Nano-radiopharmaceuticals as therapeutic agents.

In recent years, there has been an increased interest in exploring the potential synergy between nanotechnology and nuclear medicine. The application of radioactive isotopes, commonly referred to as radiopharmaceuticals, is recognized in nuclear medicine for diagnosing and treating various diseases. Unlike conventional pharmaceutical agents, radiopharmaceuticals are designed to work without any pharmacological impact on the body. Nevertheless, the radiation dosage employed in radiopharmaceuticals is often sufficiently high to elicit adverse effects associated with radiation exposure. Exploiting their capacity for selective accumulation on specific organ targets, radiopharmaceuticals have utility in treating diverse disorders. The incorporation of nanosystems may additionally augment the targeting capability of radiopharmaceuticals, leveraging their distinct pharmacokinetic characteristics. Conversely, radionuclides could be used in research to assess nanosystems pharmacologically. However, more investigation is needed to verify the safety and effectiveness of radiopharmaceutical applications mediated by nanosystems. The use of nano-radiopharmaceuticals as therapeutic agents to treat various illnesses and disorders is majorly covered in this review. The targeted approach to cancer therapy and various types of nanotools for nano-radiopharmaceutical delivery, is also covered in this article.

Size, Surface Properties, and Ion Release of Zinc Oxide Nanoparticles: Effects on Cytotoxicity, Dopaminergic Gene Expression, and Acetylcholinesterase Inhibition in Neuronal PC-12 Cells.

The extensive applications of zinc oxide nanoparticles (ZnO NPs) have resulted in a substantial risk of human exposure. However, the knowledge of the toxicity of these NPs in the nervous system is still limited. A comparative analysis of ZnO NPs of various sizes and NPs of the same size, with and without surface coating, and the potential role of released zinc ions is yet to be thoroughly explored. As a result, we have studied the cellular toxicity of two different-sized ZnO NPs, ZnO-22 (22 nm) and ZnO-43 (43 nm), and NPs with similar size but with polyvinylpyrrolidone coating (ZnO-P, 45 nm). The findings from our study suggested a time-, size-, and surface coating-dependent cytotoxicity in PC-12 cells at a concentration ≥ 10 µg/ml. ZnO NP treatment significantly elevated reactive oxygen and reactive nitrogen species, thereby increasing oxidative stress. The exposure of ZnO-22 and ZnO-43 significantly upregulated the expression of monoamine oxidase-A and downregulated the α-synuclein gene expression associated with the dopaminergic system. The interaction of NPs enzymes in the nervous system is also hazardous. Therefore, the inhibition activity of acetylcholinesterase enzyme was also studied for its interaction with these NPs, and the results indicated a dose-dependent inhibition of enzyme activity. Particle size, coating, and cellular interactions modulate ZnO NP's cytotoxicity; smaller sizes enhance cellular uptake and reactivity, while coating reduces cytotoxicity by limiting direct cell contact and potentially mitigating oxidative stress. Furthermore, the study of released zinc ions from the NPs suggested no significant contribution to the observed cytotoxicity compared to the NPs.

Homeopathic Formulations of Syzygium jambolanum Alleviate Glycation-Mediated Structural and Functional Modifications of Albumin: Evaluation through Multi-Spectroscopic and Microscopic Approaches.

BACKGROUND: The growing interest in identifying the mode of action of traditional medicines has strengthened its research. Syzygium jambolanum (Syzyg) is commonly prescribed in homeopathy and is a rich source of phytochemicals. OBJECTIVE: The present study aims to shed light on the anti-glycation molecular mechanism of Syzyg mother tincture (MT), 30c, and 200c on glycated human serum albumin (HSA) by multi-spectroscopic and microscopic approaches. METHODS: The phytochemicals and antioxidant potential of the Syzyg formulations were estimated by the high-performance liquid chromatography and spectroscopic technique, respectively. Glycation was initiated by incubating HSA with methylglyoxal, three Syzyg formulations, and the known inhibitor aminoguanidine in separate tubes at 37°C for 48 hours. The formation of glycation adducts was assessed by spectrofluorometer and affinity chromatography. The structural modifications were analyzed through circular dichroism, Fourier transform infrared spectroscopy, turbidity, 8-anilinonapthalene-1-sulfonic acid fluorescence, and nuclear magnetic resonance. Further, the formation of the aggregates was examined by thioflavin T, native-polyacrylamide gel electrophoresis, and transmission electron microscopy. Additionally, the functional modifications of glycated HSA were determined by esterase-like activity and antioxidant capacity. The binding analysis of Syzyg formulations with glycated HSA was evaluated by surface plasmon resonance (SPR). RESULTS: Syzyg formulations MT, 30c, and 200c contained gallic acid and ellagic acid as major phytochemicals, with concentrations of 16.02, 0.86, and 0.52 µg/mL, and 227.35, 1.35, and 0.84 µg/mL, respectively. Additionally, all three formulations had remarkable radical scavenging ability and could significantly inhibit glycation compared with aminoguanidine. Further, Syzyg formulations inhibited albumin's structural and functional modifications. SPR data showed that Syzyg formulations bind to glycated HSA with an equilibrium dissociation constant of 1.10 nM. CONCLUSION: Syzyg formulations inhibited the glycation process while maintaining the structural and functional integrity of HSA.

Comprehensive Analysis of Titanium Oxide Nanoparticle Size and Surface Properties on Neuronal PC-12 Cells: Unraveling Cytotoxicity, Dopaminergic Gene Expression, and Acetylcholinesterase Inhibition.

Titanium oxide nanoparticles can penetrate the blood-brain barrier, infiltrate the central nervous system, and induce neurotoxicity. One of the most often utilized nanoparticles has been investigated for their neurotoxicity in many studies. Nonetheless, there remains an unexplored aspect regarding the comparative analysis of particles varying in size and nanoparticles of identical dimensions, both with and devoid of surface coating. In the current study, we synthesized two differently sized nanoparticles, TiO2-10 (10 nm) and TiO2-22 (22 nm), and nanoparticles of the same size but with a polyvinylpyrrolidone surface coating (TiO2-PVP, 22 nm) and studied their toxic effects on neural PC-12 cells. The results highlighted significant dose- and time-dependent cytotoxicity at concentrations ≥10 µg/mL. The exposure of TiO2 nanoparticles significantly elevated reactive oxygen and nitrogen species levels, IL-6 and TNF-α levels, altered the mitochondrial membrane potential, and enhanced apoptosis-related caspase-3 activity, irrespective of size and surface coating. The interaction of the nanoparticles with acetylcholinesterase enzyme activity was also investigated, and the results revealed a dose-dependent suppression of enzymatic activity. However, the gene expression studies indicated no effect on the expression of all six genes associated with the dopaminergic system upon exposure to 10 µg/mL for any nanoparticle. The results demonstrated no significant difference between the outcomes of TiO2-10 and TiO2-22 NPs. However, the polyvinylpyrrolidone surface coating was able to attenuate the neurotoxic effects. These findings suggest that as the TiO2 nanoparticles get smaller (towards 0 nm), they might promote apoptosis and inflammatory reactions in neural cells via oxidative stress, irrespective of their size.

Antiviral and ROS scavenging potential of Carica papaya Linn and Psidium guajava leaves extract against HIV-1 infection.

Antiretroviral therapy is the only treatment option for HIV-infected patients; however, it has certain drawbacks in terms of developing multiple toxic side effects. Thus, there is a continuous need to explore safe and efficacious anti-retroviral agents. Carica papaya Linn and Psidium guajava are known for their various biological activities. In this study, we characterized the bioactive fractions of methanolic leaves extract from both plants using the High-resolution electrospray ionization mass spectrometry (HR-ESI-MS) technique, followed by the investigation of their potential as anti-HIV-1 and antioxidant agents through in vitro mechanistic assays. The anti-HIV-1 activity was examined in TZM-bl cells through luciferase gene assay against two different clades of HIV-1 strains, whereas the intracellular ROS generation was analyzed by Fluorescence-Activated Cell Sorting. Additionally, the mechanisms of action of these phyto-extracts were determined through the Time-of-addition assay. The characterization of Carica papaya Linn and Psidium guajava leaves extract through HR-ESI-MS fragmentation showed high enrichment of various alkaloids, glycosides, lipids, phenolic compounds, terpenes, and fatty acids like bioactive constituents. Both the phyto-extracts were found to be less toxic and exhibited potent antiviral activity against HIV-1 strains. Furthermore, the phyto-extracts also showed a decreased intracellular ROS in HIV-1 infected cells due to their high antioxidant potential. Overall, our study suggests the anti-HIV-1 potential of Carica papaya Linn and Psidium guajava leaves extract due to the synergistic action of multiple bioactive constituents.

Toxic implications of silver nanoparticles on the central nervous system: A systematic literature review.

Silver nanoparticles have many medical and commercial applications, but their effects on human health are poorly understood. They are used extensively in products of daily use, but little is known about their potential neurotoxic effects. A xenobiotic metal, silver, has no known physiological significance in the human body as a trace metal. Biokinetics of silver nanoparticles indicates its elimination from the body via urine and feces route. However, a substantial amount of evidence from both in vitro and in vivo experimental research unequivocally establish the fact of easier penetration of smaller nanoparticles across the blood-brain barrier to enter in brain and thereby interaction with cellular components to induce neurotoxic effects. Toxicological effects of silver nanoparticles rely on the degree of exposure, particle size, surface coating, and agglomeration state as well as the type of cell or organism used to evaluate its toxicity. This review covers pertinent facts and the present state of knowledge about the neurotoxicity of silver nanoparticles reviewing the impacts on oxidative stress, neuroinflammation, mitochondrial function, neurodegeneration, apoptosis, and necrosis. The effect of silver nanoparticles on the central nervous system is a topic of growing interest and concern that requires immediate consideration.

Amelioration of Cisplatin-induced Renal Inflammation by Recombinant Human Golimumab in Mice.

BACKGROUND: One of the most commonly used anti-cancer agents, Cisplatin (CDDP) often causes nephrotoxicity by eliciting inflammation and oxidative stress. Golimumab, an anti-TNF biologic, is prescribed for the management of numerous inflammatory ailments like psoriatic and rheumatoid arthritis, ulcerative colitis and ankylosing spondylitis. OBJECTIVE: Current study has explored the effects of anti-TNF biologics golimumab on mice due to cisplatin-induced nephrotoxicity. METHOD: Renal toxicity was caused by administration of single cisplatin injection at 22 mg/kg by intraperitoneal (i/p) route. Golimumab (24 mg/kg, s.c.) was administered consecutively for 7 days. The parameters such as renal functions, oxidative stress, inflammation, and renal damage were evaluated on the 7th day of experiments. RESULTS: Cisplatin administration caused nephrotoxicity as shown by a significant elevation of various parameters viz; serum creatinine, neutrophil gelatinase-associated lipocalin (NGAL), urea nitrogen (BUN), and cystatin C. There was a significant rise in urinary clusterin, kidney injury molecule 1 (KIM-1), and ß-N-acetylglucosaminidase (NAG) concentrations in the animals treated with cisplatin. The markers of oxidative stress (malondialdehyde, reduced glutathione, and catalase), inflammation (IL-6, TNF-α, IL-10, IL-1ß, MCP-1, ICAM-1, and TGF-ß1), and apoptosis (caspase-3) were also altered in serum and/or kidneys of cisplatin animals. Further, cisplatin-caused histopathological changes in proximal tubular cells as observed in the H&E staining of renal tissue. Golimumab treatment reduced all markers of kidney injury and attenuated cell death. Golimumab significantly reduced inflammatory cytokines TNFα, IL- 6, MCP-1, IL- 1ß, ICAM-1, and TGF-ß1 and increased anti-inflammatory cytokine IL-10 in cisplatin-intoxicated mice. CONCLUSION: The study's results suggest that golimumab prevented nephrotoxicity induced by cisplatin- through inhibition of oxidative stress, apoptotic cell death inflammatory response, thus improving renal function.

Protective effects of recombinant human golimumab and pentoxifylline in nephrotoxicity induced by cisplatin.

In cisplatin-induced nephrotoxicity, the significant role of activation of inflammatory pathways has been reported earlier. Studies indicate elevated expression and activity of tumor necrosis factor-α (TNF-α) in both, serum and kidneys of cisplatin-treated animals. Golimumab, an anti-TNF biologic, has been approved for the management of many inflammatory conditions. This experiment was planned and executed to evaluate the impact of golimumab on renal function, inflammation in cisplatin-induced nephrotoxicity in mice, and oxidative stress. Cisplatin (22 mg/kg) as a single intraperitoneal injection was used to induce nephrotoxicity in mice. Golimumab was administered at 24 mg/kg for 7 days by subcutaneous route. Pentoxifylline (PTX) was administered for 7 days (150 mg/kg) as a reference standard. Renal functions, oxidative stress, and inflammation were evaluated on the seventh day. Cisplatin administration in mice caused a significant rise in serum cystatin C, creatinine, blood urea nitrogen, and neutrophil gelatinase-associated lipocalin. A significant rise of urinary clusterin, kidney injury molecule-1, and ß-N-acetylglucosaminidase levels was also seen in cisplatin-treated animals. Furthermore, cisplatin-induced nephrotoxicity was associated with a significant increase in oxidative stress and inflammation in serum and kidneys. Golimumab treatment significantly prevented the cisplatin-induced alteration in markers of renal function and renal damage. There was a significant reduction in oxidative stress and inflammation in golimumab-treated animals. Furthermore, the histopathological evaluation also revealed inverted changes in the proximal tubules after golimumab treatment in kidneys after cisplatin toxicity. The standard drug, PTX, also prevented nephrotoxicity caused by cisplatin as indicated by the recovery in renal function, reduction in oxidative stress and inflammation. These results indicate that golimumab was effective in nephrotoxicity induced by cisplatin through inhibition of oxidative stress, and inflammatory response.

Monoclonal antibody purification and its progression to commercial scale.

With the advancements in upstream technologies, the capacity for monoclonal antibody (mAb) production has transformed from a few milligrams to grams per liter. These titers lead to enormous pressure on downstream processes (DSPs), which need to be reworked to achieve higher efficiency and better utilization of available resources. Various parameters, such as column sizing, aggregate removal, filtration and volume handling, must be considered while designing a facility for commercial scale. If any of these critical parameters are not defined during the facility design stage, collapse of the process can result, further resulting in commercial loss and delaying entry of the product into the market. Therefore, during the facility design stage, the process requirements, space utilization, process efficiency and advanced manufacturing systems must be evaluated appropriately before implementation on a commercial scale.

Nanomedicine: Bioavailability, Biotransformation and Biokinetics.

BACKGROUND: Nanomedicine is increasingly used to treat various ailments. Biocompatibility of nanomedicine is primarily governed by its properties such as bioavailability, biotransformation and biokinetics. One of the major advantages of nanomedicine is enhanced bioavailability of drugs. Biotransformation of nanomedicine is important to understand the pharmacological effects of nanomedicine. Biokinetics includes both pharmacokinetics and toxicokinetics of nanomedicine. Physicochemical parameters of nanomaterials have extensive influence on bioavailability, biotransformation and biokinetics of nanomedicine. METHODS: We carried out a structured peer-reviewed research literature survey and analysis using bibliographic databases. RESULTS: Eighty papers were included in the review. Papers dealing with bioavailability, biotransformation and biokinetics of nanomedicine are found and reviewed. Bioavailability and biotransformation along with biokinetics are three major factors that determine the biological fate of nanomedicine. Extensive research work has been done for drugs of micron size but studies on nanomedicine are scarce. Therefore, more emphasis in this review is given on the bioavailability and biotransformation of nanomedicine along with biokinetics. CONCLUSION: Bioavailability results based on various nanomedicine are summarized in the present work. Biotransformation of nanodrugs as well as nanoformulations is also the focus of this article. Both in vitro and in vivo biotransformation studies on nanodrugs and its excipients are necessary to know the effect of metabolites formed. Biokinetics of nanomedicine is captured in details that are complimentary to bioavailability and biotransformation. Nanomedicine has the potential to be developed as a personalized medicine once its physicochemical properties and its effect on biological system are well understood.

Pharmacokinetics, Metabolism, Distribution and Permeability of Nanomedicine.

BACKGROUND: Medical application of nanotechnology is termed as Nanomedicine and is widely used in healthcare industries. Nanotechnology has helped Physicians, Scientists and Technologists to understand the changes in cellular levels to develop nanomedicines and address the challenges faced by the healthcare sectors. Nanoparticles with less than 1nm in size have been used as drug delivery and gene delivery systems to accelerate the drug action in humans. Size of nanomaterials is akin to that of biomolecules and expected to have better interactions. Hence, its utility for various biomedical applications is explored. OBJECTIVE: Pharmacokinetics, metabolism, permeability, distribution and elimination studies of nanoparticles are essential to understand its potency, toxicity threshold and confirm its safe use in humans. Reports were available for toxicity studies on nanoparticles, but work on metabolism, pharmacokinetics, distribution and permeability of nanomedicine is limited. Hence, the main focus of this review article is about metabolism, pharmacokinetics, permeability and biodistribution of nanomaterials used in nanomedicine. CONCLUSION: Nanomedicine is increasingly becoming important in the treatment of diseases and diagnosis. Size of the particle plays an important role. As the particle size decreases its effect to cure the disease increases. Pharmacokinetics, bioavailability, half-life, metabolism, biodistribution and permeability of nanomedicine were found to be better than that of microsized drugs. In vitro and In vivo ADME (Absorption, Distribution, Metabolism and Excretion) studies are mandatory for pharmaceutical organic drugs. Similarly, nanomaterials should be subjected to both in vitro and in vivo ADME studies. Thus, nanomedicine can assist in the development of safe personalized medicine in humans.

Technologies and strategies to characterize and quantitate metabolites in drug discovery and development.

Pharmacokinetics and toxicokinetics studies rely on the quantitation of drugs and its metabolites. Drug metabolism studies are based on the characterization of the structure of drugs and their metabolites. Liquid chromatography coupled with mass spectrometry is an undisputed technology for the routine analysis of drugs and metabolites in the drug discovery set-up. Advancements in liquid chromatography and mass spectrometry have accelerated and improved drug discovery and development. Identification and quantitation of drugs and metabolites have gained importance because of their relevance to clinical research. In the present review, we present a bird's eye view of qualitative and quantitative analysis of drugs and their metabolites. Various technologies available to characterize and quantitate metabolites are analyzed. Characterization of metabolites of the sulfonyl urea drug glyburide and the kinetics of glyburide metabolism are discussed in detail.

Metabolite characterization of anti-cancer agent gefitinib in human hepatocytes.

Intensive Biotransformation studies on Gefitinib could play a significant role in designing and synthesizing new drugs around the core structure of Gefitinib. These studies may be useful in developing an entirely new drug by blocking the metabolic spots in Gefitinib. Gefitinib (Iressa) was the first oral epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor. Gefitinib shows toxicity to cancer cells and has the capability to inhibit the growth of cancer cells. Gefitinib is considered as one of the selective EGFR inhibitors to be available in clinical practice. In 2003, FDA had approved Gefitinib for metastatic non-small-cell lung cancer therapy (NSCLC). However, it was observed that NSCLC Patients who responded to treatment developed resistance to Gefitinib. Hence, in the present study Gefitinib was incubated with hepatocytes to identify both phase-I and Phase-II metabolites. Identified Phase -I metabolites were due to oxidative defluorination, N-dealkylation and loss of morpholine ring. One of the phase-II metabolites identified i.e. the glutathione adduct suggests the need to modify the structure of the drug for higher potency and safety.

Metabolic profile of glyburide in human liver microsomes using LC-DAD-Q-TRAP-MS/MS.

The sulfonylurea urea drug glyburide (glibenclamide) is widely used for the treatment of diabetes milletus and gestational diabetes. In previous studies monohydroxylated metabolites were identified and characterized for glyburide in different species, but the metabolite owing to the loss of cyclohexyl ring was identified only in mouse. Glyburide upon incubation with hepatic microsomes resulted in 10 metabolites for human. The current study identifies new metabolites of glyburide along with the hydroxylated metabolites that were reported earlier. The newly identified drug metabolites are dihydroxylated metabolites, a metabolite owing to the loss of cyclohexyl ring and one owing to hydroxylation with dehydrogenation. Among the 10 identified metabolites, there were six monohydroxylated metabolites, one dihydroxylated metabolite, two metabolites owing to hydroxylation and dehydrogenation, and one metabolite owing to the loss of cyclohexyl ring. New metabolites of glyburide were identified and characterized using liquid chromatography-diode array detector-quadruple-ion trap-mass spectrometry/mass spectrometry (LC-DAD-Q-TRAP-MS/MS). An enhanced mass scan-enhanced product ion scan with information-dependent acquisition mode in a Q-TRAP-MS/MS system was used to characterize the metabolites. Liquid chromatography with diode array detection was used as a complimentary technique to confirm and identify the metabolites. Metabolites formed in higher amounts were detected in both diode array detection and mass spectrometry detection.

Split calibration curve: an approach to avoid repeat analysis of the samples exceeding ULOQ.

BACKGROUND: The current practice of using calibration curves with narrow concentration ranges during bioanalysis of new chemical entities has some limitations and is time consuming. In the present study we describe a split calibration curve approach, where sample dilution and repeat analysis can be avoided without compromising the quality and integrity of the data obtained. RESULTS: A split calibration curve approach is employed to determine the drug concentration in plasma samples with accuracy and precision over a wide dynamic range of approximately 0.6 to 15,000 ng/ml for dapsone and approximately 1 to 25,000 ng/ml for cyclophosphamide and glipizide. A wide dynamic range of concentrations for these three compounds was used in the current study to construct split calibration curves and was successfully validated for sample analysis in a single run. CONCLUSION: Using this method, repeat analysis of samples can be avoided. This is useful for the bioanalysis of toxicokinetic studies with wide dose ranges and studies where the sample volume is limited.

Glyburide metabolism by placentas of healthy and gestational diabetics.

The aim of this investigation was to determine the metabolism of glyburide (GL) by microsomes prepared from placentas obtained from uncomplicated pregnancies (UP), women with gestational diabetics (GD) on a diabetic diet, and those on a diet and GL. Term placentas were obtained from UP and GD. Crude microsomal fractions were prepared by differential centrifugation and stored at -80 degrees C. The activity of the microsomes in metabolizing glyburide to the trans-4-hydroxycyclohexyl glyburide (THCGL) and cis-3-hydroxycyclohexyl glyburide (CHCGL) was determined and quantified using high-performance liquid chromatography-mass spectrometer (HPLC-MS). The activity of the placental microsomes varied widely between individual placentas in each group. The median values (pmol.mg (-1) P.min (-1)) for the rates of THCGL formation were 0.34, 0.3, and 0.23 for placentas of UP, GD on diet, and GD on GL and a diet, respectively. The median values for CHCGL formation were 0.13 for UP, 0.11 for GD on a diet, and 0.10 (pmol.mg (-1) P.min (-1)) for GD on GL and a diet. A pool of individual microsomal fractions from each group was prepared and its activity revealed the following: greater formation of THCGL in the UP (0.36 +/- 0.10) than GD (0.22 +/- 0.03) ( P = 0.058 for GD on a diet, 0.04 for GD on GL). There was greater formation of CHCGL in UP (0.26 +/- 0.04) than GD (0.12 +/- 0.003) ( P < 0.006). There was no difference in GD on a diet and GD on GL plus diet. We concluded that the apparent differences in the formation of metabolites may be statistically significant, but it is unlikely to be of physiological importance, given the sample size and other experimental factors. Therefore, a more comprehensive investigation is underway.

Kinetics of glyburide metabolism by hepatic and placental microsomes of human and baboon.

Glyburide (glibenclamide) is under investigation for treatment of gestational diabetes. Two metabolites of glyburide have been previously identified in patients, namely, 4-trans-(M1) and 3-cis-(M2) hydroxycyclohexyl glyburide. Recently, the metabolism of glyburide by microsomes of liver and placenta from humans and baboons revealed the formation of four additional metabolites: 4-cis-(M2a), 3-trans-(M3), and 2-trans-(M4) hydroxycyclohexyl glyburide, and ethyl-hydroxy glyburide (M5). The aim of this investigation was to determine the kinetics for the metabolism of glyburide by cytochrome P450 (CYP) isozymes of human and baboon placental and hepatic microsomes. The metabolism of glyburide by microsomes from the four organs revealed saturation kinetics and apparent K(m) values between 4 and 12 microM. However, the rates for formation of the metabolites varied between organs and species. M1 was the major metabolite (36% of total), formed by human hepatic microsomes with V(max) of 80+/-13 pmol mg protein(-1)min(-1), and together with M2, accounted for only 51% of the total. M5 was the major metabolite (87%) formed by human placental microsomes with V(max) of 11 pmol mg protein(-1)min(-1). In baboon liver, M5 had the highest rate of formation (V(max) 135+/-32 pmol mg protein(-1)min(-1), 39% of total), and in its placenta, was M4 (V(max) 0.7+/-0.1 pmol mg protein(-1)min(-1), 65%). The activity of human and baboon hepatic microsomes in metabolizing glyburide was similar, but the activity of human and baboon placental microsomes was 7% and 0.3% of their respective hepatic microsomes. The data obtained suggest that more than 1 CYP isozyme is responsible for catalyzing the hydroxylation of glyburide.

Identification of glyburide metabolites formed by hepatic and placental microsomes of humans and baboons.

Glyburide (glibenclamide) is a second-generation sulfonylurea used for treatment of type-2 and gestational diabetes mellitus. To date, two glyburide metabolites have been identified in maternal urine: namely, 4-trans-hydroxycyclohexyl glyburide and 3-cis-hydroxycyclohexyl glyburide. The use of glyburide to treat gestational diabetes prompted us to investigate its metabolism by the placenta. The metabolism of glyburide by microsomal preparations from human and baboon placenta was compared with metabolism by their livers. The metabolites formed by the microsomes of the four tissues were identified by high-performance liquid chromatography-mass spectrometry using retention times, ion current (extracted at m/z 510), and selected-ion monitoring. The data obtained revealed the formation of six distinct hydroxylated derivatives of glyburide by each of the four microsomal preparations. However, the amounts of the six metabolites formed by the placentas were a fraction of that formed by the livers. Moreover, the relative quantities of each metabolite formed differed between species as well as between the two tissues. Also, the structure of the unidentified metabolites was determined by comparison with synthesized standards. These metabolites were identified as the 4-cis-hydroxycyclohexyl glyburide, 3-trans-hydroxycyclohexyl glyburide, and 2-trans-hydroxycyclohexyl glyburide. Therefore, one glyburide metabolite remains to be identified, but the data we obtained allowed us to suggest its structure.