Jimmi Elers

Determination of salbutamol and salbutamol glucuronide in human urine by means of liquid chromatography-tandem mass spectrometry.

The determination of salbutamol and its glucuronide in human urine following the inhalative and oral administration of therapeutic doses of salbutamol preparations was performed by means of direct urine injection utilizing liquid chromatography-tandem mass spectrometry (LC-MS/MS) and employing d(3)-salbutamol and d(3)-salbutamol glucuronide as internal standards. Unconjugated salbutamol was detected in all administration study urine samples. Salbutamol concentrations following inhalation were commonly (99%) below 1000 ng/ml whereas values after oral administration frequently (48%) exceeded this threshold. While salbutamol glucuronide was not detected in urine samples collected after inhalation of the drug, 26 out of 82 specimens obtained after oral application contained salbutamol glucuronide with a peak value of 63 ng/ml. The percentage of salbutamol glucuronide compared to unconjugated salbutamol was less than 3%. Authentic doping control urine samples indicating screening results for salbutamol less than 1000 ng/ml, showed salbutamol glucuronide concentrations between 2 and 6 ng/ml, whereas adverse analytical findings resulting from salbutamol levels higher than 1000 ng/ml, had salbutamol glucuronide values between 8 and 15 ng/ml. The approach enabled the rapid determination of salbutamol and its glucuronic acid conjugate in human urine and represents an alternative to existing procedures since time-consuming hydrolysis or derivatization steps were omitted. Moreover, the excretion of salbutamol glucuronide in human urine following the administration of salbutamol was proven.

The pharmacokinetic profile of inhaled and oral salbutamol in elite athletes with asthma and nonasthmatic subjects.

Objective Data on pharmacokinetics of inhaled and oral salbutamol in elite athletes with asthma are needed to differentiate between therapeutic use and doping in doping control. Design An interventional open-label crossover. Setting Respiratory Research Unit, Copenhagen University Hospital, Bispebjerg. Participants Eight elite athletes with asthma and 10 nonasthmatic subjects aged 18 to 33 years. Intervention Administration of 0.8 mg of inhaled salbutamol and 8 mg of oral salbutamol separated by 14 days. Main Outcome Measures Urine concentration of free salbutamol. Results Maximum urine concentrations peaked in the period of 0 to 4 hours after the administration of inhaled and oral salbutamol in both groups. Median concentrations after inhaled salbutamol and oral salbutamol were 401.6 and 2108.1 ng/mL in healthy subjects and 334.9 and 2975.2 ng/mL in elite athletes with asthma. There were no significant statistical differences between the groups. One sample exceeded the World Anti-Doping Agency threshold value of 1000 ng/mL with a urinary salbutamol concentration of 1057 ng/mL 4 hours after inhalation, when no correction for urine specific gravity was done. When this sample was corrected for urine specific gravity, the result was 661 ng/mL. Conclusions We found no significant difference in pharmacokinetic profile of inhaled and oral salbutamol between elite athletes with asthma and nonasthmatic subjects. Our results indicate that urine salbutamol concentrations should be corrected for urine specific gravity when evaluating doping cases.

Urine and serum concentrations of inhaled and oral terbutaline.

We examined urine and serum concentrations after therapeutic use of single and repetitive doses of inhaled and supratherapeutic oral use of terbutaline. We compared the concentrations in 10 asthmatics and 10 healthy subjects in an open-label, cross-over study with 2 mg inhaled and 10 mg oral terbutaline on 2 study days. Further, 10 healthy subjects were administrated 1 mg inhaled terbutaline in 4 repetive doses with total 4 mg. Blood samples were collected at baseline and during 6 h after the first inhalations. Urine samples were collected at baseline and during 12 h after the first inhalations. Median (IQR) urine concentrations peaked in the period 0-4 h after inhalation with Cmax 472 (324) ng/mL in asthmatics and 661 (517) ng/mL in healthy subjects, and 4-8 h after oral use with Cmax 666 (877) ng/mL in asthmatic and 402 (663) ng/mL in healthy subjects. In conclusion we found no significant differences in urine and serum concentrations between asthmatic and healthy subjects. We compared urine and serum concentrations after therapeutic inhaled doses and supratherapeutic oral doses and observed significant statistical differences in both groups but found it impossible to distinguish between therapeutic and prohibited use based on doping tests with urine and blood samples.

Blood and Urinary Concentrations of Salbutamol in Asthmatic Subjects

PURPOSE Data on blood and urinary concentrations of salbutamol after inhalation and oral administration in healthy subjects are scarce. Accordingly, we examined the pharmacokinetics of inhaled and oral salbutamol in asthmatic subjects. METHODS We enrolled 10 men aged 18-45 yr in an open-label study in which 0.8 mg of inhaled or 8 mg of systemic salbutamol was administered in a crossover design. All subjects had doctor-diagnosed asthma, used beta2 agonist when needed, and abstained from any medicine, beta2 agonist inclusive, for 14 d before visit. Urine was collected from all subjects (0-4, 4-8, and 8-12 h), and blood samples were taken at 0, 0.5, 1, 2, 3, 4, and 6 h after salbutamol administration. RESULTS Maximum urine concentration was reached during the first 4 h after administration of both inhaled and oral salbutamol. We found differences in median urinary concentrations (Cmax) of 260.9 and 2422.2 ng x mL(-1), respectively (P < 0.005). Urinary concentrations show high individual variability irrespective of the route of administration. Blood analyses showed a systemic exposure of salbutamol after both inhaled and oral salbutamol with peak concentration after inhalation before the oral intake (P < 0.05). A difference in median Cmax after inhalation and oral treatment was found: 1.75 and 18.77 ng x mL(-1), respectively (P < 0.05). CONCLUSIONS Median urinary concentrations after oral administration of 8 mg of salbutamol were significantly higher than those after inhalation of 0.8 mg of salbutamol.

Urine Concentrations of Repetitive Doses of Inhaled Salbutamol

We examined blood and urine concentrations of repetitive doses of inhaled salbutamol in relation to the existing cut-off value used in routine doping control. We compared the concentrations in asthmatics with regular use of beta2-agonists prior to study and healthy controls with no previous use of beta2-agonists. We enrolled 10 asthmatics and 10 controls in an open-label study in which subjects inhaled repetitive doses of 400 microgram salbutamol every second hour (total 1600 microgram), which is the permitted daily dose by the World Anti-Doping Agency (WADA). Blood samples were collected at baseline, 30 min, 1, 2, 3, 4, and 6 h after the first inhalations. Urine samples were collected at baseline, 0-4 h, 4-8 h, and 8-12 h after the first inhalations. Median urine concentrations peaked in the period 4-8 h after the first inhalations in the asthmatics and between 8-12 h in controls and the median ranged from 268 to 611 ng×mL (-1). No samples exceeded the WADA threshold value of 1000 ng×mL (-1) when corrected for the urine specific gravity. When not corrected one sample exceeded the cut-off value with urine concentration of 1082 ng×mL (-1). In conclusion we found no differences in blood and urine concentrations between asthmatic and healthy subjects. We found high variability in urine concentrations between subjects in both groups. The variability between subjects was still present after the samples were corrected for urine specific gravity.

Asthma in elite athletes

Asthma is frequently found among elite athletes performing endurance sports such as swimming, rowing and cross-country skiing. Although these athletes often report symptoms while exercising, they seldom have symptoms at rest. Moreover, compared with nonathletic asthmatic individuals, elite athletes have been shown to have a different distribution of airway inflammation and unequal response to bronchial provocative test. Elite athletes display signs of exercise-induced symptoms, for example, nonasthmatic inspiratory wheeze, vocal cord dysfunction and cardiac arrhythmias, which could limit their physical capacity. Elite athletes should undergo comprehensive assessment to confirm an asthma diagnosis and determine its degree of severity. Treatment should be as for any other asthmatic individual, including the use of β2-agonist, inhaled steroid as well as leukotriene-antagonist. It should, however, be noted that daily use of β-agonists could expose elite athletes to the risk of developing tolerance towards these drugs. Use of β2-agonist should be replaced with daily inhaled corticosteroid treatment, the most important treatment of exercise-induced asthma. All physicians treating asthma should be aware of the doping aspects. Systemic β2-agonist intake is strictly prohibited, whereas inhaled treatment is allowed in therapeutic doses when asthma is documented and dispensation has been granted when needed.

Formoterol concentrations in blood and urine: the World Anti-Doping Agency 2012 regulations.

INTRODUCTION We examined urinary and serum concentrations of formoterol in asthmatic and healthy individuals after a single dose of 18 μg inhaled formoterol and after repeated inhaled doses in healthy individuals. Results were evaluated using the World Anti-Doping Agency (WADA) 2012 threshold for formoterol. METHODS On the day of this open-label, crossover study, 10 asthmatic subjects who regularly used beta2-agonists and 10 healthy participants with no previous use of beta2-agonists received a single dose of 18 μg formoterol. Further, 10 nonasthmatic participants inhaled 18 μg formoterol every second hour until obtaining a total of 72 μg, which is twice the maximum daily dose (36 μg formoterol) permitted by the WADA. Blood samples were collected at baseline, 30 min, 1, 2, 3, 4, and 6 h after the first inhalation. Urine samples were collected at baseline, 0-4, 4-8, and 8-12 h after the first inhalation. RESULTS Median urine concentration, corrected for specific gravity, after the single-dose administration peaked during 0-4 h after inhalation at a maximum of 7.4 ng·mL(-1) in asthmatic subjects and 7.9 ng·mL(-1) in healthy subjects. Median urine concentration after repeated doses peaked during 4-8 h after inhalation of a total of 72 μg formoterol at a maximum of 16.8 ng·mL(-1) in healthy participants. The maximum individual concentration of 25.6 ng·mL(-1) was found after inhalation of a total of 72 μg formoterol. CONCLUSIONS We found no significant differences in urinary and serum concentrations of formoterol between asthmatic and healthy subjects. We found high interindividual variability in the concentrations in all groups. Our data support the WADA 2012 urinary threshold of 30 ng·mL(-1) formoterol as being an adverse analytical finding.

Daily Use of Salmeterol Causes Tolerance to Bronchodilation with Terbutaline in Asthmatic Subjects

Background: The purpose was to assess tolerance to terbutaline after daily use of long-acting β2- agonist (LABA) and further to evaluate two designs of reversibility test widely used in research and clinic in order to demonstrate tolerance. Methods: Twenty-eight asthmatics were given daily LABA in 12 weeks and were randomized to challenge test and either conventional reversibility test with 2 puffs terbutaline or reversibility test with refracted doses (1 puff) every 5 min, total 4 puffs. FEV1 was measured pre-challenge, post-challenge, during and after reversibility test. All subjects had 3 visits: baseline, after 4 weeks and after 12 weeks of LABA treatment. All subjects were non-smokers, aged 18–45 years and had a positive methacholine challenge. Results: The analyses showed a significant fall in reversibility after 4 and 12 weeks of LABA treatment (p=0.001) in the group with the conventional reversibility test. The group with reversibility test using refracted doses also showed a significant fall in reversibility after 4 weeks of LABA treatment (p=0.017) followed by a similar trend after 12 weeks (p=0.054), however, we experienced an interfering number of dropouts at the last visit. Conclusion: The bronchodilator response to terbutaline was significantly reduced in asthmatic subjects using daily LABA. The tolerance develops rapidly and is present after 4 weeks of treatment. Our study showed that both the conventional reversibility test and the reversibility test with refracted doses, combined with methacholine challenge is able to demonstrate tolerance to bronchodilator after daily use of LABA.

Urine Concentrations of Inhaled Salmeterol and its Metabolite a-Hydroxysalmeterol in Asthmatic and Non-Asthmatic Subjects

Salmeterol is a long-acting beta2-agonist, which is on the WADA prohibited list, but can be used by athletes in therapeutic doses by inhalation. The prohibited list, however, contains no urinary threshold for salmeterol, which gives athletes the opportunity to inhale unlimited doses of salmeterol. In doping controls, metabolites may be used as markers for misuse of substances. No studies have determined urine concentrations of α-hydroxysalmeterol, the metabolite of salmeterol. Furthermore, the metabolism and excretion of salmeterol may vary between asthmatics and nonasthmatics. We determined the serum and urinary concentrations of salmeterol and its metabolite α-hydroxysalmeterol after inhalation of 100 μg salmeterol in ten asthmatics and ten non-asthmatics. Blood samples were collected at baseline and ½, 1, 2, 3, 4 and 6 hours after the administration of salmeterol. Urine samples were collected at baseline and 4, 8 and 12 hours after administration. The urinary concentration of salmeterol following enzymatic hydrolysis of the glucuronide fraction was 0.38 ± 0.26 ngmL-1 in asthmatics and 0.38 ± 0.22 ngmL-1 in non-asthmatics, 4 hours after inhalation. The highest median serum concentration (Cmax) was 0.07 ± 0.03 ngmL-1 in asthmatics after 30 minutes (Tmax) and 0.06 ± 0.03 ngmL-1 in non-asthmatics. No statistical differences were found in Cmax or Tmax of salmeterol between asthmatics or non-asthmatics in neither the serum nor urine samples. The highest median urinary concentration of α-hydroxysalmeterol following enzymatic hydrolysis of the glucuronide fraction was 2.86 ± 1.75 ngmL-1 in asthmatics, 4 hours after inhalation of salmeterol. In non-asthmatics, it was 2.73 ± 2.08 ngmL-1. The asthmatics had a significantly (p<0.05) higher concentration after 12 hours compared with non-asthmatics. In doping control, α-hydroxysalmeterol may be a more suitable marker for excessive use of inhaled salmeterol, due to its higher concentration in urine.