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Differentiation of Candida species resides on biochemical reactions and morphological features. However, phenotypic variations within some species such as Candida albicans has led to confusion in taxonomical and clinical classification of Candida species. Recent developments in analytical instrumentation and statistical methods, have enabled Fourier Transform Infrared Spectroscopy (FTIR) to emerge as a rapid, accurate and inexpensive method to characterize microorganisms [1–3]. The aim of present study is to characterize 5 reference strains of Candida species (C. albicans, C. parapsilosis, C. knisei, C. glabrata and C. tropicalis) using attenuated total-reflectance (ATR)- FTIR spectroscopy, i.e., without any preparation of the yeast colonies. These results could in turn be used to construct a data base for comparison of clinical strains.

Fast and exact identification of a great number of microorganisms is becoming a serious challenge. Differentiation and identification of microorganisms is today mainly achieved by the use of a variety of distinct techniques based on morphological, serological aspects and an array of biochemical tests. Vibrational spectroscopic techniques (FTIR and Raman) can be alternative and useful methods in this field due to their rapidity, “fingerprinting” capabilities, and the molecular information that they can provide. FTIR spectroscopy has benefited from recent advances in instrumentation and data processing techniques and has consequently progressed a lot in this field. However, direct analysis of microorganisms from culture plates, without transfer of the biological material, is still difficult to achieve. If one could do measurements directly from the agar plates, there would be essentially no sample preparation, and if a spectroscopic technique could exhibit such characteristics as easy handling, rapid identification within 24 h when starting from a single colony, and high differentiating capacity, then such a technique can be more easily accepted as a complementary biochemical method in routine clinical laboratories. In this study, we have carried out some pilot studies on E. coli in order to go in this sense. Since Raman spectroscopy itself is inherently a weak phenomenon and suffers from low sensitivity and sample fluorescence, the best way to have access to the bacterial colony spectral information was to use surface-enhanced Raman scattering (SERS) technique. In our situation, the most straightforward manner of implementing this technique was the use of colloidal suspensions.

The wheat grain consists of the endosperm and the germ, surrounded by protective bran layer. During milling, the grain is disrupted and fractionated into several different fractions. The highest quality white flour should not contain any germ. The separation of wheat germ to yield white flour is not a fully efficient process. Therefore , it is important to measure the concentration of germ in millstreams in order to estimate the purity of white flour.

An encoding α-glucosidase gene from an archaebacteria Thermococcus hydrothermalis was cloned by functional complementation of TCY70 Saccharomyces cerevisiae mutant. The obtained strain (AGLSC) exhibited an intriguing characteristic with pink-coloured cells. The possible implication of the α-glucosidase on the yeast cell wall has been followed by transmission FTIR spectroscopy. The differences between TCY70 and AGLSC strains have allowed to evaluate the cell wall structural modifications.

Confocal multispectral imaging comprises the advantages of traditional spectroscopy (Raman or fluorescence) and confocal microscopy. It is based on microspectroscopy resulted from coupling a spectrometer with a confocal microscope [1] to obtain spatially resolved and structurally specific (particularly with Raman) information. High-precision scanning of a sample along the spatial dimensions provides multispectral matrix. The latter can be used to generate, using characteristic spectral parameters, a chemicalstructural map [2] of the sample. Among the most exciting biomedical applications of such non-invasive mapping one can cite the in situ investigations of living cells: to monitor molecular composition of the cells or to follow small Raman-active chromophores such as drugs, ion-markers, etc.

A lot of work has been carried out on the second transformation of Triticum aestivum wheat, concerning the baking properties of flour. Nevertheless, to date, the knowledge on the first transformation of grains during milling is still very poor. Our study aims at characterising spectroscopically [1–3] the molecules responsible of the grain cohesion in order to define the milling value of a wheat variety. This can lead to an optimisation of the milling by selecting more interesting wheat varieties or by applying specific treatment on commonly raised wheat. The milling value takes into account both the ease of fracturing the kernel of the grain [4, 5], and the separation of the envelope and the kernel. The high spatial resolution of Raman microspectroscopy permits to differentiate the components of the starchy endosperm or kernel, and of the aleurone cells layer forming the envelope — kernel interface. Confocal Raman microscopy (Labram Dilor, France) using a He/Ne laser has been performed on grain sections (50 um thick) of various Triticum aestivum samples of different hardnesses (Soft and Hard). Samples were supplied by INRA and Champagne Cereales.

Microspectrofluorometry which has already been a proven tool for studying the behaviour of drugs in living cells allows to record confocal fluorescence spectral images [1, 2, 3]. This technique has been developed to detect the phenolic components as a whole on the surface of transversal and longitudinal sections of wheat grain, on the basis of fluorescence emission spectra.

Schizosaccharomyces is a genus of fungi belonging to the family Saccharomycetaceae that contains the fission yeasts and are found in fermented beverages and fruit juices. It invades wine fermentation medium and is discarded after identification of Saccharomyces cerevisiae by biochemical and molecular biology (pulsed-field, PCR) techniques. However, these methods are quite time consuming. We propose in this study, by joint means of ATR-FTIR spectroscopy and chemometrics methods, to discriminate in a non-invasive manner, Saccharomyces cerevisiae (α FL 200, laboratory strain) from Schizosaccharomyces.

Previous studies demonstrated that there are some similarities of action between retinoids and carotenoids. The discovery of a family of nuclear retinoic acid receptors (RAR) with a high affinity for the ligand raised the possibility that the conversion of carotenoids to retinoids could activate the RAR [1]. It was also reported that carotenoids of saffron and especially their derivatives, crocins (CRCS), or pure derivatives, crocetin (CRT) and dimethylcrocetin (DMCRT), could be used as alternative antitumour agents in cancer chemotherapy because of their lower toxicity at high doses [2]. For DMCRT, it was shown that it induces maximum differentiation almost equally effective as ATRA at a concentration of 5 μM [3].

Retinoids are a class of compounds structurally related to vitamin A which have been shown to have marked activities, experimentally as well as clinically. The most common, retinoic acid (RA), exists naturally in vertebrates as two principal forms, all-trans (ATRA) and 9-cis (9C-RA), and affects the growth of normal malignant cells [1]. It is now well established that nuclear receptors are the ultimate targets of retinoids. Most of the biological effects of retinoids are mediated by two sub-families of receptors, namely retinoic acid receptors (RARs) and retinoic X receptors (RXRs), which form part of the steroid/thyroid hormone receptor superfamily of nuclear transcription factors [2]. RA modulates the transcription of a variety of genes associated with cellular proliferation and differentiation by binding RARs and RXRs. Three types of RARs (α, β, and γ), coded by three different genes, exist and are activated by both ATRA and 9C-RA. RXRs exist also in three forms (α, β, and γ), but are activated only by the 9-cis isoform. A recent 3D 2 A crystal structure of the ligand binding domain (LBD) of the human retinoic acid receptor RARγ bound to ATRA suggests an electrostatic guidance mechanism [3]. In fact, the ATRA molecule is driven in the ligand-binding pocket and is locked there by interactions with the predominant hydrophobic residues.

The characterisation of the TEM and SHV phenotypes (bacterials strains producing plasmid-mediated penicillinases) can be performed by isoelectric focusing, determination of substrate and inhibition profile, and by hybridization or nucleotide sequencing [1]. In this work, we propose a straightforward method, by using ATR-FTIR spectroscopy and chemometrics techniques, to discriminate the transconjugants TEM-2 and SHV-3 (resistant strains) from each other and from the suceptible strain (E. coli C600 K12).

During the last few years there has been a growing interest concerning both the instrumentation and application aspect of Fourier transform Raman spectroscopy (1,2). On the other hand, little effort have been guided towards micro FT-Raman analysis since the additionnal use of a microscope relatively reduces the sensitivity of this analytical tool (3, 4). In order to improve the sensitivity with aqueous solutions (at physiological concentration) we have combined this technique with surface-enhanced Raman scattering (SERS) effect, since the resonance Raman effect usually does not exist in this domain. In this report, An FT-Raman and FT-SERS microprobe with spatial resolution on the order of 8 µm has been developed. The whole set-up is validated with a model biological compound, crocetin, using silver and gold colloids and gold island films. We have compared the enhancement levels of different SERS active substrates and evaluated the influence of different SERS active substrates on molecular structure by comparaison of states isomerisation of free crocetin and when adsorbed on the metal surface.

Planar ring systems of antitumor drugs are often the chromophores with quite strong electronic transitions in visible or near-UV regions. Hence, previous optical spectroscopy approach to the investigation of the nature of the drug-target interactions was based on resonance Raman and surface-enhanced resonance Raman (SERRS) analyses (1–3). SERRS spectroscopy enabled to work at concentrations in the range of 10-6–10-9 M and to avoid interference from the intrinsic fluorescence of the drugs. Unfortunately, this approach is not always available for the investigation of interactions of side-chain groups (usually not exhibiting strong electronic transitions in visible) of the drugs within the supramolecular complexes. Obviously, FT-Raman spectroscopy with the off-resonance near-infrared excitation could enable analysis of drug side-chain groups interactions. The problem is that the technique has quite low sensitivity, i.e., not applicable for investigation at physiological (10-5–10-7 M) concentrations of drugs in aqueous solutions. Hence, the SERS in the near-infrared could be a promisng approach for studies of drug interactions within the supramolecular complexes.

Several among the most active anticancer drugs poison cellular DNA topoisomerase I (Top I). Camptothecins (CPTs, Fig.l) inhibit Top I selectively and are very active in the clinical trials that have been initiated since the early 90‘s. After showing that active DNA replication is required for converting cleavable complexes to DNA damage and cytotoxicity, Top I was purified from several cancer cell lines, camptothecin sequence selectivity in various DNA fragments including oligonucleotides was studied, and drug structure-activity studies based on the synthesis of different derivatives were performed. CPT seems to bind to a stereospecific receptor site on the Top I - DNA complexes.

FT-IR spectroscopy has been applied to monitor molecular structural changes induced by navelbine in the resistant J82-NVB cell line. Two different sampling methods have been used: cell smears (Le Gal et al, 1993) and cells contained in KBr discs. Infrared spectroscopy combined with high pressure had been already used to differenciate between normal and malignant cells (Rigas et al, 1992; Wong et al, 1993).

One of the aminoacridine derivatives, m-AMSA (4’-(9-acridinyl-amino)-methanesulfon-m-anisidide, Figure 1), is well known to exhibit strong antitumor activity [1]. This was correlated with the drug’s ability to trap the nuclear enzyme, DNA topoisomerase II (topo II) in the form of a “cleavable complex” on cellular DNA [2]. To study the molecular interactions of m-AMSA within living K562 cells, we have applied micro-SERS spectroscopy, as a sensitive and non-destructive technique.

The anticancer agent camptothecin (CPT) as well as several related derivatives from the CPT-family of drugs, are thought to control proliferation of cancer cells by stabilization of the reversible topoisomerase (top) I - drug - DNA (cleavable) ternary complex (1). The antitumor activity of CPT and related compounds deteriorate following dissolution of the compounds in aqueous media due to the presence of a hydrolyzable a-hydroxylactone ring moiety. Intracellular interactions exhibit strong influence on the lactone-carboxylate equilibrium of the drug. Lactone and carboxylate forms exhibit different interactions with human serum albumin, hence, the protein’s preferential binding of carboxylate results in the lactone ring opening more rapidly (2). On the other hand, lipid bilayer stabilize the lactone moiety of CPT-drugs (3).

Thin Layer Chromatography (TLC) is a very simple and useful separation technique that suffers from a lack of identity-indicating information. Although non specific, comparative procedures such as Rf (relative factor) values and spray reaction are widely used, but their application in the field of unknown TLC spot identification is somewhat limited. A better approach is the ability to have direct access to spectral informations from an unknown spot on the plate and thereafter using these to characterize the substance.

It has been known that retinoic acid and carotenoid derivatives induce differentiation of human promyelocytic leukemia cell line (HL60) but not of human erythroleukemia cell line (K562) (1, 2). This suggested that this response is due to a specific retinoid binding protein (SRBP) present in the HL60 cell nucleus but not in K562 cells, which leads to a ternary complex formation (Drug-SRBP-DNA) and subsequently, specific gene expression (3). However, retinoic acid toxicity remains an important limiting factor for its use at high therapeutical doses. Previous studies showed that all-trans retinoic acid (ATRA) induces a maximal differentiation almost equally effective as dimethyl crocetin (DMCRT) at a concentration of 5 µM (4). Several studies have suggested that natural carotenoids could be used as alternative antitumour agents in cancer chemotherapy because of their lower toxicity at high doses (5). Carotenoids have been used to probe the environment of some cell compartments in vitro and in living cells (6, 7). In this work, we propose to study the molecular interactions of DMCRT in HL60 and K562 cell lines using near-infrared Fourier transform Raman microspectroscopy.

Previous work has shown that confocal spectral imaging analysis is an effective probe of antitumor drug distribution in single living cell.1 However, spectral imaging analysis itself poses only nondirect assignment of spectral images to particular cellular components.

Set of flexible, solid and liquid substrates for trace organic analysis by surface-enhanced Raman spectroscopy (SERS), comprising a flexible or solid support treated by ions irradiation, chemically etched and coated by metal or prepared in the form of metal (silver, gold or platinum) hydrosols or suspension and “activated” by the addition of counterions has been developed.

Many DNA intercalators and non-intercalators have been shown to have antitumor activity.1Despite extensive effort in analogue synthesis, function of the drugs is not understood. No single known parameter correlated with drug cytotoxicity or antitumor activity.1 However, structural specificity of the drugs was clearly noted and the biological activity has been attributed to the formation of complexes between the chromophore framework and base pairs of DNA. The changes in the overall structures of the drug/target complexes amplify small chemical differences between antibiotics and provide a possible explanation for the differences in the clinical activity of the drugs.

Infrared spectroscopy is a non-destructive and rapid tool for characterization of structural features of biological molecules and complex materials like cells and bacteria (1,2). The spectra yield a wealth of information regarding the vibrational motions in the biological molecules. In this study, an application of FT-IR technique to intact bacteria is described. FT-IR spectroscopy has shown to be sensitive to the antibiotic-induced chemical changes in bacteria. The spectral changes between untreated bacteria and bacteria treated by drugs are discussed in termes of biological effects induced by antibiotics.

The authors have endeavoured to present the material in a style understandable to the average reader. However, by means of the refs.(1 to 7) the reader will easily gain access to the original special reports on Resonance Raman pratice and to reviews of biological applications published in recent years.

In the last decade, chemotherapy has been used alone or in association with surgery, radiotherapy and lately immunotherapy in cancer treatment. This method has increased continuously and led to a great improvement in the cure of tumours, in particular, when the tumour cells have been spread all over the body.