This Article Abstract Résumé de cet Article Full Text (PDF) Submit a scholarly reply Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Mitchell, L. W. Articles by Leighton, B. L. Search for Related Content PubMed PubMed Citation Articles by Mitchell, L. W. Articles by Leighton, B. L.

Warmed diluent speeds dantrolene reconstitution

[Le diluant réchauffé accélère la reconstitution du dantrolène]

Laura W. Mitchell, PhD^* and Barbara L. Leighton, MD

¹ From the Department of Chemistry, *St. Joseph’s University, Philadelphia, Pennsylvania and
the Department of Anesthesiology, MCP Hahnemann University, Philadelphia, Pennsylvania, USA.

Address correspondence to: Dr. Barbara L. Leighton, Department of Anesthesiology, 660 S. Euclid Ave, Campus Box 8054, St Louis, Missouri 63110, USA. Phone: 314-362-2356; Fax: 314-362-1185; E-mail: bleigh{at}alumni.princeton.edu

	Abstract

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
Purpose: To define more completely the aqueous solubility of dantrolene in order to devise faster reconstitution techniques for use during malignant hyperthermia episodes.

Methods: To determine Beer’s law compliance and the extinction coefficient, we measured the spectrophotometric absorbance at 385 nm of known dantrolene solutions. We added small aliquots of sterile water USP (pH 5.3; 15–40°C) or buffered water (20°C; pH 6.8–9.1) to dantrolene, mechanically agitated and filtered the solutions, and spectrophotometrically determined concentration. To simulate clinical reconstitution conditions, we added sterile water, 60 mL, at temperatures between 15 and 40°C to dantrolene vials and measured the manual shaking time needed to create a) a suspension of small particles, and b) a clear solution.

Results: A plot of UV-vis absorbance at 385 nm vs dantrolene concentration was linear and went through the origin; the extinction coefficient is 16.1 mM^-1. At 20°C, dantrolene is nearly insoluble below pH 8.8. Dantrolene is 2.8 times more soluble in 0.1 M THAM (tris-(hydroxymethyl)aminomethane) than in sterile water at pH 9.1. Dantrolene is 6.7 times more soluble in 40°C than in 20°C water at pH 9.5 (the pH of reconstituted dantrolene). Under clinical conditions, water temperature altered the time to create a clear solution but not a suspension (60 sec).

Conclusion: Diluting dantrolene with 40°C water rather than operating-room temperature water (20°C or below) would speed dantrolene reconstitution.

	Introduction

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
RECONSTITUTING dantrolene fast enough to reverse malignant hyperthermia (MH) in an adult is a difficult task. Gronert et al. advise that this job usually requires the full-time efforts of three or four people.¹ The only iv dantrolene formulation available in North America (Dantrium®, Procter & Gamble Pharmaceuticals, Cincinnati, OH, USA; each vial contains dantrolene 20 mg, mannitol 3 g, and sodium hydroxide to ensure that the reconstituted solution has a pH of 9.5), is poorly water soluble. However, water is the recommended dantrolene diluent.

Heat speeds dantrolene dissolution. If dantrolene does not dissolve readily, textbooks recommend heating the dantrolene vial either by immersing the vial in hot water or gently autoclaving the vial.¹ This could be hazardous, for one could easily apply excessive heat (particularly with autoclaving) and produce a dantrolene solution hot enough to cause burns.^2,3 Also, both suggested heating techniques take time. An alternative way to facilitate dantrolene dissolution might be to preheat the sterile water diluent to an effective but thermally safe (i.e.,40°C) temperature. However, there is insufficient information on the temperature dependence of dantrolene solubility to determine whether such preheating might be clinically useful. In addition, the pH adjustments recommended by some authors to assist dantrolene preparation have little support.^4,5 Therefore, we systematically studied the solubility of dantrolene as a function of temperature and pH.

	Methods

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
Materials
The dantrolene used to confirm compliance with Beer’s law was obtained from Sigma-Aldrich Chemical Co., St. Louis, MO, USA. We obtained the dantrolene used in temperature, pH and THAM dissolution experiments (Dantrium® Intravenous, Procter & Gamble Pharmaceuticals, Cincinnati, OH, USA) at the time of clinical expiration (April 30, 1999) from City Avenue Hospital, Philadelphia, PA, USA and performed these experiments May 11-12, 1999. We obtained phosphate buffer (sodium potassium phosphate), concentrated HCl, THAM (tris-(hydroxymethyl)aminomethane), and an Accumet pH meter from Fisher (ACS grade; Fisher Scientific, Malvern, PA, USA). THAM and phosphate buffer solutions were diluted with house distilled water that was additionally purified by passage through a Barnstad water polisher. All other water used was sterile water, USP. Cloudy suspensions and solutions containing particles were filtered through 0.2 µM syringe filters with cellulose acetate membranes (Fisher Scientific, Malvern, PA, USA).

Measurement of ultraviolet-visible spectra of dantrolene solutions and calculation of dantrolene concentration
We obtained ultraviolet-visible (UV-vis) spectra in quartz cuvettes using a Hitachi U2001 spectrophotometer (Pleasanton, CA, USA) scanning from 190 nm to 1100 nm at a scan speed of 800 nm•min^-1 using the appropriate buffer as a reference. To determine compliance with Beer’s law and dantrolene’s extinction coefficient, we measured the maximal absorbance at 385 nm of solutions of pure dantrolene (12–120 µM) dissolved in water. We calculated the concentrations of the solutions in parts 1 and 2 using absorbance data at 385 nm.

	Part 1. Dantrolene solubility with varying pH

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
We determined the effect of solute pH on dantrolene solubility by adding 0.5–1.0 mL aliquots (up to a total volume of 10 mL) of 0.1 M THAM (pH 7.1–9.1), 0.1 M phosphate (pH 6.8 and 7.2), or sterile water, USP (pH 5.3) to 0.33–0.35 g Dantrium®. Solutions were mechanically agitated to speed dissolution. Aliquots were added until a clear solution formed or a maximum volume of 10 mL was reached. Solutions that were cloudy with 10 mL diluent were filtered. Solution pH was redetermined after reconstitution. We compared the optical densities for dantrolene dissolved in sterile water vs 0.1 M THAM at pH 9.1.

	Part 2. Dantrolene solubility with varying water temperature

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
In order to assess whether dantrolene stability depended on the temperature of the water used for reconstitution, we measured absorbance at 385 nm immediately and 24 hr after reconstitution for all solutions mixed in part 2. Solutions were stored at 20°C after reconstitution. The diluent was sterile water, USP (pH 5.3).

A. LABORATORY CONDITIONS
We determined the minimum fluid volume in which the contents of a Dantrium® vial dissolved when the initial temperature of the sterile water diluent was 20, 25, 30, 37, or 40°C. We performed one experiment at each diluent temperature. In each experiment, we incrementally added 2–5 mL aliquots of sterile water, USP, to a previously unopened Dantrium® vial, thoroughly shook the vial manually and with a mechanical agitator for at least 30 sec after each aliquot, and then visually inspected the vial to determine whether the vial contents had dissolved. If the vial contents did not fully dissolve in 60 mL of water, the resulting solution was filtered to remove insoluble material.

B. SIMULATED CLINICAL CONDITIONS
Sterile water, USP, 60 mL, was added at 15, 20, 25, 30, 37, or 40°C to Dantrium® vials and the duration of manual shaking needed to dissolve the contents was determined. We did not use a mechanical agitator as one is usually not available in the operating room. The solutions were observed continuously to determine: a) when a clear solution formed; and b) when a suspension with small but not large particles formed. Manual shaking was discontinued for five seconds every 30 sec to facilitate visual inspection. We decided that dissolution times longer than three minutes were clinically impractical and did not shake vials longer than 180 sec. Vials in which a clear solution had not formed by 180 sec were observed again after 60 min storage at 20°C.

	Results

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
A plot of UV-vis absorbance at 385 nm vs dantrolene concentration was linear and went through the origin, indicating compliance with Beer’s law. We obtained an extinction coefficient of 16.1 mM^-1.

Part 1. Dantrolene solubility with varying pH
Dantrolene is nearly insoluble at pH values below 8.8 at 20°C (Figure 1). At 20°C and pH 9.1, dantrolene is 2.8 times more soluble in 0.1 M THAM than in sterile water (2.52 vs 0.88 mM).

View larger version (10K): [in this window] [in a new window]   FIGURE 1 Dantrolene solubility vs pH. Maximal dantrolene concentrations in 0.1 M phosphate (pH 6.8 and 7.2) and 0.1 M THAM (pH 7.1–9.1) were determined spectrophotometrically. When dantrolene is reconstituted as recommended with 60 mL of water, the dantrolene concentration is about 1 mM.

A. LABORATORY CONDITIONS
Dantrolene optical density at 385 nm increased linearly with temperature between 20°C and 40°C at pH 9.5 (Figure 2). Dantrolene is 6.7 times more soluble in 40°C water than in 20°C water.

View larger version (12K): [in this window] [in a new window]   FIGURE 2 Dantrolene solubility vs temperature. Maximal dantrolene concentrations in sterile water, USP (20–40°C) were determined spectrophotometrically. The y-axis scale of this figure is the same as that in Figure 1. When dantrolene is reconstituted as recommended with 60 mL of water, the dantrolene concentration is about 1 mM.

	Discussion

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

During simulated clinical conditions, a clear solution of dantrolene never formed after the addition of 60 mL sterile water (the recommended volume) at 15°C or 20°C. It is not known if injecting a dantrolene suspension rather than a solution would present patient safety issues. In general, however, drugs administered in aqueous solution are available at cellular sites of action more quickly than drugs administered as fine particles in suspension.⁷

Warm sterile water for injection is rarely available in the operating room. Hospitals frequently store the sterile water for dantrolene reconstitution in the operating suite, which has an average air temperature of 18–21°C.⁶ Procter & Gamble recommends that dantrolene be reconstituted with water at "room temperature"; one doubts that the manufacturer fully appreciates the frosty ambiance of most North American operating rooms. An iv fluid warming device could be used to warm the sterile water for dantrolene dissolution to 40°C. Alternatively, many operating rooms maintain 37°C warming cabinets for the storage of saline and sterile water irrigating solutions and cotton blankets. Sterile water for injection, USP could be stored in one of these warming cabinets.

Heat also increases the aqueous solubility of mannitol, the other active ingredient in Dantrium®. A vial of Dantrium® contains 3,000 mg mannitol and 20 mg dantrolene; thus, the ingredients are 99% mannitol by weight. According to the mannitol package insert, mannitol crystals precipitate if the solution is chilled.^A Heating is the recommended way to redissolve crystal-containing mannitol containers.^A

Paradoxically, the time saved by using 40°C water to reconstitute dantrolene could decrease heat gain in MH patients. During fulminant MH, core temperature can increase as much as 1°C every five minutes.¹ In a 70-kg adult, this translates to about 14 kcal•min^-1 heat production.^B The 420-mL of water needed to mix a 2-mg•kg^-1 dose for a 70-kg patient contains 8.4 kcal more heat at 40°C than at 20°C. Using 40°C water would be thermally neutral if one saved 36 sec in dantrolene preparation time (5.1 sec/vial) and would result in net heat loss if more time were saved.

At 20°C, dantrolene is nearly insoluble at pH values below 8.8. pH adjustments of the dantrolene diluent are unlikely to improve the drug preparation time. Dantrium is packaged to have pH 9.5 after reconstitution with sterile water USP, and higher pH values might cause tissue damage. Although dantrolene is more soluble in THAM than in sterile water, the effect is small and THAM is quite expensive. THAM does not appear to be a cost-effective adjuvant in dantrolene preparation. In addition, the effects of THAM buffer on the stability and potency of dantrolene would need to be determined before the use of a THAM buffer diluent could be recommended.

Prompt dantrolene injection is the cornerstone of effective MH therapy. Precious time can be lost if one is not prepared for the practical problem of dantrolene reconstitution. We recommend that the sterile water for dantrolene reconstitution either be stored in a 40°C operating room warming cabinet or that equipment be available for warming the water rapidly to 40°C, since dantrolene is far more soluble in warm water than cold.

	Footnotes

 
Received from and supported only by the Department of Chemistry, St. Joseph’s University, Philadelphia, PA and the Departments of Anesthesiology, MCP Hahnemann University, Philadelphia PA and Weill Medical College of Cornell University, New York, NY, USA.

Presented in part at the 1999 annual meeting of the American Society of Anesthesiologists.

^A Mannitol iv package insert, Abbott Laboratories, North Chicago, IL, USA, ©Abbott, 1994, revised 2/1998.

^B For this rough approximation, we assumed that total body water is 100% rather than 67% of body weight. One kilocalorie increases the temperature of 1 kg by 1°C. If body temperature can increase 0.2°C per minute during fulminent MH (1°C every five minutes), then a 70-kg MH patient can produce 14 kcal of heat per minute.

Revision received August 26, 2002. Accepted for publication May 23, 2002.

	References

TOP Abstract Introduction Methods Part 1. Dantrolene solubility... Part 2. Dantrolene solubility... Results Discussion References

 
1 Gronert GA, Antognini JF, Pessah IN. Malignant hyperthermia. In: Miller RD (Ed.). Anesthesia, 5th ed. New York: Churchill Livingstone Inc., 2000: 1033–52.

2 Leaman PL, Martyak GG. Microwave warming of resuscitation fluids. Ann Emerg Med 1985; 14: 876–9.[Medline]

3 Sieunarine K, White GH. Full-thickness burn and venous thrombosis following intravenous infusion of microwave-heated crystalloid fluids. Burns 1996; 22: 568–9.[Medline]

4 Drechsler H. Zur. [On the treatment of malignant hyperthermia. A simple method of preparing a dantrolene sodium solution (author’s transl)] German. Anaesthesist 1981; 30: 257–8.[Medline]

5 Jansen ACA, Hilbers HW, Ni XR, van Helden SP, Janssen LHM. Some physical-chemical properties of dantrolene and two of its analogues. Int J Pharm 1991; 75: 193–9.

6 Blitt CD. Monitoring the anesthetized patient. In: Barash PG, Cullen BF, Stoelting RK (Eds.). Clinical Anesthesia, 3rd ed. Philadelphia: Lippencott-Raven Publishers, 1997: 563–85.

7 Benet LZ, Kroetz DL, Sheiner LB. Pharmacokinetics. The dynamics of drug absorption, distribution, and elimination. In: Hardman JG, Limbird LE (Eds.) Goodman & Gilman’s The Pharmacological Basis of Therapeutics, 9^th ed. New York: McGraw-Hill, 1996: 3–27.

This article has been cited by other articles:

				D. Chartrand Intervention rapide lors d'une crise d'hyperthermie maligne/Rapid intervention for an episode of malignant hyperthermia Can J Anesth, February 1, 2003; 50(2): 104 - 107. [Full Text] [PDF]

This Article Abstract Résumé de cet Article Full Text (PDF) Submit a scholarly reply Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Mitchell, L. W. Articles by Leighton, B. L. Search for Related Content PubMed PubMed Citation Articles by Mitchell, L. W. Articles by Leighton, B. L.