Structure and Medium-Dependent Photodecomposition of Fluoroquinolone Antibiotics

The photochemical reactivity of four fluoroquinolone antibiotics is examined. For norfloxacin (NOR), enoxacin (ENX) and lomefloxacin (LOM), the only process occurring is defluorination (from position 6 for the first two drugs, from position 8 for the last one). The quantum efficiency is both structure and medium dependent (4 close to 0.5 both in water and in 0.1 M phosphate buffer for LOM,- 0.01 for ENX and 0.004 for NOR in buffer, but more than an order of magnitude higher in neat water). Ofloxacin (OFL) is less light sensitive φ 0.001) and undergoes, in part, reactions different from defluorination. The photoreaction involves heterolytic C-F bond fragmentation and its efficiency is determined by the internal charge-transfer character of the excited state (increasing in the series OFL < NOR < ENX < LOM according to the electronegativity of the substituent in position 8) and by the stabilization of the resulting aryl cation (larger for the 8-cation than for the 6-cation). The relevance of these data for the rationalization of the known phototoxicity of these drugs is discussed.     

反相高效液相色谱法同时测定四种氟喹诺酮类药物

在ＳＨＩＭ－ＰＡＣＫ ＣＬＣ－ＯＤＳ柱上,以甲醇与０．０５ｍｏｌ／Ｌ柠檬酸＋０．０１ｍｏｌ／Ｌ醋酸铵（ＰＨ４．５）（Ｖ甲醇：Ｖ柠檬酸＋醋酸铵＝２５：７５）的混合溶液为流动相,要用反相高效相色谱法分离了测定了四种氟喹诺酮类药物;环丙沙星、氧氟沙星、诺氟沙星、依诺沙星,线必不丙沙星和氧氟沙星１０～１００μｇ／ｍＬ,诺氟沙星和依诺沙星２～８０μｇ／ｍｌ,相关系数ｒ〉０．９９９５,检出限分别为环丙沙星和依     

Assessment of DNA Damage Induced by Broadband and Narrowband UVB in Cultured Lymphoblasts and Keratinocytes Using the Comet Assay

Phototherapy with broadband UVB is an effective treatment for inflammatory dermatoses. A newly developed fluorescent UVB lamp (Philips TL01) that emits a narrowband UVB around 311 nm was shown to be superior for the phototherapy of psoriasis. In order to contribute to the knowledge about the carcinogenic potential of this UVB source, we measured the DNA damage in lymphoblasts and keratinocytes induced by narrowband UVB and compared it with that by conventional broadband UVB using the single cell gel electrophoresis (comet as-say). At equal doses, broadband UVB produced more DNA damage than narrowband UVB. However, in phototherapy of psoriasis, up to 10-fold higher doses are used with TLO1. When therapeutically equivalent doses were compared (10-fold correction for narrowband UVB), we found only slight differences in the amount of DNA damage produced by broadband and narrowband UVB. This supports the already existing evidence that for phototherapy narrowband UVB is not more carcinogenic than broadband UVB.     

Detection and Prevention of Ocular Phototoxicity of Ciprofloxacin and Other Fluoroquinolone Antibiotics†

Fluoroquinolone (FLQ) drugs are a potent family of antibiotics used to treat infections including ocular infections. To determine if these antibiotics may be phototoxic to the eye, we exposed human lens epithelial cells to 0.125–1 mm FLQs (ciprofloxacin [Cipro], lomefloxacin [Lome], norfloxacin [Nor] and ofloxacin [Ofl]), the precursor quinolone nalidixic acid (Nalid) and UVA radiation (2.5 J cm⁻²). Based on fluorescence confocal microscopy, FLQs are diffused throughout the cytoplasm and preferentially located in the lysosomes of lens epithelial cells. Neither FLQ exposure alone nor UVA exposure alone reduced cell viability. However, with exposure to UVA radiation the FLQs studied (Cipro, Nor, Lome and Ofl) induced a phototoxic reaction that included necrosis, apoptosis, loss of cell viability as measured by MTS, and membrane damage as determined by the lactate dehydrogenase assay. Both Nalid and all FLQs studied (Cipro, Nor, Lome and Ofl) photopolymerized the lens protein α-crystallin. Phototoxic damage to lens epithelial cells and/or α-crystallin will lead to a loss of transparency of the human lens. However, if precautions are taken to filter all UV radiation from the eye while taking these antibiotics, eye damage may be prevented.     

Separation of Fluoroquinolone Antibiotics by TLC on Silica Gel,Cellulose and Silanized Layers

JPC – Journal of Planar Chromatography – Modern TLC - Separation of the fluoroquinolone antibiotics has been examined using numerous mobile phases and commercially available TLC plates...     

固相萃取-高效液相色谱-串联质谱法同时测定土壤中氟喹诺酮、四环素和磺胺类抗生素

建立了固相萃取-高效液相色谱-串联质谱法同时检测土壤中氟喹诺酮类、四环素类和磺胺类18种抗生素的分析方法。土样经含50%乙腈的磷酸盐缓冲液(pH=3)提取后,以SAX-HLB串联小柱净化富集,在HPLC/MS/MS多反应监测模式下进行定性及定量分析。添加浓度为200和50μg/kg时,土壤中氟喹诺酮类、四环素类、磺胺类的加标回收率分别为67.2%～89.0%,62.2%～85.4%和55.8%～97.4%;其相对标准偏差为1.1%～17.2%。以3倍信噪比估算出氟喹诺酮类、四环素类、磺胺类的检出限分别为3.4～8.9μg/kg,0.56～0.91μg/kg和0.07～1.85μg/kg。应用此方法检测6种不同类型土壤样品,结果表明,污灌区土壤中检出有抗生素,浓度为1.72～119.6μg/kg。     

Surface‐Imprinted Nanofilaments for Europium‐Amplified Luminescent Detection of Fluoroquinolone Antibiotics

The development and characterization of novel, molecularly imprinted polymer nanofilament‐based optical sensors for the analysis of enrofloxacin, an antibiotic widely used for human and veterinary applications, is reported. The polymers were prepared by nanomolding in porous alumina by using enrofloxacin as the template. The antibiotic was covalently immobilized on to the pore walls of the alumina by using different spacers, and the prepolymerization mixture was cast in the pores and the polymer synthesized anchored onto a glass support through UV polymerization. Various parameters affecting polymer selectivity were evaluated to achieve optimal recognition, namely, the spacer arm length and the binding solvent. The results of morphological characterization, binding kinetics, and selectivity of the optimized polymer material for ENR and its derivatives are reported. For sensing purposes, the nanofilaments were incubated in solutions of the target molecule in acetonitrile/HEPES buffer (100 mM , pH 7.5, 50:50, v/v) for 20 min followed by incubation in a 10 mM solution of europium(III) ions to generate a europium(III)–enrofloxacin complex on the polymer surface. The detection event was based on the luminescence of the rare‐earth ion (λ_exc=340 nm; λ_em=612 nm) that results from energy transfer from the antibiotic excited state to the metal‐ion emitting excited state. The limit of detection of the enrofloxacin antibiotic was found to be 0.58 μM .     

Genotoxicity and Cytotoxicity of 4-Nonylphenol Ethoxylate on Lymphocytes as Assessed by the Comet Assay

Surfactants, which are prevalent at industrial sites and in the environment generally, are potential risk factors in human carcinogenesis. The widespread industrial use of surfactants such as 4-alkylphenol ethoxylates and their prevalence in many cleaning products have provoked studies about surfactant concentrations in water and their toxicity levels. Up to now, these substances have mainly been tested on aquatic organisms. Though tests on human cell lines are rare. The alkaline Comet assay was performed to evaluate the genotoxicity of 4-nonylphenol ethoxylate, a biodegradable product of 4-alkylphenol ethoxylate, in human lymphocytes. Concentrations tested ranged from 0.15 to 150 µg/mL. Test concentrations of 10 to 15 µg/mL caused an increase level of DNA migration in human cells, but without inducing excessive toxicity (viability > 80%). Though induced levels of DNA migration starting at concentrations of 30 µg/mL may have been due to excessive levels of cytotoxicity (viability < 70%). Based on these data, 4-nonylphenol ethoxylate can induce DNA damage in human lymphocytes but at higher concentrations than are normally found in river or drinking water. However, considering the prevalence of surfactants, the measured genotoxicity of these substances is of concern. Further investigations on human target cells are necessary to evaluate the carcinogenic impact of surfactants and reconsider their environmental acceptance.     

适体荧光分析法检测养殖废水中四环素类抗生素

建立了核酸适体识别-荧光探针技术检测养殖废水中土霉素、四环素、金霉素及强力霉素4种四环素类抗生素(TCs)总残留量的新方法。两段DNA对TCs共同识别后折叠成稳定的"发卡型"双链结构,核酸染料4'-6-二脒基-2-苯基吲哚(DAPI)能插入"发卡"部位产生荧光信号发射,据此可实现对TCs的定量检测。在最优实验条件下,以土霉素为目标分子建立工作曲线,在10~50 nmol/L范围内,荧光强度与对应浓度呈良好线性关系,土霉素的检出限为2.3 nmol/L。用四环素、土霉素、金霉素和强力霉素分别进行加标回收实验,平均回收率为88.5%~102.3%,相对标准偏差(n=5)为3.2%~6.7%。5个养殖场水样中均有TCs检出,检出量在7.6~42.7 nmol/L之间。该方法简单、灵敏、快速,可满足养殖场废水中TCs总残留量的测定要求。     

氟喹诺酮类抗生素与磷钨酸体系的共振瑞利散射光谱研究及其分析应用

在5.0 mol/L的HCl缓冲介质中,磷钨酸（Pwa）与莫西沙星(MXFX)和加替沙星（GTF）等氟喹诺酮类抗生素(FLQs)相互作用形成摩尔比1∶1离子缔合物,导致体系的共振瑞利散射(RRS)显著增强并出现新的RRS光谱。 MXFX和GTF的反应产物具有相似的光谱特征,最大散射波长位于320 nm附近,且药物浓度与散射增强(ΔI)成正比,2种氟喹诺酮类药物的线性范围分别为0.025~6.0 mg/L（MXFX）和0.023~9.0 mg/L（GTF）。 据此可建立用于测定氟喹诺酮类药物的简捷快速灵敏的新方法,方法用于胶囊和人尿液中的FLQs测定并取得满意结果。 并对反应机理和RRS增强的原因进行了讨论。     

Lysosomes Are Sites of Fluoroquinolone Photosensitization in Human Skin Fibroblasts: A Microspectrofluorometric Approach*

The fluoroquinolone antibiotics are widely used despite their strong phototoxicity under solar UV irradiation. Although they are known as good photodynamic photosensitizers, other factors than production of activated oxygen species may play a role in the effectiveness of the phototoxic effect. Subcellular localization is one of the important parameters that may determine this strength. Using microspectrofluorometry, it is shown that norfloxacin, ofloxacin, lomefloxacin, ciproflaxin and BAYy3118 are readily incorporated into lysosomes of HS68 human skin fibroblasts although weak staining of the whole cytoplasm also occurs especially with norfloxacin. Consistent with their photoinstability in solutions, the fluoroquinolones under study are readily photobleached by UVA in the HS68 fibroblasts. The BAYy3118 derivative that has the fastest bleaching rate also shows the strongest phototoxicity toward HS68 fibroblasts. Photosensitization with these fluoroquinolones induces lysosomal membrane damage as shown by the increased rate of leakage of the lysosomal probe lucifer yellow as compared to that observed with untreated cells.     

Isopimpinellin Is Not Phototoxic in a Chick Skin Assay

Abstract— Synthetic isopimpinellin (5,8-dimethoxypsoralen), confirmed to contain as impurities only trace quantities at most of psoralen, bergapten (5-methoxypsoralen) and xanthotoxin (8-methoxypsoralen), is not phototoxic when tested in a chick skin bioassay system. These findings are at variance with earlier studies showing isopimpinellin to be phototoxic against chick skin and support the conclusion that isopimpinellin is photobiologically inactive. As recently proposed by others, the several reports of isopimpinellin photoactivity are most likely attributable to contamination by small amounts of highly active psoralens such as bergapten or xanthotoxin.     

Wound Healing Assay Using Microfluidic Channels

An approach was described to produce a wound area in vitro using laminar flow technique to selectively remove cells in microfluidic channels. Cell migration which plays an important role in the process of wound healing can be observed dynamically by this method. In order to prove our system, we have studied its properties by injecting NIH-3T3 cells into main channel and added several reagents to observe the mediatory influence. The results reveal that fibroblast growth factor-2(FGF-2), insulin-like growth factor 1(IGF-I), platelet-derived growth factor BB(PDGF-BB) and ascorbic acid(Vc) groups can promote cell migration compared with the control group. But the average migration distance was diminished in dexamethasone group. It can be concluded that this system can be used to analyze the process of wound healing.     

Spectrophotometric Assay of Certain Aminoglycosides Using Cyanoacetamide

Abstract

A simple and sensitive ultraviolet spectrophotometric method has been devised for the determination of amikacin, kanamycin, neomycin and streptomycin in pure form and in some pharmaceutical preparations. The method depends on the nitrosation of the primary amino groups followed by reaction with cyanoacetamide in ammonia medium at 100°C for 30 min, The reaction products exhibit a characteristic absorption. maximum at 270 nm. Beer's law is obeyed within the concentration ranges 4–40 μg ml^?1 for amikacin, kanamycin and neomycin and 8–80 μg ml^?1 for streptomycin with apparent molar absorptivities of about 1.85 × 10⁴ for the first three compounds and 1.3×10⁴ 1. mole^?1.cm^?1 for streptomycin.     

Determination of Toxins Using Enzyme-Amplified Receptor Assay

Abstract

A new receptor based assay is described for the determination of toxins which have high affinities for the acetylcholine receptor. The method is based upon the hindrance of the normal binding of a synthetic enzyme-drug conjugate with a high affinity for the acetylcholine receptor protein by the presence of toxins acting as antagonists. The activity of the enzyme marker system, glucose-6-phosphate dehydrogenase covalently conjugated to desipramine, is monitored by colorimetric detection of the rate for NADH formation at 340 nm. The procedure proposed is designed to provide a simple toxin screen which can be done in a minimally equipped laboratory while achieving the required sensitivity. The technique is illustrated for snake venoms from Bungarus multicintus, Naja naja, and the alkaloid tubocurarine. Aspecific binding responses are shown to have minimal effect on the assay.     

Fluorometric Assay of Alcohols Using Alcohol Oxidase

Abstract

Fluorometric methods are described for the assay of ethanol, methanol, allyl alcohol and n-propanol using the enzyme alcohol oxidase. The enzyme catalyzes the oxidation of the alcohols to aldehydes with concomitant formation of hydrogen peroxide. The hydrogen peroxide formed is measured fluorometrically using p-hydroxy-phenylacetic acid and peroxidase. From 0.1–100 μg/ml of the alcohols and from 0.0002 to 0.050 unit/ml of alcohol oxidase can be assayed in 2 minutes with a precision of about ± 2%.     

Photogenotoxicity of mammalian cells: a review of the different assays for in vitro testing

During the past several years, phototoxicity has been studied at the molecular level, and these studies have provided new insights in the field of DNA lesion characterization, DNA repair and cell response to ultraviolet (UV)-induced stress. The development of new antibiotics and antiinflammatory drugs has highlighted the necessity to develop the assessment of phototoxicity in the safety evaluation of new chemical compounds. This paper aims at reviewing the known molecular mechanisms of the cellular response to UV-induced stress, the in vitro methods that can be proposed and used to screen for toxicity of sunlight and the photosensitization process resulting from the activation of drugs by light. UV sources, biological systems and endpoints of interest in that particular objective are listed. Phototoxic effects span from the cytotoxic-apoptotic effect to the induction of primary DNA damage, DNA repair and a variety of stress genes acting on the cell cycle and the fate of the cell. Ultimately, it can lead to the induction of hereditary DNA modification. A variety of assays are proposed to specifically address all these particular consequences of UV-induced toxicity.     

Spectrophotometric Determination of Selected Antibiotics Using Prussian Blue Reaction

A simple, sensitive and accurate spectrophotometric method has been described for the determination of ampicillin(I), amoxicillin trihydrate(II) and cefazolin sodium(III). The procedure is based on the formation of Prussian Blue (PB) complex. The reaction between the acidic hydrolysis products of antibiotics (T = 60 °C) with mixture of Fe³⁺ and hexacyanoferrate(III) ions was evaluated for the spectrophotometric determination of the mentioned drugs. The maximum absorbance of the colored complex occurs at λ = 700 nm and the molar absorptivity is 3.0 × 10⁴ 1 mol^?1cm^?1. The effect of various parameters such as concentration of K₃Fe(CN)₆ and Fe³⁺, nature and amount of acids used, temperature and time of heating were investigated. Under optimum conditions the linear range of calibration graph was 2.0–12.0, 5.0–13.5 and 3.0–12.0 μg mL^?1 for ampicillin, amoxicillin and cefazolin, respectively. The relative standard deviation for the determination of 10 μg mL^?1 of antibiotics was about 0.5–1.5%. The proposed method was successfully applied to the determination of selected antibiotics from pharmaceutical preparations. The validity of the method was tested by the official methods and by the recovery studies of standard addition to pharmaceuticals.     

Spectrophotometric Assay of Certain Aminoglycosoide Antibiotic Using Chloranil

Abstract

Chloranil (2,3,5,6-teterachloro-p-benzoquinone) reacts with certain aminoglycosoide antibiotics in borate buffer at pH 9.0 to give charge-transfer complexes. The complexes possese an absorption maxmum at about 350 nm with molar absorptivities of 3.78 × 10⁵, 4.69 × 10⁵, 5.0 × 10⁵ and 7.08 × 10⁵ for kanamycin, neomycin B, amikacin and tobramycin reepectively, These high intensity of the chargetransfor band and confirmity to beer's law enabled assay of these antibiotics with appreciable sensitivity and good precision.

Aminoglycosoide antibiotics have a broad spectrum of activity against both gram-negative and gram-positive bacteria.

The official compendia^1,2 recommend the microbiological assay for their evalution. The high experimental errors in microbiological estimation heae been reported³. Numerous spectrophotometric, fluorometric and chromatographic techniques for their quantitation have been thoroughly reviewed^4,5.

Substituted quinones such as 2,5 dichloroquinone⁶, 7,7,8,8-tetracyanoqulnone dimethane⁷, flouranil⁸ are used as TT- acceptors with various electron donors to form charge-transfer(CT) complexes and radical ions^9,10. Some aromatic amines^11,12 and aminoacids have been determined with chloranil^13–15.

The present work describes the utility of chloranil for the spectrophotometric determination of kanamycin sulphate, amikacin sulphate tobramycin aulphate, and neorycin B sulphate (framycetine sulphate) either in bulk or in their reepective doaage forms.