Combined environmental risk assessment for the antiviral pharmaceuticals ganciclovir and valganciclovir in Europe.

Potential environmental risks of the old antiviral pharmaceuticals ganciclovir (GCV) and valganciclovir (VGCV) were reassessed based on new environmental fate and chronic ecotoxicity tests and on actual use data for Europe. Valganciclovir is hydrolyzed to GCV by intestinal and hepatic esterases, and hence the new environmental tests only refer to GCV. A sorption study showed that GCV will not sorb significantly, excluding the soil as a relevant environmental compartment. Despite earlier data suggesting nondegradability, a new water/sediment fate test showed GCV to be primarily and ultimately degraded and to be nonpersistent. The chronic ecotoxicity tests with algae and daphnids resulted in no inhibition at the highest tested concentrations, whereas a fish partial life cycle test, selected in view of mammalian mutagenicity and reprotoxicity data, showed effects on growth of the young fish, but not on gametogenesis, fertilization, embryogenesis, or teratogenicity. Predicted environmental concentrations were derived based on actual per capita use data for European countries for 2004 to 2014, and the highest was selected for the risk assessment. A comparison of predicted environmental concentrations with predicted no-effect concentrations shows no significant risk for wastewater treatment, surface waters, groundwater, or sediment. In addition, potential risks to (semi)aquatic top predators or to human consumers of water and fish are exceedingly low. Environ Toxicol Chem 2017;36:2205-2216. © 2017 The Author. Environmental Toxicology and Chemistry Published by Wiley Periodicals, Inc. on behalf of SETAC.

Caracterização do estado sólido de ganciclovir / Solid state characterization of ganciclovir

O presente trabalho teve como objetivo o estudo do estado sólido do ganciclovir (GCV) e suas diferentes formas polimórficas. O GCV é um fármaco antiviral útil no tratamento de infecções por citomegalovírus (CMV). Embora seja um fármaco amplamente usado, poucos estudos têm sido realizados sobre seu estado sólido. Atualmente, o GCV é conhecido por apresentar quatro formas cristalinas, duas anidras (Forma I e II) e duas hidratas (III e IV). Neste trabalho, nós reportamos a solução da estrutura cristalográfica da Forma I do GCV, que foi encontrado durante o screening de cristalização do fármaco, em que nove ensaios de cristalização (GCV-1, GCV-A, GCV-B, GCV-C, GCV-D, GCV-E, GCV-F, GCV-G e GCV-H) foram realizados e os materiais resultantes foram caracterizados por Difratometria de raios X (DRX), análise térmica (DTA/TG) e Hot Stage Microscopy. De todas as cristalizações realizadas foram obtidas quatro formas sólidas, denominadas como Forma I (GCV-1, GCV-B e GCV-H), Forma III (GCV-C, GCV-D, GCV-F e GCV-G), Forma IV (GCV-A) e Forma V (GCV-E). Esta última está sendo descrita pela primeira vez na literatura e indica a presença de outra forma hidratada de GCV. As Formas I, III e IV corresponderam a forma anidra e as duas formas hidratadas do fármaco, respectivamente. Além disso, foi evidenciado por experimentos de conversão de slurry e análise térmica que o cristalizado de GCV-1 (Forma I) foi o mais estável entre os materiais obtidos, e este deu origem ao monocristal da Forma I de GCV, estrutura cristalina anidra do fármaco. Neste trabalho, pela primeira vez, a estrutura cristalina deste composto foi definida por cristalografia de raios X de monocristal. A análise estrutural mostrou que a Forma I do fármaco cristaliza no grupo espacial monoclínico P21/c e está composta por quatro moléculas de GCV na sua unidade assimétrica. Cada molécula está unida intermolecularmente por ligações de hidrogênio, que dão lugar à formação de cadeias infinitas e estas por sua vez se arranjam de maneira a formar uma estrutura tridimensional.

This presented work aims to study the solid state of ganciclovir (GCV) and its different polymorphic forms. GCV is an antiviral drug useful in the treatment of cytomegalovirus (CMV) infections. Although it is a widely-used drug, few studies have been conducted on its solid state. Currently, GCV is known to have four crystalline forms, two anhydrous (Form I and II) and two hydrates (III and IV). In this investigation, we report a successful preparation of GCV Form I and its crystallographic structure, which was found during the crystallization of the drug, in which nine crystallization tests (GCV-1, GCV-A, GCV-B, GCV- D, GCV-E, GCV-F, GCV-G and GCV-H) were performed and the resulting materials were characterized by X-ray diffractometry (XRD), thermal analysis (DTA/TG) and Hot Stage Microscopy. Of all the crystallizations performed, four solid forms were obtained, denoted as Form I (GCV-1, GCV-B and GCV- H), Form III (GCV-C, GCV-D, GCV-F and GCV-G), Form IV (GCV-A) and Form V (GCV-E). The latter is being described for the first time in the literature and indicates the presence of another hydrated form of GCV. Forms I, III and IV corresponded to the anhydrous form and the two hydrated forms of the drug, respectively. In addition, it was evident by both the slurry conversion and the thermal analysis methods that the GCV-1 crystallized (Form I) was indeed the most stable amongst the materials obtained. This gave rise to GCV Form I monocrystal, anhydrous crystalline structure of the drug. The compound was characterized by monocrystal X-ray crystallography. The structural analysis showed that Form I of the drug crystallized in the monoclinic system space group P21/c is composed of four molecules of GCV in its asymmetric unit. Each molecule is linked intermolecularly by hydrogen bonds, which give rise to the formation of infinite chains arranged in a way that form a three-dimensional structure.

The fabrication of a new electrochemical sensor based on electropolymerization of nanocomposite gold nanoparticle-molecularly imprinted polymer for determination of valganciclovir.

A new sensitive and selective electrochemical sensor was successfully developed for the determination of valganciclovir. It is based on one-step electropolymerization of the molecularly imprinted polymer composed from 2,2'-dithiodianiline, gold nanoparticles, and valganciclovir on a glassy carbon electrode modified with carboxyl-functionalized multiwalled carbon nanotubes via cyclic voltammetry. The gold nanoparticles were introduced into the polymer composite for the development of electrical response by facilitating charge transfer. The fabrication process of the sensor was characterized by cyclic voltammetry and electrochemical impedance spectroscopy. Under the optimized condition calibration curve of the imprinted sensor has two linear concentration ranges from 1.0 to 500.0nM and 500.0 to 2000.0nM, with the limit of detection of 0.3nM. The relative standard deviation (RSD) for seven parallel determination of 1.0µM valganciclovir at optimum conditions was found to be 2.9%. The imprinted sensor has the advantages of high porous surface structure, ease of preparation, good reproducibility, good repeatability and high selectivity and sensitivity. Furthermore, the proposed method was successfully intended for the determination of valganciclovir in real samples.

Liquid chromatography tandem mass spectrometry quantitation of intracellular concentrations of ganciclovir and its phosphorylated forms.

Ganciclovir (GCV) is prescribed for cytomegalovirus infection which is a major issue in immunodepressed patients. It is however characterized by hematological toxicity. A better understanding of GCV concentration-effects relationships implies the measurement of intracellular forms. The objective of this study was to develop a method to measure GCV and its derivatives in cells. A four-stage procedure was developed with the following strategy: (1) to separate into different fractions the different intracellular forms of GCV (GCV itself and its phosphorylated forms) by solid-phase extraction (SPE) from blood cells, (2) to dephosphorylate the different phosphorylated forms into GCV, (3) to perform a second SPE to desalt samples and concentrate GCV, and (4) to measure GCV concentrations in the different extracts using a triple-quadrupole, linear ion trap mass spectrometer. Finally, the procedure was tested in 17 patients receiving GCV. From lysed cells, the different forms of GCV were fractionated, the phosphorylated forms were eluted with different KCl solutions, and the obtained fractions were treated with acid phosphatase to transform the phosphorylated metabolites back into GCV. The method was validated from 5 to 500 µg L(-1) with a limit of detection of 1 µg L(-1). The whole procedure was validated according to the US Food and Drug Administration guidelines and successfully applied in 17 patients receiving GCV. The method liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) allowing the measurement of GCV and its phosphorylated forms in blood cells was developed and can be used in developing clinical studies to explore the role of these biomarkers in the event of toxicity.

Development of a RP-LC method for a diastereomeric drug valganciclovir hydrochloride by enhanced approach.

Two methods, prescribed in USP, for the analysis of related substances of valganciclovir hydrochloride drug substance, were evaluated in terms of selectivity and ease of use. A new, simple, selective, stability indicating and user friendly RP-LC method was developed for related substances analysis. The developed single method is capable of separating all known impurities, which are quantified by two methods of USP. A central composite design was applied to optimize the critical chromatographic parameters. A multi step gradient program was strategically designed and a part of the program was optimized through Design of Experiments. Separation was achieved with a Zorbax SB C18 column with 0.1% trifluoro acetic acid and methanol in gradient elution. Design space is proposed graphically for the robust operation of the method. The method is linear, precise and accurate from LOQ level to 150% level of specification limit of impurities. Simple modification in the gradient program with reduced run time is also proposed for assay and diastereomer ratio.

Utility of certain sigma- and pi-acceptors for the spectrophotometric determination of ganciclovir in pharmaceutical formulations.

Studies were carried out, for the first time, to investigate the charge-transfer reactions of ganciclovir as n-electron donor with the sigma-acceptor iodine and various pi-acceptors: 7,7,8,8-tetracyanoquinodimethane; tetracyanoethylene; 2,3-dichloro-5,6-dicyano-1,4-benzoquinone; p-chloranilic acid; 2,3,5,6-tetrabromo-1,4-benzoquinone; 2,3,5,6-tetrachloro-1,4-benzoquinone and 2,4,7-trinitro-9-fluorenone. The formation of the colored charge-transfer complexes was utilized in the development of simple, rapid and accurate spectrophotometric methods for the analysis of ganciclovir in pure form as well as in its pharmaceutical formulation (capsules). Different variables affecting the reactions were studied and optimized. Under the optimum reaction conditions, linear relationships with good correlation coefficients (0.9993-0.9998) were found between the absorbance and the concentration of ganciclovir in the range of 2.0-240 microg mL(-1). For more accurate analysis, Ringbom optimum concentration range was found to be between 5.0 and 225 microg mL(-1). The limits of detection ranged from 0.36 to 2.45 microg mL(-1) and the limits of quantification ranged from 1.20 to 8.17 microg mL(-1). A Job's plot of the absorbance versus the molar ratio of ganciclovir to each of acceptors under consideration indicated (1:1) ratio. The proposed methods were applied successfully for simultaneous determination of ganciclovir in capsules with good accuracy and precision and without interferences from common additives. The recovery percentages ranged from 99.45+/-0.73% to 100.35+/-1.40%. The results were compared favourably with the reported method.

Determination of acyclovir in horse plasma and body fluids by high-performance liquid chromatography combined with fluorescence detection and heated electrospray ionization tandem mass spectrometry.

Two methods are presented for the determination of 'respectively' the plasma protein unbound and total concentration of acyclovir in horse plasma and body fluids: first, a liquid-liquid extraction was performed on plasma, combined with HPLC-fluorescence detection for the total plasma concentration; second a more sensitive method using high-performance liquid chromatography combined with heated electrospray ionization tandem mass spectrometry (LC-HESI-MS/MS) was described for plasma and for body fluids analysis. To obtain the unbound concentration of acyclovir in plasma, a simple deproteinization step using a Microcon filter was performed. Ganciclovir was used as an internal standard. Analysis was carried out on an Inertsil 5 ODS-3 column for the HPLC-fluorescence method. For the LC-HESI-MS/MS method a PLRP-S column was used. The limit of quantification (LOQ) for the total concentration was set at 50 and 2 ng mL(-1) for the HPLC-fluorescence method and the LC-HESI-MS/MS method, respectively. The limit of quantification for the unbound concentration was set at 5 ng mL(-1) and at 2 ng mL(-1) for body fluids. The methods were successfully used to perform pharmacokinetic and clinical studies in horses after intravenous and oral dosage of acyclovir and its prodrug valacyclovir.

Determination of ganciclovir in different matrices from solid organ transplanted patients treated with a wide range of concomitant drugs.

The aim of the present study was to develop a time-efficient chromatographic method for the analysis of therapeutic concentrations of ganciclovir (GCV) in plasma, urine as well as dialysate (from continuous renal replacement therapy) from solid organ transplant recipient treated with either GCV or its prodrug valganciclovir (VGCV) in combination with a wide variety of other concomitant drugs. Sample preparation was performed by reversed phase solid phase extraction and was followed by separation of the analytes on a reversed phase column using isocratic elution with a mobile phase consisting of acetonitrile-a counter ion (50 mM 1-heptanesulfonic acid) in an aqueous sodium dihydrogen phosphate buffer (pH 2.1; 10 mM) (10:90 v/v) and a fluorescence detector. Validation of the method showed linearity within the concentration range of 0.1-40 microg/mL for plasma and 0.1-120 microg/mL for urine and dialysate (R(2)>0.99, n> or =5). Accuracy and precision (evaluated at 0.1, 5 and 40 microg/mL) were both satisfactory. The LLOQ was determined to be 0.1 microg/mL. The method was successfully applied on clinical samples from renal transplant recipients treated with VGCV in combination with a variety of usually used concomitant drugs for solid organ transplant recipients.

Development and validation of a new reversed-phase ion pairing liquid chromatographic method with fluorescence detection for penciclovir analysis in plasma and aqueous humor.

A simple, sensitive and reliable HPLC ion-pairing method with fluorescence detection, was developed for penciclovir determination in plasma and aqueous humor, with a Zorbax SB-aq C18 (100 mmx2.1 mm) column. Plasma samples were treated by solid-phase extraction with Oasis MCX (30 mg) cartridges. Ganciclovir, an antiviral drug structurally related to penciclovir, was used as internal standard (I.S.). Aqueous humor samples were directly injected into the chromatographic system. Separation was performed by a gradient elution with a mobile phase consisting of a mixture of acetonitrile and phosphate buffer 50mM containing 5mM of sodium octanesulfonate, pH 2.0, at a flow rate of 0.3 ml/min. The method was validated and showed good performances in terms of linearity, sensitivity, precision and trueness. Quantification limit was obtained at 0.05 microg/ml for aqueous humor and at 0.1 microg/ml for plasma. Finally, the proposed analytical method was used to measure penciclovir in clinical samples for a pharmacokinetic study, after oral administration of famciclovir.

Sustained release of ganciclovir from poly(lactide-co-glycolide) microspheres.

Biodegradable poly(lactide-co-glycolide) microspheres loaded with ganciclovir were produced using the emulsification/solvent evaporation technique. The effects of drug-to-polymer ratio and dispersion time on the drug content in the microspheres were investigated. The release rate of the drug was studied for 20 weeks in a phosphate buffered solution of pH 7 at 37 degrees C. Data revealed that lower drug content was obtained with increasing drug-to-polymer ratio and decreasing dispersion time. The release of the drug followed a triphasic release pattern, i.e. an initial burst, a diffusive phase and a second burst. The initial burst occurred within the first 2 days of immersion. After the burst, the release was by diffusion for up to 13 weeks, followed by another burst release, which signals the onset of bulk degradation of the polymer. Gel permeation chromatography (GPC), differential scanning calorimetry (DSC), scanning electron microscopy (SEM) and ultraviolet spectroscopy (UV) were used to follow the hydrolytic degradation and drug release rate of the microspheres.

Positron-emission tomography reporter gene expression imaging in rat myocardium.

BACKGROUND: This study examines the quantitative accuracy, detection sensitivity, and time course of imaging the expression of a mutant herpes simplex type-1 virus thymidine kinase (HSV1-sr39tk) PET reporter gene in rat myocardium by using the PET reporter probe 9-(4-[18F]-Fluoro-3-Hydroxymethylbutyl)-Guanine ([18F]-FHBG) and a small-animal PET (microPET). METHODS AND RESULTS: In 40 rats, adenovirus expressing HSV1-sr39tk driven by a cytomegalovirus promoter (Ad-CMV-HSV1-sr39tk, 1x10(6) to 1x10(9) pfu) was injected through a thoracotomy directly into the left ventricular myocardium. After 3 days, myocardial perfusion was imaged with [13N]-ammonia for delineating the left ventricular myocardium, followed by imaging the expression of the reporter gene with intravenous [18F]-FHBG. The total myocardial [18F]-FHBG accumulation was quantified in percent of injected dose (%ID). Immunohistochemistry and autoradiography demonstrated HSV1-sr39tk enzyme (HSV1-sr39TK) and accumulation of [18F]-FHBG in the inoculated myocardium in 3 rats each. In 24 rats with various viral titers, the %ID was correlated with ex vivo well counting (r2=0.981, P<0.0001) and myocardial HSV1-sr39TK activity by tissue enzyme activity assay (r2=0.790, P<0.0001). Myocardial [18F]-FHBG accumulation was identified at viral titers down to 1x10(7) pfu. In 6 rats serially imaged up to day 17, myocardial [18F]-FHBG accumulation on microPET peaked on days 3 to 5 and was no longer identified on days 10 to 17. CONCLUSIONS: HSV1-sr39tk reporter gene expression can be monitored with [18F]-FHBG and microPET in rat myocardium quantitatively and serially with high detection sensitivity. Cardiac PET reporter gene imaging offers the potential of monitoring the expression of therapeutic genes in cardiac gene therapy.

Sustained release of low-dose ganciclovir from a silicone formulation prolonged the survival of rats with gliosarcomas under herpes simplex virus thymidine kinase suicide gene therapy.

A silicone formulation of ganciclovir (GCV-pellet) was developed to enhance the cytotoxic effects of herpes simplex virus thymidine kinase suicide gene therapy. The effectiveness of this drug delivery system was assessed in a rat 9L gliosarcoma model. The GCV-pellets (1 mm in length and in diameter) used in this experiment contained a total amount of 0.15 mg of GCV. In vitro experiments demonstrated that GCV was gradually released over a period of 7 days. Five days after stereotactic tumor inoculation into the right caudate nucleus, a herpes simplex virus type 1 (HSV-1) vector expressing herpes simplex virus thymidine kinase (HSV-tk) (T1, 2x10(6) pfu) was administered at the same location. The survival rate of the group treated with the GCV-pellet was compared with that of the T1 group injected intraperitoneally (IP) with GCV (30 mg/kg/day for 7 days). The GCV-pellet-treated group had a significantly prolonged survival (a median of more than 80 days) compared with the GCV IP group (a median of 65 days) and with control groups (P<0.05). The control groups (untreated or receiving only the virus vector) had a survival of 35-38 days. The survival rate of the GCV-pellet group over 80 days was 75%, and all the rats that survived more than 80 days and did not show tumors upon histological examination of the brain were deemed cured. No toxic effects or immunological reactions were observed histologically around the pellet in brain sections from the rats treated with the GCV-pellet. After GCV-pellet inoculation into the tumor, drug concentrations were kept at 1-10 microg/g tissue for 3-4 days. When the same dose of GCV (0.15 mg) in aqueous solution was injected into the tumor, GCV concentrations reached a peak of 0.5 mg/g tissue after 30 min and decreased below measurable level within 12 h. After IP injections of 3 mg GCV, GCV concentrations in the tumor reached a peak of 5.7 microg/g tissue after 30 min and also decreased below measurable level within 12 h. This sustained release of a low and effective GCV dose with the silicone formulation significantly prolonged survival in combinations with HSV-tk expression if compared to IP administration of GCV. Histological examination suggests that the treatment appears to be safe.

Development of a sensitive method for the determination of ganciclovir by reversed-phase high-performance liquid chromatography.

Ganciclovir is a nucleoside analogue widely used in the treatment of cytomegalovirus infections, which affects mainly immunocompromised patients. Recently, new pharmaceutical dosage forms based on the use of albumin nanoparticles have been developed for improving the efficacy of this drug. The aim of this study was to develop an analytical HPLC method for the determination of ganciclovir in both pharmaceuticals (i.e. albumin nanoparticles) and biological medium samples. The chromatography was performed on a reversed-phase encapped column (LiChrospher Select B C8) with a mobile phase consisting of acetonitrile in 0.05 M ammonium acetate (pH 6.5; 2: 98, v/v). Acyclovir was used as internal standard and the detection wavelength was 254 nm. The limit of quantitation of ganciclovir was 50 ng/ml and the average recoveries over a concentration range of 0.05-10 microg/ml ranged from 98 to 102%. Precision did not exceed 5%. In summary, this assay is a selective, sensitive and reproducible method for the determination of the ganciclovir in albumin nanoparticles. It can be successfully applied to the estimation of the ganciclovir uptake by cultured human corneal fibroblasts.

An HPLC method for the determination of diastereomeric prodrug RS-79070-004 in human plasma.

Ganciclovir is an antiviral nucleoside analogue approved for treatment and prevention of cytomegalovirus infections in immunocompromised subjects. RS-79070-194, a diastereomeric monovalyl ester of ganciclovir (hydrochloride salt), is under evaluation as a prodrug to increase the bioavailability of ganciclovir. An HPLC method with column switching has been developed and validated for quantification of the corresponding free base RS-79070-004 in human plasma. In the method, proteinaceous material in 0.25 ml of plasma is precipitated by trichloroacetic acid. An aliquot of the supernatant is analyzed by HPLC, with automated column switching to remove late-eluting materials that might interfere with the analyte peaks in subsequent runs. Detection of RS-79070-004 is by UV lambda = 254 nm). The peak areas for each isomer are summed to generate a value for total RS-79070-004. The method has a validated range of 0.0400-4.00 microg/ml and a lower limit of quantification of 0.0400 microg/ml. All intra- and inter-assay %CVs were < 7.5%, and all recoveries (accuracy) were within 6% of nominal values. No interference was observed by ganciclovir, caffeine, acetaminophen, or ibuprofen. Analyte stability in plasma and in the sample extracts is adequate for the specified collection, storage, and analysis conditions. The validated method has been successfully used to analyze clinical study samples.

Immunohistochemical localization of ganciclovir in the human retina.

PURPOSE: To localize ganciclovir in the retina of human eyes treated with intravenous or intravitreal ganciclovir for cytomegalovirus (CMV) retinitis. METHODS: Paraffin-embedded five-micron sections of autopsy eyes were obtained from seven patients as follows: two patients with CMV retinitis treated with intravenous ganciclovir; two patients with CMV retinitis treated with an intravitreal sustained-release ganciclovir device; one patient with CMV retinitis treated with intravenous foscarnet; and two patients with AIDS without CMV retinitis who did not receive any anti-CMV therapy. The paraffin was removed from the sections, and indirect immunofluorescent staining was performed, using an antiserum to ganciclovir. RESULTS: Bright fluorescent staining was noted in the retinal pigment epithelium (RPE) and photoreceptor outer segments of eyes treated with intravenous or intravitreal ganciclovir, but not in eyes treated with foscarnet or without CMV retinitis. In addition, patches of bright fluorescent staining of the internal limiting membrane was noted in eyes treated with intravitreal ganciclovir. CONCLUSIONS: Ganciclovir is detected in the outer retina of patients with CMV retinitis treated with intravenous or intravitreal therapy. The drug is detected also in the internal limiting membrane in eyes treated with the intravitreal sustained-release ganciclovir device.

Release kinetics of liposome-encapsulated ganciclovir after intravitreal injection in rabbits.

This study was undertaken to establish experimentally whether the intravitreal application of liposomally-entrapped ganciclovir could prolong intraocular therapeutic levels when it is compared to the intravitreal injection of a simple solution of the drug. New Zealand white rabbits were given an intravitreal injection of the drug solution and of liposome-encapsulated ganciclovir. The intravitreal clearance of ganciclovir was determined after a single injection of either the drug solution (200 micrograms/0.1 mL) or the liposomally-entrapped (with 41% load; 82 micrograms drug load and 118 micrograms free) ganciclovir. The ganciclovir vitreal concentrations were measured at various time intervals for a period up to 43 days using an HPLC method. The results of this study clearly demonstrated that prolonged intravitreal drug levels (above the mean inhibitory dose of cytomegalovirus of 1 microgram/mL) after administration of the liposome-entrapped ganciclovir and estimated to continue beyond 30-43 days. The injection of the 200 micrograms/0.1 mL of drug solution showed a mean vitreous concentration which was higher than the ID50 only for 55 h. The disappearance rate constant for the liposome-encapsulated injections was approximately 22 x slower than simple drug solution injections (controls). No evidence of retinal toxicity was found by clinical or light microscopy examination of the treated eyes.

Sample preparation for the determination of purine nucleotide analogues in tissues.

A sample treatment procedure for the determination of thiopurine and ganciclovir nucleotides in human tissues was developed. Owing to the lack of suitable standards for most of the active nucleotide analogues, the procedure was based on two steps: (1) perchloric acid homogenization and deproteinization of the tissue specimen and (2) conversion of purine nucleotides into parent drug or free bases by enzymatic or acid hydrolysis. The parent drug or purine bases formed were then analyzed on a Hypersil ODS column using isocratic elution with dihydrogenphosphate buffer for ganciclovir nucleotides or the gradient elution mode with dihydrogenphosphate buffermethanol for thiopurine nucleotides. The sample treatment procedure was evaluated using guanosine triphosphate (GTP), 6-thioinosinic acid (6TIMP) and 6-thioguanosine monophosphate (6TGMP) as standards. Mean analytical recoveries determined by adding known concentrations of standards to the tissue specimen before sampling processing were higher than 97%. The sample preparation described is simple and represents a suitable method for the investigation of active nucleotide pool in tissues.

High-performance liquid chromatographic determination of ganciclovir nucleotides in human myocardial tissue.

A method for the analysis of ganciclovir nucleotides in myocardial tissues was developed. The antiviral effect of ganciclovir is attributed to intracellular ganciclovir nucleotides. The procedure is based on perchloric acid deproteinization and enzymatic hydrolysis of the ganciclovir nucleotides to ganciclovir. Then, the parent drug was analyzed on a Hypersil ODS column using potassium dihydrogenphosphate buffer as mobile phase. The mean analytical recovery of ganciclovir from myocardial tissue was 101 +/- 2% and the detection limit was 2 pmol. The sample treatment procedure described is simple and presents a suitable analytical tool for the investigation of the ganciclovir nucleotides pool in tissues.

Stability of ganciclovir sodium and amino acids in parenteral nutrient solutions.

The stability of ganciclovir sodium and amino acids in parenteral nutrient solutions was studied. Three admixtures of ganciclovir sodium plus parenteral nutrient solution were prepared, one containing ganciclovir sodium 0.83 mg/mL, 1% amino acids, and 10% dextrose injection; one containing ganciclovir sodium 1.4 mg/mL, 2.5% amino acids, and 10% dextrose injection; and one containing ganciclovir sodium 1.4 mg/mL, 5% amino acids, and 25% dextrose injection. The solutions were visually inspected for precipitates, color change, and gas formation and were tested for pH. High-performance liquid chromatography was used to measure the concentration of ganciclovir and 16 amino acids in each admixture immediately and one, two, and three hours after preparation. There was no evidence of visual incompatibility in any of the admixtures, and pH did not vary appreciably during the study. The mean ganciclovir sodium concentration remaining was greater than 100% of the initial concentration for all the admixtures at one, two, and three hours. The mean amino acid concentration remaining in the admixtures with 2.5% or 5.0% amino acids was greater than 90% of the initial concentration for each amino acid at one, two, and three hours. The mean amino acid concentration remaining in the admixture with 1% amino acids was greater than 90% of the initial level at one and two hours. Ganciclovir sodium 0.83 mg/mL was stable for at least three hours in parenteral nutrient solution with 1% amino acids, and ganciclovir sodium 1.4 mg/mL was stable for at least three hours in admixtures with 2.5% or 5% amino acids.(ABSTRACT TRUNCATED AT 250 WORDS)