during intensive insulin therapy reduces the incidence and seriousness of chronic complications in T1D patients. The UKPDS⁵ demonstrated similar results in the treatment of type 2 diabetes. However, in the DCCT⁴ and UKPDS,⁵ the diabetic patients presented an increased frequency of hypoglycemia, defined by the American Diabetes Association³ as a condition in which glycemia is <70 mg/dL.

In general, during hypoglycemia, autonomic symptoms (profuse sweating, tremors, tachycardia, and facial flushing, etc.) and neuroglycopenic symptoms (lack of attention, confusion, exhaustion, weakness, headaches, inappropriate behavior, vision abnormalities, convulsions, and coma) occur.^6
–8

The occurrence, frequency, and intensity of episodes of severe hypoglycemia are strongly interconnected with an earlier introduction of insulin therapy, alterations in dose and type of insulin, insulin injection regime, and exercise.^4,8,9

It is interesting to note that when the episodes are repeated in short periods, there is a loss of perception of the symptoms of hypoglycemia. The mechanisms of this phenomenon are not yet completely understood and may disappear as adjustments in insulin therapy prevent the occurrence of new episodes of hypoglycemia.^10

–13

Thus, the risk of hypoglycemia represents the main limiting factor in the effort in seeking ideal glycemic control.^14,15 Therefore, it is important to treat hypoglycemia during insulin therapy.

Considering that parenteral glucagon is very expensive and has reduced availability, glucose is the main antidote to treat hypoglycemia. However, oral glucose shows limited effects and new treatments must be considered.

In this context, we verified the possibility of improving glycemic recovery using combined administration of oral glucose + glycerol instead of oral glucose alone to treat hypoglycemia not only in animal laboratories but also in T1D patients.

Materials and Methods

Animals

We used the same experimental procedure established in our laboratory 26 years ago.¹⁶ Before the experiments, adult male Wistar rats, weighing ∼200 g, were housed under a controlled 12-h light/12-h dark cycle, at a temperature of 23 ± 2°C, and with free access to water and food (Nuvilab^®, Curitiba, Brazil). All animals were maintained in these conditions until the day before the experiment when they were fasted for 15 h (5 p.m. to 9 a.m.) with the purpose of reducing the intestinal absorption of glucose.

Hypoglycemia was induced by intraperitoneal injection of lispro insulin (1 IU/kg). The insulin (Humalog^®) was not diluted and it was injected with an infusion pump. We chose the dose of insulin based on previous studies.^16,17

The animals (total = 16) were randomly selected to receive water (water group), glucose 0.25 g/kg dissolved in water (glucose group), glycerol 2.5 g/kg dissolved 1:1 in water (glycerol group), or glucose 0.25 g/kg + glycerol 2.5 g/kg dissolved 1:1 in water (glucose + glycerol group). Water, glucose, and glycerol (total volume = 1 mL) were orally administered (gavage) 25 min after insulin injection.

Glycemia was measured by analyzing blood obtained from the tail of each animal at 0, 25, and 60 min after insulin injection. The time, 0 min, provides blood glucose values before administration of insulin. Thus, a drop of blood was obtained after a small incision was made at the tip of the tail, and glycemia was evaluated using a home glucometer (Diagnostics Accu-Chek^® Active glucometer—Roche Diagnostics, Jaguaré, Brazil). This method was chosen because it is extremely rapid and does not appreciably influence glycemia.¹⁸ Moreover, it permits obtaining glycemia values using a limited number of rats, following the tendency to drastically reduce the number of animals.

During the time required for the experiment, all rats were maintained in the fasted state.

The protocol was approved by the Animal Ethics Committee of our University following the Brazilian Directives on Research Animal Protection and Experimentation.

Patients

A double-blind, randomized, crossover clinical trial was conducted in the municipality of Maringá, Paraná, Brazil. Participants were selected from a total of 164 eligible T1D patients of Clinics and Research in Endocrinology of Maringá city (CLIPEM). Institutional review board (IRB) approval: 86984418.0.0000.0104.

The ethical principles set forth by Resolution 196/96 of the Brazilian National Health Council were followed and the clinical protocol was evaluated by the State University of Maringá Standing Committee on Human Subject Research Ethics (COPEP).

Eligibility criteria

Patients of either gender were invited based on their medical records, particularly the history of hypoglycemia. The eligibility criteria also included patients aged 18–80 years and who agree to participate in all proposed study activities and sign two copies of the Informed Consent Form.

Exclusion criteria

Exclusion criteria included the following: type 2 diabetes or any other type of diabetes that cannot be classified as type 1 diabetes; liver disease and/or aspartate aminotransferase (ALT) and/or alanine aminotransferase (AST) levels ≥3 times the upper limit of normal; infectious or contagious disease; cardiovascular disease, including a history of stroke, decompensate hypertension, or heart failure; severe kidney disease; smoking, alcoholism, or illicit drug use; severe psychiatric disorder; and inability to understand the study protocol.

Thirty prediagnosed T1D patients were recruited. Five patients did not meet the inclusion criteria, two subjects were excluded after starting the study, and 23 patients concluded the study.

Preparations containing glucose or glucose + glycerol were used alternately (2 weeks/2 weeks) in a double-blind crossover scheme. For this purpose, the patients were randomized into groups 1 and 2. After implantation of a sensor to measure glucose levels, group 1 received glucose (n = 12) and group 2 received glucose + glycerol (n = 11) during the 1st and 2nd weeks. After this period, a new sensor was implanted in each patient and group 1 received glucose + glycerol (n = 12) and group 2 received glucose (n = 11) for two additional weeks. Glucose and glucose + glycerol were used only after symptoms of hypoglycemia. All this information has been condensed in a flow diagram (Fig. 1).

FIG. 1.

Study flow.

Considering that the detection and treatment of hypoglycemia are indicated for patients with glycemia <70 mg/dL, this value was considered as a criterion of identification and treatment.

All individuals were submitted to the same clinical protocols and ethical procedures, including guidelines regarding (1) standard diet—isocaloric diet [25 kcal/(kg/d)], divided into six daily meals, consisting of 55% carbohydrates, 30% fat (10% saturated), and 15–20% protein; during the intervention period, subjects should refrain from consuming alcoholic beverages or rapidly absorbed simple sugars; and (2) aerobic physical activity—preferably walking, 30–60 min/day, at a speed of 4–6 km/h.

Patients: anthropometric and laboratory data and medical records

Anthropometric data (body–mass index [BMI] and abdominal circumference), and medical records (age, gender, race, duration of diabetes, the total daily dose of insulin, mean arterial pressure, the existence of hypertension, and other chronic complications of diabetes) were evaluated.

The results of laboratory tests performed up to 6 months before starting the study were accepted, which include fasting glycemia, glycated hemoglobin (HbA1c), C-peptide, creatinine, total cholesterol, high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, triacylglycerol, thyroid-stimulating hormone (TSH), AST, ALT, hemogram, and urinalysis.

A survey of the medical records of T1D subjects was performed, and the following data were collected and recorded in a specific worksheet: disease duration, clinical and laboratory data, and intercurrent health events emerging in the last 24 months.

Preparation of glucose and glucose + glycerol solutions

Glucose solution and glucose + glycerol solution were obtained from a compounding pharmacy (Pharmacy School of Maringá State University, Maringá, Paraná State, Brazil), in compliance with the current applicable legislation. The glucose solution comprised 15 g of glucose dissolved in 30 mL of water packaged in unit-dose vials.

The glucose + glycerol solution comprised 15 g of glucose and 9.45 g of glycerol (total volume 30 mL) packaged in unit-dose vials. Thus, a glucose + glycerol solution was prepared as follows: 15 g glucose +7.5 mL glycerol +22.5 mL water.

Evaluation of glucose concentrations

A continuous blood glucose monitoring device (FreeStyle Libre^®; Abbott) was used to monitor trends of the glucose level in the interstitial fluid, with measurements every 15 min after implantation of a subcutaneous sensor that records glucose values for 14 days. Moreover, a capillary blood glucose monitoring device (FreeStyle^® Optium; Abbott) was used to compare with glucose results of the FreeStyle Libre monitor in case of questionable glucose measurements or suspected hypoglycemia.

Patients: intervention protocols and procedure

At visit 1, the first sensor was implanted and each participant received a kit containing one monitor, 50 test strips for capillary blood glucose measurement, an autolancing device, and material for skin preparation. Each sensor was used for 14 days.

In addition, each patient of group 1 received a kit containing 15 unit-dose vials with glucose solution and each patient of group 2 received a kit containing 15 unit-dose vials with glucose + glycerol solution for treatment of hypoglycemia.

Then, they received instructions on the use of the FreeStyle Libre monitor and sensors, as well as about the preparations for treatment of any episodes of hypoglycemia that they may experience.

All hypoglycemia symptoms were self-reported and treated by the patients. For this reason, during visits 1 and 2, all patients received a sheet to inform details about the hypoglycemic episode (symptoms of hypoglycemia, capillary glucose levels, time of hypoglycemic symptoms, and use of an additional food).

At visit 2, 14 days after starting the study, the first sensor was removed and returned to the research team for analysis of monitor records. The second sensor was implanted. All participants had their clinical data reassessed and were invited to report any hypoglycemia and/or adverse or intercurrent events that may have occurred. Considering that the preparations containing glucose alone or glucose + glycerol were used alternately (2 weeks/2 weeks), each patient of group 1 received a kit containing 15 unit-dose vials with glucose + glycerol solution and each patient of group 2 received a kit containing 15 unit-dose vials with glucose solution.

At visit 3, 28 days after starting the study, the second sensor was removed and returned to the investigators for assessment and interpretation of the records as well as a review of the answers and intercurrent events reported by each patient. Clinical data from each participant were reassessed at visit 3.

We did not observe severe hypoglycemia characterized by hospital internment and/or use of parenteral glucose or glucagon.

The primary efficacy endpoint was glucose recovery (glycemia above 100 mg/dL).

The study design is shown in Figure 2.

FIG. 2.

Study algorithm. ALT, aspartate aminotransferase; AST, alanine aminotransferase; BMI, body–mass index; HbA1c, glycosylated hemoglobin; TSH, thyroid-stimulating hormone.

Statistical analyses

Analysis of variance was used for intragroup (one-way) and intergroup (two-way) comparisons of glycemia values in rats. The Wilcoxon nonparametric test was used for comparisons in T1D patients. A 95% level of confidence (P < .05) was accepted for all comparisons. Results are reported as mean ± standard deviation.

The statistical analyses were performed in R language (version 4.0) using the stats package, R Core Team (2020); R: a language and environment for statistical computing, R Foundation for Statistical Computing, Vienna, Austria.

Results

The results of experiments in rats are given in Table 1, in which glucose recovery rates after water, glucose, glycerol, or glucose + glycerol are compared.

Table 1.

Glucose Levels (mg/dL) at 0, 25, and 60 min After 1 IU/kg Insulin Injection

	Water	Glucose	Glycerol	Glucose + glycerol
0 min	70.5 ± 4.6	70.6 ± 6.4	74.2 ± 7.2	68.2 ± 16.0
25 min	48.3 ± 11.4^*	45.0 ± 5.8^*	45.3 ± 4.4^*	37.2 ± 12.0^*
60 min	33.7 ± 5.0^* ^, ^**	40.2 ± 11.0^*	45.5 ± 4.8^*	52.4 ± 9.0^**

Water (water group), 0.25 g/kg glucose (glucose group), 2.5 g/kg glycerol (glycerol group), or 0.25 g/kg glucose +2.5 g/kg glycerol (glucose + glycerol group) was orally administered 25 min after insulin injection in Wistar rats. Values are expressed as mean ± standard deviation (n = 4).

^* P < .05 in comparison with 0 min (same column).

^** P < .05 (25 vs. 60 min—same column).

Insulin administration caused a rapid and marked fall (P < .05) in glycemia between 0 and 25 min. Glycemia recovery (P < .05) was observed 35 min after oral administration of glucose + glycerol, that is, 60 min after the insulin injection. However, no glycemia recovery was observed after oral administration of water or glucose (Table 1).

The results of clinical studies are given in Tables 2 –5. The general characteristics of patients are condensed in Table 2. Patients (65.2% women and 34.8% men) had a mean age of 38.5 ± 12.1 years (range 18–66 years). The duration of diabetes was 23.2 ± 10.3 years (range 4–38 years). The total daily dose of insulin was 0.77 ± 0.19 IU/kg (range 0.48–1.23). The average BMI for women and men was 25.2 and 25.7 kg/m², respectively. The average abdominal circumference for women and men was 87.4 and 94.1 cm, respectively. Mean arterial pressure (MAP) was 85.9 ± 5.01 mmHg. Regarding the presence of chronic complications of diabetes, 13.0% had retinopathy, 8.7% nephropathy, 4.3% retinopathy plus nephropathy. The most prevalent comorbidities were primary hypothyroidism (39.1%) and arterial hypertension (17.4%).

Table 2.

General Characteristics of the Patients

Patient characteristics	n = 23
AC (cm)^a	89.7 ± 9.9
Age (years)	38.5 ± 12.1
Arterial hypertension	4 yes/19 no (17.4%/82.6%)
BMI (kg/m²)^a	25.4 ± 3.6
DD (years)^a	23.2 ± 10.3
Gender (male/female)	8/15 (34.8%/65.2%)
MAP (mmHg)^a	85.9 ± 5.0
Primary hypothyroidism	9 yes/13 no (39.1%/61.9%)
Race (white/nonwhite)	21/2 (91.3%/8.7%)
TDDI (IU)^a	0.77 ± 0.19
CDC (No/Ret/Neph/Ret plus Neph)	17/3/2/1 (74.0%/13.0%/8.7%/4.3%)

AC, age, BMI, DD, MAP, and TDDI are means ± standard deviations considering visits 1, 2, and 3.

AC, abdominal circumference; BMI, body–mass index; CDC, chronic diabetes complications; DD, duration of diabetes; MAP, mean arterial pressure; Neph, nephropathy; Ret, retinopathy; TDDI, total daily dose of insulin.

The results of the laboratory evaluation are given in Table 3. The mean values ± standard deviations of the HbA1c level, fasting glycemia, and C-peptide were 7.32 ± 0.87%, 137 ± 51.1 mg/dL, and 0.08 ng/mL ±0.07, respectively. It must be emphasized that all individual values of C-peptide are compatible with absolute insulinopenia (results not shown).

Table 3.

Laboratory Evaluation

Laboratory data	n = 23
Fasting glycemia (mg/dL)	137.0 ± 51.1
HbA1c (%)	7.32 ± 0.87
C peptide (ng/mL)	0.08 ± 0.07
Creatinine (mg/dL)	0.87 ± 0.19
Total cholesterol (mg/dL)	169.9 ± 30.0
HDL (mg/dL)	55.9 ± 13.1
LDL (mg/dL)	97.8 ± 22.6
Triacylglycerol (mg/dL)	79.0 ± 42.2
TSH (μIU/mL)	2.46 ± 1.29

Data are expressed as the means ± standard deviations.

HbA1c, glycosylated hemoglobin; HDL, high-density lipoprotein; LDL, low-density lipoprotein; TSH, thyroid-stimulating hormone.

The mean values ± standard deviations of total cholesterol, HDL and LDL, and triacylglycerol were 170.0 ± 30.0, 55.9 ± 13.1 and 97.8 ± 22.6 mg/dL, and 79.0 ± 42.2 mg/dL, respectively. The creatinine and TSH levels were 0.87 ± 0.19 mg/dL and 2.46 ± 1.29 μUI/mL, respectively. Blood count with platelet count, ALT, AST, and urinalysis did not show significant changes (results not shown).

Glucose recovery (blood concentration >100 mg/dL) after ingestion of glucose or glucose plus glycerol was observed in 52% of the episodes of hypoglycemia (total = 187).

The majority of episodes of glucose recovery (glycemia >100 mg/dL) were observed after ingestion of glucose + glycerol (59%) in comparison with the isolated ingestion of glucose (41%) (Table 4).

Table 4.

The Number of Diurnal and Nocturnal Episodes of Hypoglycemia and the Number of Episodes of Glucose Recovery Where Oral Glucose or Oral Glucose + Glycerol Increased Glucose Levels Above 100 mg/dL

	ND	NG	NGG		NN	NG	NGG
06:00–06:30	12	1	6	18:00–18:30	3	0	1
06:30–07:00	8	0	5	18:30–19:00	0	0	0
07:00–07:30	5	1	2	19:00–19:30	6	2	3
07:30–08:00	8	2	3	19:30–20:00	1	1	0
08:00–08:30	4	0	3	20:00–20:30	4	3	1
08:30–09:00	4	0	2	20:30–21:00	0	0	0
09:00–09:30	5	2	2	21:00–21:30	3	0	1
09:30–10:00	3	0	2	21:30–22:00	5	2	1
10:00–10:30	5	1	2	22:00–22:30	5	4	1
10:30–11:00	1	1	0	22:30–23:00	3	0	1
11:00–11:30	6	3	0	23:00–23:30	1	0	0
11:30–12:00	4	1	0	23:30–00:00	0	0	0
12:00–12:30	4	0	1	00:00–00:30	8	2	0
12:30–13:00	1	0	0	00:30–01:00	7	2	1
13:00–13:30	4	0	0	01:00–01:30	4	1	1
13:30–14:00	2	0	0	01:30–02:00	1	1	0
14:00–14:30	3	0	1	02:00–02:30	0	0	0
14:30–15:00	4	1	2	02:30–03:00	2	0	0
15:00–15:30	10	1	2	03:00–03:30	1	1	0
15:30–16:00	11	3	2	03:30–04:00	2	0	2
16:00–16:30	4	0	1	04:00–04:30	2	2	0
16:30–17:00	4	0	0	04:30–05:00	6	0	2
17:00–17:30	2	1	0	05:00–05:30	2	0	2
17:30–18:00	2	0	2	05:30–06:00	5	1	2
	116	18	38		71	22	19
Total: 187

Glucose concentration values of 23 type 1 diabetic patients evaluated for 4 weeks.

ND, number of diurnal episodes; NG, number of oral glucose; NGG, number of oral glucose + glycerol; NN, number of nocturnal episodes.

We observed an elevation (P < .05) of glucose concentrations 30 and 60 min after ingestion of glucose or glucose + glycerol. Patients who ingested glucose + glycerol showed higher (P < .05) glucose levels in comparison with those patients who ingested glucose alone. The elevation of glucose levels (time 0 min vs. time 60 min) after oral ingestion of glucose or glucose + glycerol was 102% and 137%, respectively (Table 5).

Table 5.

Glucose Levels (mg/dL) at 0, 15, 30, and 60 min After Ingestion of Glucose (15 g) or Glucose (15 g) + Glycerol (9.45 g) at Any Time of the Day Over 4 Weeks of Evaluation in Type 1 Diabetic Patients

	Glucose	Glucose + glycerol
0 min	50.8 ± 9.8	49.3 ± 9.3
15 min	58.8 ± 17.8	56.2 ± 16.5
30 min	81.0^* ^, ^** ±29.2	87.1^* ^, ^** ±27.2
60 min	102.7^* ^, ^ ^, ^* ±39.6	116.9^* ^, ^ ^, ^* ^,† ± 39.1

Evaluations were done immediately before (0 min), 15 min, 30 min, and 60 min after ingestion of glucose (n = 70) or glucose + glycerol (n = 70). Values are expressed as mean ± standard deviation.

P < .05 in comparison with 0 min (same column).

P < .05 in comparison with 15 min (same column).

***

P < .05 in comparison with 30 min (same column).

†

P < .05 (glucose vs. glucose + glycerol − same line).

Discussion

Glycerol is a pharmaceutical excipient for several active substances, including a majority of commercial preparations of insulin (Tresiba^®, Lantus^®, Levemir^®, Humulin R^®, Novolin N^®, Humalog, and NovoRapid^®). However, an increased amount of glycerol changes its role from a single excipient to an important caloric source (4.32 kcal/g) and gluconeogenic precursor.

The best performance of the combination of glucose + glycerol in comparison with glucose in our preclinical and clinical studies could be explained, partly at least, by the difference in the number of calories. For example, the number of calories used in the clinical study was 60 kcal and 100.8 kcal for glucose alone and glucose + glycerol, respectively, which represents 40.8 additional kilocalories. Furthermore, glycerol has free permeability across cell membranes¹⁹ being used in the glycolysis and Krebs cycle to produce energy in the brain,²⁰ erythrocytes,²¹ and skeletal muscle.²² In addition, the contribution of renal glucose production from glycerol must be considered.²³

However, the main mechanism by which glycerol promotes glucose recovery probably is its role as a liver glucose precursor.^24,25

It is well established that insulin inhibits liver glycogenolysis and gluconeogenesis.^26

–29 However, during hypoglycemia, the availability of liver glucose precursors in the blood and the liver's ability to produce glucose from gluconeogenic substrates are maintained, opening the possibility of using liver glucose precursors (glycerol, lactate, alanine, and glutamine) to treat hypoglycemia.^{30

–38}

Interestingly, we previously reported (in livers of hypoglycemic rats) that glucose administration decreased glucose production from lactate, alanine, or glutamine, but glucose production from glycerol is maintained.²⁴ This peculiar response could be attributed to the fact that glycerol enters this metabolic pathway at the triose phosphate step, after the phosphoenolpyruvate carboxylase step,^24,39,40 which is inhibited by insulin.

Therefore, the best glucose recovery during hypoglycemia, not only in rats (Table 1) but also in T1D patients (Table 5), promoted by oral glucose + glycerol in comparison with oral glucose alone is a consequence of the maintained liver conversion of glycerol to glucose after glucose ingestion.

In summary, rats that received oral glucose + glycerol or patients who ingested glucose + glycerol showed higher glucose levels, that is, greater glucose recovery, compared with isolated glucose intake. These results demonstrated that glycerol potentiates the effects of glucose in promoting glucose recovery during hypoglycemia in rats and T1D patients. The better glucose recovery during hypoglycemia promoted by oral glucose + glycerol was a consequence of the maintained conversion of glycerol to glucose in the liver, after glucose intake, as we previously demonstrated.²⁴

The main limitation of this study was the small sample size not only of rats but also T1D patients. Additional limitations of the study include the use of a unique dose, evaluations of glycemia restricted to a short period (60 min), only 52% of glucose recovery (glycemia >100 mg/dL) after ingestion of glucose or glucose + glycerol, lack of generalizability to other hypoglycemic conditions, and absence of information about blood parameters other than glucose.

The importance of this study is shown in Table 4, describing 187 episodes of hypoglycemia (116 diurnal +71 nocturnal episodes) with ingestion of glucose or glucose + glycerol. It means two episodes per week, that is, about 100 episodes per T1D patient for 1 year.

Taken together, our preclinical model and clinical investigation reported the best performance of oral administration of glucose + glycerol in comparison with isolated glucose in the treatment of hypoglycemia. Further studies will elucidate more precisely how oral administration of glucose + glycerol alleviates hypoglycemia.

Footnotes

Acknowledgment

The authors are grateful to Mr. Carlos Eduardo de Oliveira for his technical assistance.

Author Disclosure Statement

There is a patent application forwarded on behalf of the State University of Maringá, Brazil.

Funding Information

Research supported by the Brazilian research agencies: Nucleus of Excellence Support Program (PRONEX)/National Council for Scientific and Technological Development (CNPq)/Araucaria Foundation - grant number 249/2013, Coordination for the Improvement of Higher Education Personnel (CAPES) - Grant number 88882.448882/2019-01.

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