Combined assessment of lysine and N-acetyl cadaverine levels assist as a potential biomarker of the smoker periodontitis.

Periodontitis is an inflammatory condition of supporting structures of teeth leading to attachment and bone loss. Cigarette smoking is the single most important and modifiable risk factor with 5 to 20-fold susceptibility for periodontal diseases. Reverse smoking is a peculiar habit of smoking where the lit end is kept inside the mouth, which is predominant in the northern coastal districts of Andhra Pradesh. Polyamines are biologically active amines involved in tissue regeneration and modulation of inflammation. The study aimed to evaluate polyamines and check their utility as a marker in detection of periodontitis among different groups. Total polyamine levels showed significant increase in reverse smokers with periodontitis when compared to the other groups. Qualitative analysis by thin layer chromatography showed three polyamine bands with varying intensity among the different groups. Mass spectrometric and NMR analyses of the three bands identified them as N1, N8-diacetyl spermidine, N-acetyl cadaverine and lysine. Most significantly elevated levels of lysine was observed in the smoker and reverse smoker periodontitis groups when compared to healthy and non-smoker periodontitis groups. The significantly elevated levels of N-acetyl cadaverine could be responsible for the more destruction of periodontium in the reverse smoker group. Antioxidant potential decreased significantly in different smoker periodontitis groups. The present study suggests that the quantitative analysis of salivary polyamines, lysine and N-acetyl cadaverine can aid as an easy noninvasive diagnostic method for assessing the periodontal status, especially in smokers.

Assessment the influence of salt and polyphosphate on protein oxidation and Nε-(carboxymethyl)lysine and Nε-(carboxyethyl)lysine formation in roasted beef patties.

The impact of NaCl and tripolyphosphate (TPP)/pyrophosphate (PP) on protein oxidation and Nε-(carboxymethyl)lysine (CML) and Nε-(carboxyethyl)lysine (CEL) formation in roasted beef patties was investigated. The content of CML and CEL in patties treated with salts was approximately 1.1-1.7 and 1.2-3.2 times higher than that of the control samples, respectively. An increase in salt content caused higher oxidation of tryptophan and protein carbonylation with a decrease in Schiff bases (P < 0.05) and a slight decrease in lipid oxidation (P < 0.05). Significant correlations (P < 0.05) between CML, CEL, and protein oxidation measurements was found. The higher salts content, causing less cooking loss and higher moisture content, significantly correlated (P < 0.05) with CML, CEL content, and protein oxidation of the patties. The increase in CML and CEL content and protein oxidation in roasted patties with salts might be related to the pro-oxidation of salts, and also partly due to the temperature changes caused by the water-holding capacity of salts.

Assessment of Statin Interactions With the Human NTCP Transporter Using a Novel Fluorescence Assay.

Sodium taurocholate cotransporting polypeptide (NTCP), which is highly expressed in the sinusoidal membrane of hepatocytes, maintains bile acid homeostasis and participates in the hepatic disposition of a variety of endogenous substances as well as xenobiotics. Manifested by the involvement of organic anion-transporting polypeptides 1B1 and 1B3 (OATP1B1 and OATP1B3) in the hepatic uptake of statin drugs, sinusoidal membrane transporters play an important role in the pharmacokinetics and pharmacodynamics of these agents. It has been speculated that NTCP may function as an alternative pathway for statin hepatic uptake, complementary to OATP1B1 and OATP1B3. In the current study, we produced stable NTCP-expressing human embryonic kidney 293 (HEK293) cells and developed a fluorescence-based assay using flow cytometry for measuring NTCP transport with chenodeoxycholyl-(Nε-7-nitrobenz-2-oxa-1,3-diazole)-lysine (CDCA-NBD) as the substrate. NTCP-mediated CDCA-NBD transport was time-dependent and exhibited typical Michaelis-Menten kinetics, with a Km of 6.12 µM. Compounds known to interact with NTCP, including chenodeoxycholic acid and taurocholic acid, displayed concentration-dependent inhibition of NTCP-mediated CDCA-NBD transport. We report here a systematic evaluation of the interaction between statins and the NTCP transporter. Utilizing this system, several statins were either found to inhibit NTCP-dependent transport or act as substrates. We find a good correlation between the reported lipophilicity of statins and their ability to inhibit NTCP. The objective was to develop a higher-throughput system to evaluate potential inhibitors such as the statins. The in vitro assays using CDCA-NBD as fluorescent substrate are convenient, rapid, and have utility in screening drug candidates for potential drug-NTCP interactions.

Free Pentosidine Assessment Based on Fluorescence Measurements in Spent Dialysate.

The aim of this study was to primarily explore the relationship between free pentosidine and the fluorescence properties of spent dialysate, and also to develop a model to assess the levels of free pentosidine in spent dialysate based on the fluorescence measurements. First, 40 patients (20 females and 20 males) were examined during 40 dialysis sessions. High-pressure liquid chromatography (HPLC) was used to measure the free pentosidine concentrations from the spent dialysate. The full fluorescence spectra of the spent dialysates were recorded and single- and multi-wavelength (MW) models were developed. The average free pentosidine concentrations in the spent dialysate measured by HPLC at the start and end of the dialysis session were (mean ± SD) 4.25 ± 3.11 and 0.94 ± 0.69 µg/L respectively. The removal ratios (RRs) between RR_lab and RR_MW were statistically similar (p > 0.2). The concentration of free pentosidine and the RR can therefore be estimated from the spent dialysate when utilising fluorescence measurements.

Quantitative and Structural Assessment of Histone Methyllysine Analogue Engagement by Cognate Binding Proteins Reveals Affinity Decrements Relative to Those of Native Counterparts.

Methyllysine analogues (MLAs), furnished by aminoethylation of engineered cysteine residues, are widely used surrogates of histone methyllysine and are considered to be effective proxies for studying these epigenetic marks in vitro. Here we report the first structure of a trimethyllysine MLA histone in complex with a protein binding partner, quantify the thermodynamic distinctions between MLAs and their native methyllysine counterparts, and demonstrate that these differences can compromise qualitative interpretations of binding at the nucleosome level. Quantitative measurements with two methyllysine binding protein modules reveal substantial affinity losses for the MLA peptides versus the corresponding native methyllysine species in both cases, although the thermodynamic underpinnings are distinct. MLA and methyllysine adopt distinct conformational geometries when in complex with the BPTF PHD finger, a well-established H3K4me3 binding partner. In this case, an ∼13-fold Kd difference at the peptide level translates to nucleosomal affinities for MLA analogues that fall outside of the detectable range in a pull-down format, whereas the methyllysine species installed by native chemical ligation demonstrates robust binding. Thus, despite their facile production and commercial availability, there is a significant caveat of potentially altered binding affinity when MLAs are used in place of native methyllysine residues.

Assessment of collagen quality associated with non-enzymatic cross-links in human bone using Fourier-transform infrared imaging.

Aging and many disease conditions, most notably diabetes, are associated with the accumulation of non-enzymatic cross-links in the bone matrix. The non-enzymatic cross-links, also known as advanced glycation end products (AGEs), occur at the collagen tissue level, where they are associated with reduced plasticity and increased fracture risk. In this study, Fourier-transform infrared (FTIR) imaging was used to detect spectroscopic changes associated with the formation of non-enzymatic cross-links in human bone collagen. Here, the non-enzymatic cross-link profile was investigated in one cohort with an in vitro ribose treatment as well as another cohort with an in vivo bisphosphonate treatment. With FTIR imaging, the two-dimensional (2D) spatial distribution of collagen quality associated with non-enzymatic cross-links was measured through the area ratio of the 1678/1692cm-1 subbands within the amide I peak, termed the non-enzymatic crosslink-ratio (NE-xLR). The NE-xLR increased by 35% in the ribation treatment group in comparison to controls (p<0.005), with interstitial bone tissue being more susceptible to the formation of non-enzymatic cross-links. Ultra high-performance liquid chromatography, fluorescence microscopy, and fluorometric assay confirm a correlation between the non-enzymatic cross-link content and the NE-xLR ratio in the control and ribated groups. High resolution FTIR imaging of the 2D bone microstructure revealed enhanced accumulation of non-enzymatic cross-links in bone regions with higher tissue age (i.e., interstitial bone). This non-enzymatic cross-link ratio (NE-xLR) enables researchers to study not only the overall content of AGEs in the bone but also its spatial distribution, which varies with skeletal aging and diabetes mellitus and provides an additional measure of bone's propensity to fracture.

Quantitative assessment of organ distribution of dietary protein-bound 13 C-labeled Nɛ -carboxymethyllysine after a chronic oral exposure in mice.

SCOPE: Nɛ -Carboxymethyl-lysine (CML) is a prominent advanced glycation end-product which is not only found in vivo but also in food. It is known that a percentage of the dietary CML (dCML) is absorbed into the circulation and only partly excreted in the urine. Several studies have tried to measure how much dCML remains in tissues. However obstacles to interpreting the data have been found. METHODS AND RESULTS: A new protocol which discriminates dCML from native CML (nCML) has been developed. Three CML isotopes with different mass-to-charge ratios were used: nCML Nε -carboxymethyl-L-lysine, dCML Nε -[13 C]carboxy[13 C]methyl-L-lysine and internal standard Nε -carboxymethyl-L-[4,4,5,5-2 H4 ]lysine. Wild-type (n = 7) and RAGE-/- (n = 8) mice were fed for 30 days with either a control, or a BSA-bound dCML-enriched diet. Organs were analyzed for nCML and dCML using liquid chromatography-tandem mass spectrometry. Mice exposed to dCML showed an accumulation in all tissues tested except fat. The rate of deposition was high (81-320 µgdCML /g dry matter) in kidneys, intestine, and lungs and low (<5 µg/g) in heart, muscle, and liver. This accumulation was not RAGE dependent. CONCLUSION: The kidney is not the only organ affected by the accumulation of dCML. Its high accumulation in other tissues and organs may also, however, have important physiological consequences.

Assessment of protein quality of soybean meal and 00-rapeseed meal toasted in the presence of lignosulfonate by amino acid digestibility in growing pigs and Maillard reaction products.

An experiment was conducted to determine protein quality in processed protein sources using the content of AA, -methylisourea (OMIU)-reactive Lys, Maillard reaction products (MRP), and cross-link products; the standardized ileal digestibility (SID) of CP and AA; and growth performance in growing pigs as criteria. Differences in protein quality were created by secondary toasting (at 95°C for 30 min) of soybean meal (SBM) and rapeseed meal (RSM) in the presence of lignosulfonate resulting in processed SBM (pSBM) and processed RSM (pRSM). The processing treatment was used as a model for overprocessed protein sources. Ten growing pigs were each fed 1 of the 4 diets containing SBM, pSBM, RSM, or pRSM in each of 3 periods. Ileal chyme was collected at the end of each period and analyzed for CP, AA, and OMIU-reactive Lys. Diets were analyzed for furosine and carboxymethyllysine (CML) as an indicator for MRP and lysinoalanine (LAL), which is a cross-link product. The SBM and RSM diets contained furosine, CML, and LAL, indicating that the Maillard reaction and cross-linking had taken place in SBM and RSM, presumably during the oil extraction/desolventizing process. The amounts of furosine, CML, and LAL were elevated in pSBM and pRSM due to further processing. Processing resulted in a reduction in total and OMIU-reactive Lys contents and a decrease in G:F from 0.52 to 0.42 for SBM and 0.46 to 0.39 for RSM ( = 0.006), SID of CP from 83.9 to 71.6% for SBM and 74.9 to 64.6% for RSM ( < 0.001), and SID of AA ( < 0.001), with the largest effects for total and OMIU-reactive Lys. The effects of processing could be substantial and should be taken into account when using processed protein sources in diets for growing pigs. The extent of protein damage may be assessed by additional analyses of MRP and cross-link products.

Correlation between different markers for the assessment of red chilli pepper powders stability during shelf-life.

Pungency and red colour of Capsicum powders deteriorate during processing and storage, resulting in a decrease in market value. Two varieties of pepper with different pungencies were monitored for capsaicinoids, colour and furosine. Aliquots were stored at room and at low temperature during one year. At low temperature all indicators were stable in both varieties, while at room temperature, redness and capsacinoids decreased significantly, while furosine increased. High correlation was found between those markers. The more pungent variety exhibited higher stability in terms of all parameters. Differences observed suggest a potential protective effect exerted by capsaicinoids on powder stability. The decrease in capsaicinoids and redness accompanied by furosine increase showed a linkage between those markers never reported before. Considering that capsaicinoids and furosine occurrence have strong impact on the nutritional profile, the findings of this work show relevant changes in the nutritional value of chilli pepper powder after storage.

[The assessment of bone quality in lifestyle-related diseases].

Type 2 diabetes mellitus(DM)and other lifestyle-related diseases are associated with an increased risk of bone quality deterioration-type osteoporosis. The deterioration of bone quality in type 2 DM involves factors such as qualitative changes of collagens, reduction in bone turnover, narrow cortical bone diameter, increased cortical bone porosity, and destruction of trabecular bone microarchitecture. In mild to moderate chronic kidney disease and chronic obstructive pulmonary disease, the factors involved are thought to be hyperhomocysteinemia and deterioration of trabecular bone microarchitecture as well as cortical bone structure. Investigations of the usefulness of bone quality assessment using approaches such as the following are under way : biocheminal markers such as pentosidine and homocysteine, bone structure assessment methods such as hip structure analysis, trabecular bone score, and high-resolution peripheral quantitative computed tomography.

Rapid and scalable synthesis of innovative unnatural α,ß or γ-amino acids functionalized with tertiary amines on their side-chains.

We report a selective ruthenium catalyzed reduction of tertiary amides on the side chain of Fmoc-Gln-OtBu derivatives, leading to innovative unnatural α,ß or γ-amino acids functionalized with tertiary amines. Rapid and scalable, this process allowed us to build a library of basic unnatural amino acids at the gram-scale and directly usable for liquid- or solid-phase peptide synthesis. The diversity of available tertiary amines allows us to modulate the physicochemical properties of the resulting amino acids, such as basicity or hydrophobicity.

Assessment of the concentrations of various advanced glycation end-products in beverages and foods that are commonly consumed in Japan.

Dietary consumption has recently been identified as a major environmental source of pro-inflammatory advanced glycation end-products (AGEs) in humans. It is disputed whether dietary AGEs represent a risk to human health. Nε-(carboxymethyl)lysine (CML), a representative AGE compound found in food, has been suggested to make a significant contribution to circulating CML levels. However, recent studies have found that the dietary intake of AGEs is not associated with plasma CML concentrations. We have shown that the serum levels of glyceraldehyde-derived AGEs (Glycer-AGEs), but not hemoglobin A1c, glucose-derived AGEs (Glu-AGEs), or CML, could be used as biomarkers for predicting the progression of atherosclerosis and future cardiovascular events. We also detected the production/accumulation of Glycer-AGEs in normal rats administered Glu-AGE-rich beverages. Therefore, we assessed the concentrations of various AGEs in a total of 1,650 beverages and foods that are commonly consumed in Japan. The concentrations of four kinds of AGEs (Glu-AGEs, fructose-derived AGEs (Fru-AGEs), CML, and Glycer-AGEs) were measured with competitive enzyme-linked immunosorbent assays involving immunoaffinity-purified specific antibodies. The results of the latter assays indicated that Glu-AGEs and Fru-AGEs (especially Glu-AGEs), but not CML or Glycer-AGEs, are present at appreciable levels in beverages and foods that are commonly consumed by Japanese. Glu-AGEs, Fru-AGEs, CML, and Glycer-AGEs exhibited concentrations of ≥85%, 2-12%, <3%, and trace amounts in the examined beverages and ≥82%, 5-15%, <3%, and trace amounts in the tested foods, respectively. The results of the present study indicate that some lactic acid bacteria beverages, carbonated drinks, sugar-sweetened fruit drinks, sports drinks, mixed fruit juices, confectionery (snacks), dried fruits, cakes, cereals, and prepared foods contain markedly higher Glu-AGE levels than other classes of beverages and foods. We provide useful data on the concentrations of various AGEs, especially Glu-AGEs, in commonly consumed beverages and foods.

Design and evaluation of a novel trifluorinated imaging agent for assessment of bile acid transport using fluorine magnetic resonance imaging.

Previously, we developed a trifluorinated bile acid, CA-lys-TFA, with the objective of noninvasively assessing bile acid transport in vivo using (19) F magnetic resonance imaging (MRI). CA-lys-TFA was successfully imaged in the mouse gallbladder, but was susceptible to deconjugation in vitro by choloylglycine hydrolase (CGH), a bacterial bile acid deconjugating enzyme found in the terminal ileum and colon. The objective of the present study was to develop a novel trifluorinated bile acid resistant to deconjugation by CGH. CA-sar-TFMA was designed, synthesized, and tested for in vitro transport properties, stability, imaging properties, and its ability to differentially accumulate in the gallbladders of normal mice, compared with mice with known impaired bile acid transport (deficient in the apical sodium-dependent bile acid transporter, ASBT). CA-sar-TFMA was a potent inhibitor and substrate of ASBT and the Na(+) /taurocholate cotransporting polypeptide. Stability was favorable in all conditions tested, including the presence of CGH. CA-sar-TFMA was successfully imaged and accumulated at 16.1-fold higher concentrations in gallbladders from wild-type mice compared with those from Asbt-deficient mice. Our results support the potential of using MRI with CA-sar-TFMA as a noninvasive method to assess bile acid transport in vivo.

Quantitation of Maillard reaction products in commercially available pet foods.

During processing of pet food, the Maillard reaction occurs, which reduces the bioavailability of essential amino acids such as lysine and results in the formation of advanced Maillard reaction products (MRPs). The aim of this study was to quantitate MRPs (fructoselysine (FL), carboxymethyllysine (CML), hydroxymethylfurfural (HMF)) and the cross-link lysinoalanine (LAL) in commercial pet foods. Sixty-seven extruded, canned, and pelleted dog and cat foods for growth and maintenance were analyzed using UPLC-MS. Canned pet foods contained on average the most FL, CML, and HMF (4534, 37, and 1417 mg/kg dry matter, respectively) followed by pelleted and extruded foods. Average daily intake (mg/kg body weight(0.75)) of HMF is 122 times higher for dogs and 38 times higher for cats than average intake for adult humans. As commercial pet foods are most often the only source of food for dogs and cats, future research focus should be on the bioavailability and long-term health implications of MRP consumption by dogs and cats.

Assessment of heat treatment of various types of milk.

Raw milk (RM), reconstituted condensed milk (CM) and three types of reconstituted milk powders (SMPs) were heated indirectly at 80-140°C for 4 s. Native ß-lactoglobulin after 90°C treatment of RM was 1132±167 mg/L but no reliable quantities were estimated at temperatures >100°C, whereas 218±43 mg/L residual α-lactalbumin were found at 130°C. Average lactulose contents from 51 to 1549 mg/L were detected at ⩾100°C; average furosine was 1.9 and 126.5 mg/L in raw and 140°C treated milks respectively. The behaviour of heated CM was similar to that of heated RM except for higher furosine concentration. Reconstituted SMPs contained high quantities of lactulose and furosine, the ratio of which was lower than in similarly treated RM. Among the market milks analysed, the group of high-pasteurised milks was highly variable; i.e. native ß-lactoglobulin was 69-2831 mg/L, lactulose 0-824 mg/L and furosine 3.3-68.8 mg/L.

The Maillard reaction and pet food processing: effects on nutritive value and pet health.

The Maillard reaction, which can occur during heat processing of pet foods or ingredients, is known to reduce the bioavailability of essential amino acids such as lysine due to the formation of early and advanced Maillard reaction products (MRP) that are unavailable for utilisation by the body. Determination of the difference between total and reactive lysine by chemical methods provides an indication of the amount of early MRP present in foods, feeds and ingredients. Previous research reported that the difference between total and reactive lysine in pet foods can be up to 61.8%, and foods for growing dogs may be at risk of supplying less lysine than the animal may require. The endogenous analogues of advanced MRP, advanced glycation endproducts, have been associated with age-related diseases in humans, such as diabetes and impaired renal function. It is unknown to what extent advanced MRP are present in pet foods, and if dietary MRP can be associated with the development of diseases such as diabetes and impaired renal function in pet animals. Avoidance of ingredients with high levels of MRP and processing conditions known to favour the Maillard reaction may be useful strategies to prevent the formation of MRP in manufactured pet food. Future work should further focus on understanding the effects of ingredient choice and processing conditions on the formation of early and advanced MRP, and possible effects on animal health.

The balance of striatal feedback transmission is disrupted in a model of parkinsonism.

Inhibitory connections among striatal projection neurons (SPNs) called "feedback inhibition," have been proposed to endow the striatal microcircuit with computational capabilities, such as motor sequence selection, filtering, and the emergence of alternating network states. These properties are disrupted in models of Parkinsonism. However, the impact of feedback inhibition in the striatal network has remained under debate. Here, we test this inhibition at the microcircuit level. We used optical and electrophysiological recordings in mice and rats to demonstrate the action of striatal feedback transmission in normal and pathological conditions. Dynamic calcium imaging with single-cell resolution revealed the synchronous activation of a pool of identified SPNs by antidromic stimulation. Using bacterial artificial chromosome-transgenic mice, we demonstrate that the activated neuron pool equally possessed cells from the direct and indirect basal ganglia pathways. This pool inhibits itself because of its own GABA release when stimuli are frequent enough, demonstrating functional and significant inhibition. Blockade of GABAA receptors doubled the number of responsive neurons to the same stimulus, revealing a second postsynaptic neuron pool whose firing was being arrested by the first pool. Stronger connections arise from indirect SPNs. Dopamine deprivation impaired striatal feedback transmission disrupting the ability of a neuronal pool to arrest the firing of another neuronal pool. We demonstrate that feedback inhibition among SPNs is strong enough to control the firing of cell ensembles in the striatal microcircuit. However, to be effective, feedback inhibition should arise from synchronized pools of SPNs whose targets are other SPNs pools.

Radiosynthesis of 68Ga-labelled DOTA-biocytin (68Ga-r-BHD) and assessment of its pharmaceutical quality for clinical use.

OBJECTIVES: Biocytin analogues labelled with indium-111, yttrium-90 and lutetium-177 have shown their effectiveness in the imaging of infections/inflammation in patients with osteomyelitis and function as efficient tools in pretargeted antibody-guided radioimmunotherapy. In this study, the labelling of a biocytin analogue coupled with DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid), namely, r-BHD, with gallium-68 (68Ga) was optimized, and the quality and stability of the preparations were assessed for clinical use. MATERIALS AND METHODS: Synthesis of 68Ga-r-BHD was carried out by heating a fraction of the 68Ge/68Ga eluate in a reactor containing the biocytin analogue with the appropriate buffer. The influence of the precursor amount (from 2.5 to 140 nmol), the pH of the reaction (from 2 to 5.5) and the buffer species (1.5 mol/l sodium acetate, 1.5 mol/l sodium formate, 4.5 mol/l HEPES) on radiochemical yield and radiochemical purity was assessed. Studies on stability and binding to avidin (Av) were also conducted in different media. RESULTS: Under the best labelling condition (56 nmol of precursor, 3.8 pH, sodium formate buffer) synthesis of 68Ga-r-BHD resulted in a yield of 64 ± 3% (not decay corrected). Radiochemical purity was around 95% because a 68Ga-coordinated sulfoxide form of the ligand was detected as a by-product of the reaction (68Ga-r-SBHD). The by-product was identified and characterized by liquid chromatography-electrospray ionization tandem mass spectrometry. At the natural 1 : 4 Av/68Ga-r-BHD molar ratio, affinity results were 62 ± 2 and 80 ± 2% in saline and human serum, respectively. Stability of 68Ga-r-BHD and of the radiotracer/Av complex remains almost constant over 180 min. 68Ga-r-BHD appears to be a good candidate for clinical applications.

Assessment of free energy predictors for ligand binding to a methyllysine histone code reader.

Methyllysine histone code readers constitute a new promising group of potential drug targets. For instance, L3MBTL1, a malignant brain tumor (MBT) protein, selectively binds mono- and di-methyllysine epigenetic marks (KMe, KMe(2) ) that eventually results in the negative regulation of multiple genes through the E2F/Rb oncogenic pathway. There is a pressing need in potent and selective small-molecule probes that would enable further target validation and might become therapeutic leads. Such an endeavor would require efficient tools to assess the free energy of protein-ligand binding. However, due to an unparalleled function of the MBT binding pocket (i.e., selective binding to KMe/KMe(2) ) and because of its distinctive structure representing a small aromatic "cage," an accurate assessment of its binding affinity to a ligand appears to be a challenging task. Here, we report a comparative analysis of computationally affordable affinity predictors applied to a set of seven small-molecule ligands interacting with L3MBTL1. The analysis deals with novel ligands and targets, but applies widespread computational approaches and intuitive comparison metrics that makes this study compatible with and incremental to earlier large scale accounts on the efficiency of affinity predictors. Ultimately, this study has revealed three top performers, far ahead of the other techniques, including two scoring functions, PMF04 and PLP, along with a simulation-based method MM-PB/SA. We discuss why some methods may perform better than others on this target class, the limits of their application, as well as how the efficiency of the most CPU-demanding techniques could be optimized.

[Is bone biopsy necessary for the diagnosis of metabolic bone diseases? Non- invasive assessment of bone turn over markers could define the cause of metabolic bone diseases].

Recent advances of the measurement of bone turn over markers contribute to non-invasive assessment of bone-metabolic disorders. We can detect the cause of the metabolic disorders with bone turn over markers and hormonal profiles more easily than before. Today, we can diagnose and treat metabolic bone diseases without invasive procedure such as bone biopsy.