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Evaluation of the sedative effects of diazepam, midazolam, and xylazine after intranasal administration in juvenile ostriches (Struthio camelus).

Abstract: The sedative effects of diazepam, midazolam, and xylazine after intranasal administration were evaluated in 72 (36 male and 36 female) juvenile healthy ostriches (Struthio camelus), weighing 50-61 kg and aged 4-5 months. The birds were randomly divided into 3 groups (n = 24), then each group was further subdivided to 4 subgroups (n = 6). For each drug, 4 different doses were chosen and the total calculated dose was equally administered into either naris of the individual bird. The appropriate dose of each drug to produce standing chemical restraint or sternal recumbency was evaluated based on the onset time, the duration of maximum effect, and the duration of sedation. Midazolam showed significantly shorter onset time (2.9 [+ or -] 1.2 minutes) compared with xylazine (4.4 [+ or -] 1 minute) and diazepam (4.3 [+ or -] 0.4 minutes). Longer duration of sedation was also achieved with midazolam compared with xylazine and diazepam. Moderate sedation was achieved with diazepam (0.8 mg/kg), midazolam (0.4 mg/kg), and xylazine (2 mg/kg) for standing chemical restraint, with the maximum duration effects of 7.0 [+ or -] 1.4, 17.7 [+ or -] 4.1, and 9.2 [+ or -] 2.5 minutes, respectively. Deep sedation was also achieved with midazolam (0.8 mg/kg) and xylazine (4 mg/kg), with sternal recumbency duration of 21.7 [+ or -] 4.9 and 13.5 [+ or -] 2.6 minutes, respectively. The results of the present study show that intranasal administration can be an effective route for delivery of sedatives in juvenile ostriches. Intranasal midazolam and xylazine could be suggested for standing chemical restraint or inducing sternal recumbency in juvenile ostriches.

Key words: intranasal, sedation, diazepam, midazolam, xylazine, avian, ostrich, Struthio camelus


The ostrich (Struthio camelus) is the largest living bird and belongs to the ratite family. (1,2) Ostrich farming is an economic activity worldwide. (1) Ostriches are highly susceptible to stress, and handling of adult birds, especially males, can be very difficult. (1-3) Physical examination is commonly done by hooding, a well-known restraint procedure in ostriches. However, to perform surgical procedures or for more invasive or potentially painful procedures, chemical restraint is necessary. (4,5) Chemical immobilization is usually achieved by the use of sedatives administered by intramuscular or intravenous routes, which require technical expertise and can result in economic losses due to injury. (2,5,6)

The benzodiazepines diazepam and midazolam have anxiolytic, anticonvulsant, amnesic, and sedative effects in birds. (2) Although benzodiazepines are used for premedication, they are commonly used for procedural sedation. (7-9) Xylazine, an [[alpha].sub.2]-adrenergic agonist, has been used in birds for its sedative and analgesic properties. (10) Although [[alpha].sub.2]-adrenergics, benzodiazepines, and opioids have been used as preanesthetic agents in ostriches, (4,11-14) there are few experimental studies concerning sedatives in clinically normal ostriches. (5,6)

The intranasal route is an acceptable alternative for administration of sedatives in veterinary and human medicine. (15-26) Furthermore, sedation has been achieved by the intranasal route in canaries CSerinus canarius), ring-necked parakeets (Psittacula krameri), pigeons (Columba lima domestica), Hispaniolan Amazon parrots (Amazona ventralis), budgerigars (Melopsittacus undulatus), and zebra finches (Taeniopygia guttata). (7-9,27-29) The aims of the present study were 2-fold: to investigate whether the intranasal route is effective for delivery of sedatives and to determine the appropriate dose of sedatives needed to produce moderate sedation for standing chemical restraint and deep sedation with sternal recumbency after intranasal administration of diazepam, midazolam, or xylazine in juvenile ostriches.

Materials and Methods


The birds used in this study were kept at a commercial farm in Mashhad, northeast of Iran. Seventy-two (36 males and 36 females) healthy juvenile ostriches 4-5 months of age and weighing 50-61 kg were selected randomly based on a random number table. The birds were transferred to 6 separate smaller paddocks of 120 [m.sup.2] each, with a controlled temperature of 18[degrees]C-20[degrees]C for 1 week of acclimatization before the beginning of the experiment. They had free access to water and a commercial diet for growing ostriches composed of alfalfa, barley, wheat, corn, and soybean, with an added supplement for growing ostriches of minerals, vitamins, and amino acids. Food was withheld for 12 hours before the beginning of the experiments. During the experiments, the birds were treated under standard conditions consistent with the Recommendations and Minimum Standards for the Welfare of Ostrich and Emu. (30) All procedures were done in accordance with the guidelines of the University Ethics Committee, and experiments were approved by the University Research Council.


This experiment was designed to assess the sedative effects of intranasal administration of diazepam (5 mg/mL), midazolam (5 mg/mL), and xylazine hydrochloride (20 mg/mL) in ostriches. The birds were randomly distributed (random number table) into 3 groups of 24 birds each to receive either diazepam, midazolam, or xylazine intranasally (IN). Then, each group was divided into 4 subgroups of 6 birds. For each drug in the present study, the lowest intramuscular or intravenous dose that has been previously used as a preanesthetic agent in ostriches was selected as an initial dose for intranasal administration. (4,11,31) For each drug, 4 different doses were selected (initial dose multiplied by 1,2, 4, and 8) and administered to a group of 6 birds. Each bird was used once.

Each ostrich was physically restrained by 2 experienced handlers; one held the head and neck and the other provided pressure to the bird's pelvis in an upward and forward direction. The total dose for each bird was calculated as follows: diazepam and midazolam administered at 0.2-0.8 mg/kg (1-4.9 mL/nostril) and xylazine administered at 1-4 mg/kg (1.3-6.1 mL/nostril). Approximately half the dose was administered over the course of 2 minutes into each naris with a syringe while the head of the ostrich was restrained in a slight upright position to prevent nasal runoff of drugs from the nares. The bird was then released and left alone in the pen for observation, without any further stimulation.

The sedative effects of each drug after intranasal administration were subjectively evaluated based on defined criteria. The onset time was defined as the time between drug administration and early signs of the bird's reaction to sedation, including closure of the eyelids, changes in posture (S-shaped neck, drooping wings and tail), loss of coordination, and ataxia. Duration of sedation was defined as the time between initial signs of sedation and full recovery, characterized by return to normal standing posture, alertness, and voluntary movements. The maximum duration effect of sedation was defined as the time span from when the bird did not show reaction to touch until the time that this effect was missing. The quality of sedation was graded as mild, moderate, and deep. Mild sedation was defined as a stage in which the bird showed early signs of sedation (closure of the eyelids; changes in neck, wings, and tail posture; and mild ataxia); however, it still responded to finger snap and handling. Moderate sedation was defined as the stage in which the bird allowed handling and physical examination without struggling. Deep sedation was defined as the stage in which the bird showed depression of consciousness, satisfactory muscle relaxation, and sternal recumbency.

Statistical analysis

Statistical analysis of data was performed by 2-way (3 x 3) analysis of variance (ANOVA). The data were initially checked for potential outliers by visual inspection of the boxplots. The assumptions for analysis were tested before running the ANOVA model by the Shapiro-Wilk test for normality and Levene's test for heteroscedasticity and then further rechecked graphically in the final models by inspection of residual plots. The Bonferroni test was used for post hoc analysis to adjust for multiple comparisons. A value of P < .05 was considered significant. The results are presented as mean [+ or -] standard deviation. Statistical analyses were carried out with IBM SPSS Statistics for Windows (Version 22, SPSS Inc, Chicago, IL).


Intranasal administration of diazepam, midazolam, or xylazine caused sedative effects in ostriches. The route of administration was well tolerated by the ostriches. No complications were observed during the period of study, and a smooth recovery from sedation was achieved; hence, we did not use any reversal agent (flumazenil/yohimbine).

Table 1 represents the onset time and the duration of sedation for drugs causing mild sedation. Mild sedation with a relatively short duration of action was achieved with diazepam at 0.2 and 0.4 mg/kg, midazolam at 0.2 mg/kg, and xylazine at 1 mg/kg. In the diazepam group, the onset time was significantly shorter (P <.001) at 0.4 mg/kg (4.9 [+ or -] 0.4 minutes) than at 0.2 mg/kg (6.1 [+ or -] 0.6 minutes). Duration of sedation was significantly longer (P <.05) with midazolam at 0.2 mg/kg (12.7 [+ or -] 3.7 minutes) than with diazepam at 0.2 (6.4 [+ or -] 0.6 minutes) and 0.4 mg/kg (9.0 [+ or -] 0.9 minutes) and xylazine at 1 mg/kg (8.3 [+ or -] 2.2 minutes). Overall, the mild sedation was not adequate for proper chemical restraint, and these differences were considered not clinically important.

Table 2 represents the onset time, maximum effect, and duration of sedation for drugs causing moderate sedation. Moderate sedation that was suitable for standing chemical restraint, and physical examination was achieved with diazepam at 0.8 mg/kg, midazolam at 0.4 mg/kg, and xylazine at 2 mg/kg. Birds in the midazolam group at 0.4 mg/kg showed a significantly shorter onset time (3.9 [+ or -] 0.2 minutes, P =.01) and longer maximum duration effect (17.7 [+ or -] 4.1 minutes, P < .001) than birds in the xylazine group at 2 mg/kg (4.9 [+ or -] 0.4 and 9.2 [+ or -] 2.5 minutes, respectively). Birds in the midazolam group at 0.4 mg/kg showed a significantly longer (P <.001) maximum duration effect (17.7 [+ or -] 4.1 minutes) than birds in the diazepam group at 0.8 mg/kg (7.0 [+ or -] 1.4 minutes).

Table 3 represents the onset time, the maximum effect, the duration of sternal recumbency, and the duration of sedation for drugs causing deep sedation. Deep sedation that was adequate for sternal recumbency was achieved with midazolam at 0.8 mg/kg and xylazine at 4 mg/kg. Midazolam showed significantly shorter onset time (1.9 [+ or -] 0.2 minutes, P <.001) and longer maximum duration effect (38.0 [+ or -] 2.8 minutes, P =.003) compared with xylazine (3.8 [+ or -] 0.4 and 31.5 [+ or -] 3.6 minutes, respectively).

In the midazolam group, the onset time was significantly (P <.001) shorter at 0.8 mg/kg (1.9 [+ or -] 0.2 minutes) compared with 0.4 mg/kg (3.9 [+ or -] 0.2 minutes). Maximum duration effect was significantly (P <.001) longer at 0.8 mg/kg (38.0 [+ or -] 2.8 minutes) compared with 0.4 mg/kg (17.7 [+ or -] 4.1 minutes). In the xylazine group, the onset time was significantly (P =. 004) shorter at 4 mg/kg (3.8 [+ or -] 0.4 minutes) compared with 2 mg/kg (4.9 [+ or -] 0.4 minutes), and the maximum duration effect was significantly (P <.001) longer at 4 mg/kg (31.5 [+ or -] 3.6 minutes) compared with 2 mg/kg (9.2 [+ or -] 2.5 minutes).

Intranasal administration of diazepam and midazolam at 1.6 mg/kg (8-9.5 mL/naris) was not applicable because the volume of the drugs was not tolerated by the ostriches and head shaking caused the drugs to drip out of the nares. No sedative effect with xylazine was observed at 0.5 mg/kg.


The results of the present study showed that intranasal administration can be an effective route for delivery of sedatives in juvenile ostriches. The use of different intranasal doses of diazepam, midazolam, and xylazine provides a dose-dependent sedation that may be applicable clinically for standing chemical restraint or preanesthesia.

In this study, the precise site of drug absorption after intranasal administration was not determined; hence, the short onset time may have resulted from absorption of the drugs across the nasal, oral, or pharyngeal mucosa, or across all 3. (9,27) However, in our pilot study, oral administration of midazolam at 0.8 mg/kg and xylazine at 4 mg/kg in 4 ostriches did not produce desirable sedation during the 4 hours of observation, which was in agreement with the results from the use of midazolam in ring-necked parakeets and canaries. (9,27) The short onset time may have resulted from rapid transportation of the drugs into the bloodstream and brain. (20) The nasal mucosa represents a highly permeable route of drug absorption, and many drugs are absorbed systemically at this site. (27) The highly vascularized nasal mucosa and the olfactory tissue that is in direct contact with the central nervous system allow intranasal administration to be a well-established alternative method for drug delivery that is often as fast in onset as intravenous medication in a variety of acute pediatric medical conditions. (20) Also, the terminal half-life of intranasal midazolam is reported to be comparable to that of the intravenous route (25); sedation via the intranasal route may be the result of high bioavailability of the administrated drugs by avoiding the first-pass drug metabolism via the liver. (20,32)

In the present study, proper standing chemical restraint was achieved with diazepam, midazolam, and xylazine at 0.8, 0.4, and 2 mg/kg, respectively. Also, midazolam at 0.8 mg/kg and xylazine at 4 mg/kg resulted in deep sedation with sternal recumbency. Because of the lower dose, the shorter onset time, and the longer maximum effect, midazolam could be suggested as a preferred sedative at 0.4 mg/kg for standing chemical restraint and at 0.8 mg/kg for recumbency.

In this study, shorter onset time was achieved with intranasal midazolam compared with diazepam or xylazine, which is similar to the results of using these medications intranasally in budgerigars and zebra finches. (7,29) Also, shorter onset time was reported by intranasal detomidine and midazolam compared with xylazine in ring-necked parakeets. (9) However, in canaries, a comparable onset time with intranasal diazepam, midazolam, detomidine, and xylazine was reported. (27) The shorter onset time of midazolam compared with diazepam and xylazine may result from rapid entrance of midazolam into the central nervous system, twice the affinity of midazolam for the benzodiazepine receptors versus diazepam, and highly lipophilic characteristics of midazolam at physiologic pH versus xylazine. (10) Furthermore, onset of sedation and muscle relaxation is rapid after intravenous or intramuscular administration of midazolam in most species. (10) Despite significant differences in onset time of sedation, the differences observed (1-2 minutes) are not considered to be of clinical significance.

In the present study, the longer duration of sedation with midazolam compared with diazepam is in agreement with the results of Olivos et al, (33) who reported significantly longer sedation with midazolam compared with diazepam after intravenous or intramuscular administration in domestic ostriches. However, longer duration of sedation after intranasal diazepam compared with midazolam is reported in budgerigars, zebra finches, and canaries. (7,27,29)

The duration of sedation was longer with midazolam compared with xylazine in this study, which is contrary to the results of studies comparing administration of these drugs in budgerigars, zebra finches, canaries, and ring-necked parakeets. (7,9,27,29) The longer duration of sedation with xylazine compared with diazepam in the present study is in agreement with the results of the use of these drugs in budgerigars, zebra finches, and canaries. (7,27,29) However, prolonged recovery time and inadequate sedation have been reported with xylazine, which are important undesired consequences in these birds. (7,9,27,29) These differences in duration of sedation may be attributed to the pharmacokinetic profile of administered drugs, which can differ from one avian species to another because of the relationship between biotransformation and excretion that is determined by metabolic factors and heredity. (34) Also, the activity of the drug-metabolizing enzyme system in ostriches is quantitatively different from that of chickens. (35) Although clinically important adverse effects associated with xylazine use have been reported in many species, (2) xylazine has been used safely as a preanesthetic in ostriches. (5,36)

In this study, the higher the dose of midazolam or xylazine used, the more accelerated the onset time, the longer the duration of sedation, and the longer the maximum duration effect achieved. This finding could be the result of the influence of the instilled dose to the relative distribution of substances onto nasal mucosa. (37) Minimizing drug volume while maximizing drug concentration could be a successful delivery technique to optimize medication bioavailability and effectiveness via the intranasal route. However, there is a limit to the volume of drug that can be instilled intranasally that allows an adequate dose of medication to be administered. (20) In the present study, only commercially available parenteral drugs were used. To prevent nasal mucosal surface saturation, the volume administered was minimized by using both nostrils, to double the absorptive surface. Also, the drugs were administered for a duration of 2 minutes to enhance mucosal contact time and, subsequently, absorption. Although the surface area and volume of the nasal cavity were not investigated in this study, the volume of 4-6 mL per nostril administered IN over 2 minutes was well tolerated by the ostriches.

In conclusion, the intranasal route of administration appears to be an acceptable noninvasive alternative to the intravenous or intramuscular method for delivery of sedatives in juvenile ostriches. This method requires minimal training and seems to be safe, fast, and effective. Intranasal midazolam at 0.4 mg/kg or xylazine at 2 mg/kg could provide adequate sedation needed for standing chemical restraint in juvenile ostriches. Midazolam at 0.8 mg/kg administered intranasally could be suggested as a proper sedative preanesthetic agent with sternal recumbency in juvenile ostriches.

Acknowledgments: We thank Dr Toktam Heidari for technical assistance. Special thanks to Mr Amir Abedian, ostrich farm owner, for his kindly cooperation.


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Mostafa Araghi, DVM, Saeed Azizi, DVM, DVSc, Nasser Vesal, DVM, MVetSc, and Bahram Dalir-Naghade, DVM, DVSc

From the Department of Surgery and Diagnostic Imaging (Araghi, Azizi) and the Department of Internal Medicine and Clinical Pathology (Dalir-Naghade), Faculty of Veterinary Medicine, Urmia University, Urmia, Iran, and the Department of Veterinary Clinical Sciences, School of Veterinary Medicine, Shiraz University, Shiraz, Iran (Vesal).
Table 1. Onset time and duration of sedation (mean [+ or -]
standard deviation) with intranasal diazepam,
midazolam, and xylazine for mild sedation in juvenile
ostriches (n = 6).

            Dose,          Onset               Duration of
  Drug      mg/kg        time, min            sedation, min

Diazepam    0.2     6.1 [+ or -] 0.6       6.4 [+ or -] 0.6
            0.4     4.9 [+ or -] 0.4 *     9.0 [+ or -] 0.9
Midazolam   0.2     4.8 [+ or -] 0.8 **   12.7 [+ or -] 3.7 ***
Xylazine    1       5.9 [+ or -] 0.4       8.3 [+ or -] 2.2

* Significantly different from diazepam at 0.2 mg/kg (P < .05).

** Significantly different from diazepam (0.2 mg/kg) and xylazine
(P < .05).

*** Significantly different from other doses of diazepam and
xylazine (P < .05).

Table 2. Onset time, maximum effect, and duration of sedation
(mean [+ or -] standard deviation) with intranasal diazepam,
midazolam, and xylazine for moderate sedation in juvenile
ostriches (n = 6).

  Drug      Dose, mg/kg    Onset time, min

Diazepam        0.8       4.3 [+ or -] 0.4
Midazolam       0.4       3.9 [+ or -] 0.2 *
Xylazine         2        4.9 [+ or -] 0.4

                Duration of           Maximum effect
  Drug         sedation, min         of sedation, min

Diazepam    20.8 [+ or -] 1.9       7.0 [+ or -] 1.4
Midazolam   31.8 [+ or -] 4.0 **   17.7 [+ or -] 4.1 **
Xylazine    20.2 [+ or -] 2.8       9.2 [+ or -] 2.5

* Significantly different from xylazine {P < .05).

** Significantly different from diazepam and xylazine (P < .05).

Table 3. Onset time, maximum effect, duration of sedation, and
sternal recumbency (mean [+ or -] standard deviation) after use
of intranasal midazolam and xylazine for deep sedation in
juvenile ostriches (n = 6).

                                                   Duration of
  Drug      Dose, mg/kg    Onset time, min        sedation, min

Midazolam       0.8       1.9 [+ or -] 0.2 *   54.5 [+ or -] 4.8 *
Xylazine         4        3.8 [+ or -] 0.4     45.5 [+ or -] 2.9

              Maximum effect      Duration of sternal
  Drug       of sedation, min       recumbency, min

Midazolam   38.0 [+ or -] 2.8 *   21.7 [+ or -] 4.9 *
Xylazine    31.5 [+ or -] 3.6     13.5 [+ or -] 2.6

* Significantly different from xylazine (P < .05).
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Title Annotation:Original Study
Author:Araghi, Mostafa; Azizi, Saeed; Vesal, Nasser; Dalir-Naghade, Bahram
Publication:Journal of Avian Medicine and Surgery
Date:Sep 1, 2016
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