Collaborative study for the establishment of the 3rd international standard for amphotericin B.

An international collaborative study was organised to establish the 3rd World Health Organization (WHO) International Standard (IS) for amphotericin B. Sixteen laboratories from different countries participated. Potencies of the candidate material were estimated by microbiological assays with sensitive micro-organisms. To ensure continuity between consecutive batches, the 2nd IS for amphotericin B was used as a reference. Based on the results of the study, the 3rd IS for amphotericin B was adopted at the meeting of the WHO Expert Committee for Biological Standardization (ECBS) in 2019 with an assigned potency of 953 International Units (IU) per mg. The 3rd IS for amphotericin B is available from the European Directorate for the Quality of Medicines & HealthCare (EDQM).

Influence of 5% dextrose volume on amphotericin B deoxycholate preparation.

OBJECTIVES: Preparation of amphotericin B deoxycholate (AmB-d) in different volumes of 5% dextrose (D5W) was studied to investigate a interesting phenomenon that AmB-d was easy to bring pipe blockage when diluted in 500 ml but not in 50 ml. METHODS: AmB-d (25 mg/vial) in 50 ml, 250 ml or 500 ml D5W was prepared. Fluids were collected before and after infusion, then were assayed by validated high-performance liquid chromatography (HPLC) method. Light obscuration assay was used to detect the particles in transfusions. KEY FINDINGS: pH values of different volumes of D5W were all about 3.7, which was lower than the requirement of AmB-d package insert (pH > 4.2). The number of insoluble particles >10 µm/25 µm in 25 mg/500 ml infusions exceeded China Pharmacopoeia limit. Filters in 25 mg/500 ml infusion set were full of AmB-d after dripping slowly for 6 h, and 331.3 ml solution was left in the bottles and only 11.3% of AmB-d could flow out. Whereas the AmB-d infusion consists of 25 mg/50 ml, 25 mg/250 ml and 50 mg/500 ml could meet with China Pharmacopoeia standards, and they flowed out easily and completely. CONCLUSIONS: In practice, 25 mg/250 ml and 50 mg/500 ml would be more suitable for clinical use, rather than 25 mg/500 ml. We provided a convenient method for AmB-d preparation.

Collaborative study for the establishment of the second international standard for amphotericin B.

An international collaborative study was organised by the European Directorate for the Quality of Medicines & HealthCare (EDQM) to establish the WHO second International Standard (IS) for amphotericin B as the stocks of the 1st IS, established in 1963, were close to depletion. The candidate material did not show any sign of potency loss when kept at elevated temperatures of + 4 degrees C, + 20 degrees C, + 37 degrees C and + 45 degrees C for 3 months. Ten laboratories from 8 countries participated in the collaborative study. Potencies of the candidate material were estimated by microbiological assays with sensitive micro-organisms. To ensure continuity between consecutive batches, the 1st IS for amphotericin B was used as the standard. Based on the results of the study, the 2nd IS for amphotericin B was adopted at the meeting of the WHO Expert Committee on Biological Standardisation (ECBS) in 2007 with an assigned potency of 944 International Units per mg (IU/mg). The 2nd IS for Amphotericin B is available from the EDQM.

[Clinical audit on the use of expensive systemic antifungals in the Besançon University Hospital]. / Audit clinique des prescriptions d'antifongiques systémiques couteux au centre hospitalier universitaire de Besançon.

UNLABELLED: The continuous improvement policy for healthcare quality requires practice evaluation. The principle of a clinical audit is to compare practice to guidelines. Prescription guidelines on antifungal agent use has been available in our hospital since 2003. It was updated in 2005 and 2006. OBJECTIVE: The aim of this study was to assess compliance to guidelines, with an audit of prescriptions: amphotericin B lipid formulation, voriconazole and caspofungin, expensive antifungals concerned by the budget allowance correlated to activity, subject to supplementary reimbursement to the coded Homogeneous Group of Diseases. METHOD: The assessment criteria were: relevance of the indication, absence of a better alternative, complying to recommended dosage, loading dose and timing. This retrospective study dealt with all prescriptions of all departments, from January to May 2007. RESULTS: Hundred and eighteen prescriptions were retrospectively analyzed for 81 patients. The rate of overall conformity was 54%. Antifungal therapy was justified for 113 prescriptions (96%). In 30% of the cases, a more efficient alternative was advised, cheaper or less toxic. The dosage and the charge dosing were right in 92% and 80% of the cases respectively. CONCLUSION: This audit allowed assessing good-use of antifungals. We showed an over-prescription of caspofungin and sometimes insufficient regimen of voriconazole dosages for children. Reporting these audit results and development of new international guidelines stress the need to update local recommendations regularly.

Correlation between CLSI, EUCAST and Etest methodologies for amphotericin B and fluconazole antifungal susceptibility testing of Candida spp. clinical isolates.

This study analyzed the correlation between the results obtained through two microdilution methods: Clinical and Laboratory Standard Institute (CLSI) (M27-A2) and European Committee on Antibiotic Susceptibility Testing (EUCAST) (document E. Dis. 7.1) and an agar base method Etest for determining minimmun inhibitory concentration (MIC) for amphotericin B and fluconazole against 30 clinical isolates of Candida spp. The agreement between Etest, CLSI and EUCAST MICs within +/- 2 log2 dilutions was higher for amphotericin B than for fluconazole However, Pearson correlation demonstrated a greater agreement for fluconazole. The categorical agreement between MICs provided by the Etest/ CLSI and Etest/EUCAST methodologies was high for both amphotericin B (100%) and fluconazole (> or = 96.66%). This study demonstrated the adequacy of Etest method using Mueller Hinton agar to evaluate amphotericin B and fluconazole susceptibility of clinical isolates of Candida spp.

Particulate contamination of lyophilized amphotericin B preparation during reconstitution process.

OBJECTIVE: To investigate the effect of the reconstitution methods for the commercial amphotericin B preparation with respect to particulate contamination. METHODS: The particle counts in amphotericin B solutions reconstituted according to three different methods and amphotericin B fluids made with intravenous fluids after reconstitution were performed using a light extinction method. The particle contaminants were identified with X-ray emission spectrometry attached to a scanning electron microscope. RESULTS: Amphotericin B in a vial induced particle contamination during the reconstitution process, and the contamination was especially marked by shaking vigorously after injecting water into the vial. From the X-ray analysis, it appeared that the increased number of particles was derived from the amphotericin B-deoxycholate complex containing substances such as silicone released from the vial components. Amphotericin B fluid made with intravenous fluids after reconstitution also contained particles over the acceptable limits according to the Japanese or US pharmacopoeia. CONCLUSION: These findings suggest that reconstituted solutions should be filtered with membrane filters and diluted fluids with in-line filters.

Rapid determination of amphotericin B in serum and urine by third-order derivative spectrophotometry.

A derivative spectrophotometric method for rapid monitoring of amphotericin B in serum and urine down to 30 ng/mliters is described. Samples are treated with acetonitrile, and amphotericin B is directly quantified in the crude extracts on the basis of the intensity of the peak that appears at 402 nm when the normal absorption spectrum is submitted to third-order derivative processing. Accuracy data suggested recoveries in the range of 84.3-94.9% for serum and 85.6-93.4% for urine. The precision of the method was better than 11.3% for serum and 9.2% for urine when samples contained as low as 29.6 ng/mliters of amphotericin B. Ease of applicability, short analysis time, low cost, and reliability are the main advantages of the method.

[Study of components of standard samples of polyene macrolide antibiotics by the methods of high performance liquid chromatography and SIEAP mass spectrometry]. / Izuchenie komponentnogo sostava standartnykh obraztsov polienovykh makrolidnykh antibiotikov metodom vysokoéffektivnoi zhidkostnoi khromatografii i mass-spektrometrii ERIAD.

The component composition of reference samples of polyenic macrolide antibiotics such as nystatin, mycoheptin, amphotericin B and levorin was studied by HPLC and chromatographic mass spectrometry in comparison to the WHO standards. It was shown that the samples were close by their component composition to the analogous samples of the WHO standards.

Terapia de la paracoccidioidomicosis: valor actual de los antiguos tratamientos / Therapy of the paracoccidioidomycosis: actual value of the treatment old

Para establecer la debida perspectiva, he tratado de evocar las condiciones de la era pre-terapéutica, citando autores clásicos, que describen el resignado dolor de los pacientes y la humillada impotencia de los médicos. La época de los tratamientos antiguos fue de esperanza parcialmente frustada. Considero antiguos los tratamientos de la paracoccidioidomicosis anteriores a los imidazólicos de uso oral. Antiguos y obsoletos. De ellos nos queda el valor de la experiencia que nos permitieron adquirir. Creo que fue posible en teoría erradicar la infección de todos los pacientes mediante la administración de sulfas en dosis correctas durante dos o tres años, pero en la práctica, no se logró la suficiente claridad de criterios, ni hubo, ni hay manera de acompañar a los pacientes durante ese tiempo. Las sulfas nos enseñaron que la paracoccidioidomicosis debe tratarse ininterrumpidamente por largo tiempo. Estimo que la anfotericina permitió prolongar la vida de la mayoría de los pacientes y erradicar la infección de la mitad de los pacientes tratados correctamente. Por el alto costo y la gran atención que requiere su administración, nunca ha sido una opción liberadora en paracoccidioidomicosis. La anfotericina nos enseñó que la calidad del médico es fundamental para la conducción exitosa del tratamiento. Finalmente, en muchos países del área endémica, persiste la negligencia del estado para con la asistencia a los pacientes después de su salida del hospital, igual que en los tiempos aciagos de los tratamientos antiguos. De todos modos, considero un evento feliz el haber superado los tratamientos antiguos

An internally-standardized assay for amphotericin B in tissues and plasma.

A high-performance liquid chromatographic (HPLC) method with p-nitrophenol as internal standard is described for the rapid analysis of amphotericin B recovered by methanolic extraction from tissues and plasma. Programmed, gradient elution of the ODS column was used with detection by tungsten light at 388 nm. Standard curves were derived based on the peak height ratios. The lowest reproducible limit of the assay was 0.04 micrograms/ml with plasma. The extraction and chromatographic procedures recovered 53-71% of the amphotericin B from each of these sources. The coefficient of variation of the recovery ratios was less than 18% from plasma over a range of concentrations of amphotericin B from 0.08 to 10.0 micrograms/ml. Recovery from tissues, studied over a narrower concentration range, showed a similar degree of precision. Variations in precolumns apparently resulting in selective binding of the amphotericin B were found to have a systematic but important influence on recovery efficiency. No substances were detected which interfered with the assay procedures as described. By incorporating an internal standard we have enhanced the reliability and flexibility of the HPLC assay for amphotericin B especially for assay of tissues.

International standards and international reference preparations: amphotericin B, vancomycin, capreomycin, cefalotin, demethylchlortetracycline, gentamycin, gramicidin S, kanamycin and kanamycin B, lincomycin, lymecycline, methacycline, paromomycin, rifamycin SV, ristocetin and ristocetin B, spiramycin, and triacetyloleandomycin.

Each of the preparations described here was obtained and evaluated at the request of a WHO Expert Committee on Biological Standardization. Unless otherwise stated, a standard procedure was used to distribute the material into individual ampoules. The procedure was as follows. Upon receipt by the National Institute for Medical Research (NIMR), London, materials were stored temporarily in the dark at a temperature of -10 degrees C or lower, and protected from moisture. At a convenient time they were brought back to room temperature, mixed, and distributed into individual neutral glass ampoules so that each ampoule contained 50-100 mg of powder. If it was known that the material was light-sensitive non-actinic glass ampoules were used. After exhaustive drying in vacuum over phosphorus(V) oxide, the ampoules were either constricted (up to 1963) or fitted with capillary leak plugs, dried for a further period under the same conditions, filled with dry nitrogen, and sealed by fusion of the glass. The total drying period varied from 8 to 38 days according to the nature of the material. After they had been tested for leaks, the ampoules were stored in the dark at -20 degrees C.