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Original Papers

In vitro changes in the transparency and pH of cerebrospinal fluid caused by adding midazolam

Nishiyama, T.; Sugai, N.; Hanaoka, K.

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European Journal of Anaesthesiology: January 1998 - Volume 15 - Issue 1 - p 27-31

Abstract

Introduction

Midazolam causes analgesia at the spinal level, possibly via γ-aminobutyric acid (GABA) receptors [1]. Midazolam does not cause histopathological changes when given intrathecally in rats [2], and has been used both intrathecally [3] and epidurally [4] for pain relief in humans without adverse effects.

This does not mean that perispinal midazolam is harmless. Malinovsky and colleagues [5] showed microscopic changes in the blood-brain barrier of rabbits. Local anaesthetics, especially chloroprocaine, can be neurotoxic, possibly because of their low pH [6], and midazolam solution is acidified.

Clouding of a solution indicates the formation of particles which might be neurotoxic, or some other change in its characteristics. Assuming that clouding could indicate neurotoxicity, we examined the effect on pH and transparency of adding midazolam to human cerebrospinal fluid.

Materials and methods

The protocol was approved by the ethics committee of The University of Tokyo, Tokyo, Japan, and informed consent was obtained from all patients.

The transparency and pH of the test drugs were checked by taking six samples from each drug preparation: up to 10 mg midazolam (Dormicum™, Yamanouchi Pharmaceutical Co. Ltd, Tokyo, Japan; 10 mg 2 mL−1) was added to 10 mL of saline solution (Otsuka Pharmaceutical Co. Ltd, Tokyo, Japan) and 6 mL of 0.25% bupivacaine (Marcain™, Astra Japan Co. Ltd, Osaka, Japan). The tested combinations of the agents, and their doses and concentrations, were chosen because these are used in epidural injection for post-operative pain relief [7,8] in human subjects. The preparation of bupivacaine contains preservatives, but is the only preparation available in Japan.

Transparency was checked by the naked eye. The solutions were placed in glass test-tubes and were checked against a black background. Without knowing the contents of the test-tubes, a single observer with good near vision compared the test solutions with the original drugs and with the cerebrospinal fluid [9]. The pH was measured by pH Strip™ (Merck Co. Ltd, Darmstadt, Germany), which has a detectable range of 0-14 and an interval of 0.2-0.3.

A sample of 1.5 mL of cerebrospinal fluid was drawn from each patient. This volume was chosen because it is the same as the normal intrathecal dose of local anaesthetic. The sample was poured into a clear glass test-tube. Four groups of tests were done. In the first group(group M), midazolam was added in increments of 0.1 mg to a total of 10 mg; in the second group (MS), 5 mg of midazolam in 10 mL of saline was added in increments of 0.02 mL to a total of 2 mL; in the third group (B), 0.25% of bupivacaine was added in increments of 0.04 mL to a total of 3 mL; and in the fourth group (MB), 5 mg of midazolam in 6 mL of 0.25% bupivacaine was added in increments of 0.04 mL to a total of 3 mL. The transparency and pH were checked after each increment in all groups.

Statistical analysis of the changes in pH was by analysis of variance for repeated measures followed by Fisher's protected least significant difference; P<0.05 was considered statistically significant.

Results

We studied samples from 24 patients (10 male and 14 female, aged between 30 and 65 years, weighing between 40 and 72 kg) who were scheduled for elective operations suitable for spinal anaesthesia. There were six samples in each of the tested groups.

The addition of midazolam decreased the pH of the saline and 0.25% bupivacaine solutions (Fig. 1). No clouding was observed.

Fig. 1
Fig. 1:
pH changes when midazolam was added to (a) 10 mL of saline or (b) 6 mL of 0.25% bupivacaine:(○) pH of midazolam; (*)P<0.05 vs. pH of solution without midazolam; bars indicate SD.

Adding 3 mg of midazolam to 1.5 mL of cerebrospinal fluid (group M) decreased the pH to below 7.0, but even after adding 10 mg of midazolam, the pH was still above 6.0 (Fig. 2). Clouding of the cerebrospinal fluid was first seen in some of the samples at 0.7 mg midazolam and all the samples were clouded after the addition of more than 1 mg midazolam (Fig. 2).

Fig. 2
Fig. 2:
pH changes and clouding when midazolam added to 1.5 mL of cerebrospinal fluid (CSF): (○) pH of midazolam; (*)P<0.05 vs. pH of CSF without midazolam; bars indicate SD; (in the centre) number of samples clouded by midazolam.

Adding the solution of midazolam (group MS) did not lower the pH to below 7.0 (Fig. 3) and there was no clouding.

Fig. 3
Fig. 3:
pH changes and clouding when 5 mg of midazolam in 10 mL of saline were added to 1.5 mL of cerebrospinal fluid (CSF): (○) pH of midazolam solution; (*)P<0.05 vs. the pH of CSF without added solution; bars indicate SD; there was no clouding.

Adding 0.25% of bupivacaine(group B) lowered the pH to below 7.0 after 1.9 mL had been added. Clouding was first seen in some of the samples with 1.1 mL of bupivacaine, and all the samples were clouded at the 1.4 mL level (Fig. 4). Adding the mixture of midazolam and bupivacaine (group MB) lowered the pH to below 7.0 after 1.3 mL; clouding first occurred at 0.6 mL and all the samples were clouded after the addition of 1.1 mL(Fig. 4).

Fig. 4
Fig. 4:
pH changes and clouding when 0.25% of bupivacaine (○) or 5 mg of midazolam in 6 mL of 0.25% bupivacaine (•) were added to 1.5 mL of cerebrospinal fluid (CSF): (open triangle) pH of bupivacaine; (closed triangle) pH of mixture; (*)P<0.05 vs. the pH without addition; bars indicate SD; (in the centre) number of samples clouded by the additions.

Discussion

Solutions with a low pH are thought to be neurotoxic [6]. Bupivacaine has a pH of 4 and is not neurotoxic intrathecally [10], but the reported pH was of the bupivacaine solution, not the pH when added to cerebrospinal fluid.

We have shown in this study that the pH of a sample of 1.5 mL of cerebrospinal fluid was decreased below 7.0 by more than 3 mg midazolam, more than 1.9 mL 0.25% bupivacaine or more than 1.3 mL of midazolam in 0.25% bupivacaine solution. Clouding was seen after adding more than 0.7 mg of midazolam, more than 1.1 mL of bupivacaine or more than 0.6 mL of the combined solution. Midazolam in saline had little effect on pH and there was no clouding.

If drugs injected into the cerebrospinal fluid dilute rapidly, we can extrapolate to what would happen to the whole cerebrospinal fluid volume, which is about 70-80 mL based on the standard textbooks published in Japan. This gives a factor of 46.7 (70 divided by 1.5), and means that more than 32.6 mg of midazolam, more than 51.3 mL of 0.25% bupivacaine or more than 28 mL of the combined solution would be needed to cause clouding of the cerebrospinal fluid, and that more than 140 mg of midazolam, more than 88.7 mL of 0.25% bupivacaine or more than 60.7 mL of the combined solution would be needed to lower the pH below 7.0. Even 93 mL of midazolam in saline would not have any important effect.

Intrathecally, 2 mg of midazolam is effective in relieving chronic low back pain without any side effects [11]. The effective dose of epidural midazolam for post-operative analgesia is 0.05 mg kg−1 in 10 mL of saline [7] or in 6 mL of 0.25% bupivacaine [8]. A dose of 1.25-3.75 mg in 3 mL of 5% dextrose is effective for the relief of spasticity[12]. Clouding and lowering pH to below 7.0 needed doses far in excess of these. We are presuming that neurotoxicity will not occur unless the pH is less than 7, but we have also ignored the secretion of fresh cerebrospinal fluid (0.35 mL min−1[13]), which is an extra diluent of any injected drug.

In experimental studies, injection of midazolam intrathecally in rats [2] and epidurally in rabbits [14], midazolam caused no histopathological changes in the spinal cord. In only one study (Malinovsky et al.) [5], necrosis, haemorrhage and other histopathological changes occurred in three out of the nine spinal cords of rabbits that had received single-shot intrathecal injections of midazolam [1-4].

There have been no studies of the effects of the speed of injection of midazolam, either epidurally or intrathecally; at the moment of injection, the drug will be in its highest concentration and the possibility of neurotoxicity may be greater. However, from our study, and presuming that clouding and changes in pH are indications of possible neurotoxicity, we suggest that perispinally injected midazolam is unlikely to be neurotoxic.

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Keywords:

cerebrospinal fluid; in vitro changes; midazolam

© 1998 European Academy of Anaesthesiology