Exploring novel A2AAR antagonists: Design, synthesis, and evaluation of 2,6,9-trisubstituted purine derivatives as promising antifibrotic agents.

A series of 2,6,9-trisubstituted purine derivatives were designed and synthesized with diverse chemical moieties. Through a comprehensive biological evaluation, we identified 4-(6-(methylamino)-2-(phenylethynyl)-9H-purin-9-yl)phenol (6a) as a promising A2AAR antagonist with potent antifibrotic properties. Compound 6a demonstrated significant efficacy in inhibiting CRE promoter activity and in reducing the expression of fibrogenic marker proteins and downstream effectors of A2AAR activation, surpassing the A2AAR antagonist ZM241385 and initial screening hits, 9-benzyl-N-methyl-2-(phenylethynyl)-9H-purin-6-amine (5a) and 9-((benzyloxy)methyl)-N-methyl-2-(phenylethynyl)-9H-purin-6-amine (5j). Further validation revealed that compound 6a effectively inhibited fibrogenic marker proteins induced by A2AAR overexpression or TGF-ß1 treatment in hepatic stellate cells, alongside reducing PKA and CREB phosphorylation. These findings suggest that compound 6a exerts its antifibrotic action by modulating the cAMP/PKA/CREB pathway through A2AAR inhibition. Overall, our study provides valuable insights for the development of novel therapeutics that target hepatic fibrosis through A2AAR antagonism.

New Smoothened ligands based on the purine scaffold as potential agents for treating pancreatic cancer.

Aberrant activation of the Hedgehog (Hh) signalling pathway has been associated with the development and progression of pancreatic cancer. For this reason, blockade of Hh pathway by inhibitors targeting the G protein-coupled receptor Smoothened (SMO) has been considered as a therapeutic target for the treatment of this cancer. In our previous work, we obtained a new SMO ligand based on a purine scaffold (compound I), which showed interesting antitumor activity in several cancer cell lines. In this work, we report the design and synthesis of 17 new purine derivatives, some of which showed high cytotoxic effect on Mia-PaCa-2 (Hh-dependent pancreatic cancer cell lines) and low toxicity on non-neoplastic HEK-293 cells compared with gemcitabine, such as 8f, 8g and 8h (IC50 = 4.56, 4.11 and 3.08 µM, respectively). Two of these purines also showed their ability to bind to SMO through NanoBRET assays (pKi = 5.17 for 8f and 5.01 for 8h), with higher affinities to compound I (pKi = 1.51). In addition, docking studies provided insight the purine substitution pattern is related to the affinity on SMO. Finally, studies of Hh inhibition for selected purines, using a transcriptional functional assay based on luciferase activity in NIH3T3 Shh-Light II cells, demonstrated that 8g reduced GLI activity with a IC50 = 6.4 µM as well as diminished the expression of Hh target genes in two specific Hh-dependent cell models, Med1 cells and Ptch1-/- mouse embryonic fibroblasts. Therefore, our results provide a platform for the design of SMO ligands that could be potential selective cytotoxic agents for the treatment of pancreatic cancer.

Glutathione-Mediated Neuroprotective Effect of Purine Derivatives.

Numerous basic studies have reported on the neuroprotective properties of several purine derivatives such as caffeine and uric acid (UA). Epidemiological studies have also shown the inverse association of appropriate caffeine intake or serum urate levels with neurodegenerative diseases such as Alzheimer disease (AD) and Parkinson's disease (PD). The well-established neuroprotective mechanisms of caffeine and UA involve adenosine A2A receptor antagonism and antioxidant activity, respectively. Our recent study found that another purine derivative, paraxanthine, has neuroprotective effects similar to those of caffeine and UA. These purine derivatives can promote neuronal cysteine uptake through excitatory amino acid carrier protein 1 (EAAC1) to increase neuronal glutathione (GSH) levels in the brain. This review summarizes the GSH-mediated neuroprotective effects of purine derivatives. Considering the fact that GSH depletion is a manifestation in the brains of AD and PD patients, administration of purine derivatives may be a new therapeutic approach to prevent or delay the onset of these neurodegenerative diseases.

Nutrient intake, digestibility, and utilization in goats fed graded levels of hempseed cake finisher diets.

Globally, the price of soybean meal, the most common proteinaceous ingredient in livestock diets, has become highly expensive prompting a search for alternative ingredients. Hemp seed cake is a promising alternative but could be limited by its high neutral detergent fiber and ether extract contents which impede nutrient intake and digestibility. However, some ruminant species such as goats have superior ability to digest high fiber and ether extract diets. Thus, the current research evaluated nutrient intake and digestibility, rumen fermentation, and microbial protein synthesis of goats fed hempseed cake as a substitute for soybean meal in finisher diets. A total of 25 Kalahari Red castrates (27 ± 3 kg, 4-5 months old) were assigned to five dietary treatments (5 goats/ diet) in a completely randomized design. A maize-lucerne-based finishing diet was formulated with hempseed cake substituting soybean meal as the primary protein ingredient at 0, 25, 50, 75, or 100 g/kg dry matter. Ether extract intake exhibited a positive linear trend (P ≤ 0.05) while crude protein intake and microbial nitrogen supply exhibited a negative linear trend (P ≤ 0.05) with dietary inclusion of hempseed cake. However, feeding hempseed cake did not influence (P > 0.05) apparent nutrient digestibility, rumen fermentation parameters and nitrogen use efficiency. In conclusion, the substitution of soybean meal for hempseed cake decreased crude protein intake and microbial nitrogen supply in goat finisher diets without compromising nutrient digestibility and nitrogen use efficiency. The study recommends partial or full replacement of soybean meal with hempseed cake in goat finisher diets.

Brushite nanoparticles based electrochemical sensor for detection of uric acid, xanthine, hypoxanthine and caffeine.

The research in the field of biosensors has recently been focused on the design and development of functional electrode materials that can respond to changes in their biochemical environment. Here, we report the synthesis of dicalcium phosphate dihydrate (DCPD), also known as brushite (CaHPO4·2H2O) by soft chemical method and its application for electrochemical sensing of four different analytes. Phase purity, structure, chemical composition and surface morphology of the synthesized nanoparticles have been investigated using powder XRD, FTIR, SEM, XPS and HRTEM methods. Electrochemical sensor was prepared by modifying GCE with brushite and the modified electrodes were successfully used for either independent or simultaneous determination of uric acid, xanthine, hypoxanthine and caffeine in their mixture. The brushite/GCE exhibited four strong well-defined separate peaks corresponding to the oxidation of UA, XN, HXN and CF in phosphate buffer saline (PBS) at pH 7.4. The fabricated electrode showed low detection limits (S/N = 3) of 0.576, 1.0, 0.076 and 1.26 µM for UA, XN, HXN and CF respectively. Practical application of the fabricated electrode has been demonstrated by determining UA, XN, HXN and CF in human urine and coffee samples by direct method. The brushite offers scope for fabrication of sensor systems for implantable medical applications.

Milk feeding level and starter protein content: Effects on growth performance, blood metabolites, and urinary purine derivatives of Holstein dairy calves.

The objective of this study was to investigate the effects of milk allowances equal to 526 g/d as moderate (MOD) versus 790 g/d of milk dry matter as high (HI), and starter diets containing 18% or 23% crude protein (CP), on growth performance, blood metabolites, and purine derivative (PD) excretion in the urine of dairy calves. A total of 52 female Holstein dairy calves (40.8 kg of body weight) were randomly assigned to the experimental diets. The treatments were (1) moderate milk and 18% CP starter diet (MOD-18CP); (2) MOD and 23% CP starter diet (MOD-23CP); (3) high milk and 18% CP starter diet (HI-18CP); and (4) HI and 23% CP starter diet (HI-23CP). Calves had free access to a starter feed and water and were weaned on d 53 but remained in the study until d 73. Urine samples were collected during the preweaning period (for 6 consecutive days between d 35 and 40) and postweaning period (for 6 consecutive days between d 65 and 70) to investigate urinary excretion of PD. Starter feed intake, ß-hydroxybutyrate (BHB), and blood urea concentrations were reduced; however, average daily gain (ADG) and blood glucose levels increased in calves fed HI before weaning compared with MOD. During the preweaning period, high milk feeding increased total urinary PD excretion but decreased it after weaning. The 23CP diet resulted in higher feed intake and ADG before weaning and higher excretion of allantoin and total excretion of PD compared with the 18CP diet. The HI-23CP treatment resulted in the greatest withers and hip heights at weaning and final measurement, as well as the highest preweaning blood insulin concentrations. In terms of rumen development, MOD-23CP showed the greatest benefits based on starter intake, blood BHB concentration, and urinary excretion of PD. Based on the higher urinary excretion of PD found in HI-fed calves before weaning, it is possible that milk feeding overestimates estimated microbial yield. The results suggest that feeding starters with a higher proportion of CP may help maintain a more balanced ratio of CP to ME during high milk feeding, to avoid protein deficiency due to low starter intake. When calves are fed a high milk allowance, urine excretion of PD may be misinterpreted as a measure of estimated microbial growth and rumen development; this should be considered during calculations of estimated microbial yield in milk-fed calves.

A2A Adenosine Receptor Antagonists: Are Triazolotriazine and Purine Scaffolds Interchangeable?

The A2A adenosine receptor (A2AAR) is one of the four subtypes activated by nucleoside adenosine, and the molecules able to selectively counteract its action are attractive tools for neurodegenerative disorders. In order to find novel A2AAR ligands, two series of compounds based on purine and triazolotriazine scaffolds were synthesized and tested at ARs. Compound 13 was also tested in an in vitro model of neuroinflammation. Some compounds were found to possess high affinity for A2AAR, and it was observed that compound 13 exerted anti-inflammatory properties in microglial cells. Molecular modeling studies results were in good agreement with the binding affinity data and underlined that triazolotriazine and purine scaffolds are interchangeable only when 5- and 2-positions of the triazolotriazine moiety (corresponding to the purine 2- and 8-positions) are substituted.

Defatted silkworm pupae meal as an alternative protein source for cattle.

Silkworm pupae meal (SWP) is a protein-rich by-product of the silk reeling industry, available in a significant quantity. However, there has been little and insignificant research into the use of SWP in ruminants to date. In this view, the present study was conducted in two phases to evaluate the effect of different inclusion levels of defatted silkworm pupae meal (DSWP) on rumen fermentation, microbial protein synthesis and nutrient utilisation in cattle fed on finger millet straw (FMS)-based diet. Four isonitrogenous concentrate mixtures (CM) were prepared with DSWP replacing soybean meal (SBM) protein at 0 (T0), 10 (T1), 20 (T2) and 30% (T3). In phase I, a rumen fermentation experiment was conducted in a 4 × 4 Latin square design using four crossbred steers to study the effect of different levels of DSWP on rumen fermentation. No significant difference (P > 0.05) was observed in rumen fermentation parameters such as pH, ammonia nitrogen (NH3-N) and total volatile fatty acids (VFA) among the experimental groups. In phase II, the digestibility trial was conducted in 20 crossbred cattle (311.2 ± 4.81 kg), which were divided into four experimental groups of five animals each in a completely randomised design to study the effect of different rations (T0, T1, T2, T3) on microbial protein synthesis and nutrient utilisation. The intake and digestibility of nutrients, excretion of urinary purine derivatives and microbial protein synthesis were not significantly different among the experimental groups. In addition, feeding DSWP revealed no significant (P > 0.05) change in the blood biochemical parameters of animals. Furthermore, at the same price as SBM, DSWP provides two units more crude protein. Therefore, the results of the present study indicated that DSWP can be incorporated into the ration of cattle up to 30% by replacing SBM without affecting rumen fermentation pattern and nutrient utilisation.

Effect of supplementing sulphate-reducing bacteria along with sulphur on growth performance, nutrient utilization and methane emission in goats.

A competitive relationship exists between sulphate-reducing bacteria and methanogens in the anaerobic environment including rumen for hydrogen where sulphate is not limiting growth and consequently inhibit enteric methane emission as thermodynamically energetic sulphate reduction (∆Go = - 21.1 kJ/mole of H2) is more favourable than methanogenesis (∆Go = - 16.9 kJ/mole H2). To validate this hypothesis, a study was designed to investigate the effect of supplementation of sulphate-reducing bacteria (SRB) identified as Streptococcus caviae RM296 as microbial feed additives alone or along with sulphur (as sodium sulphate) on methane production, live weight gain, feed intake, nutrient digestibility and energy metabolism in goats. The experiment was conducted on growing kids (n = 36, 5-6 months of age) with average body weight of 10.08 ± 0.21 kg, divided into six groups (n = 6). The duration of the feeding trial was of 150 days. The six treatments were control fed a basal diet (T1), SRB 0.5 ml/kg BW (T2), sulphur (as sodium sulphate) 0.095% of DMI (total sulphur level in the diet 1.5 times the requirement) (T3), sulphur (as sodium sulphate) 0.095% of DMI + SRB 0.5 ml/kg BW (T4), sulphur (as sodium sulphate) 0.19% of DMI (total sulphur level in the diet 2 times the requirement) (T5) and sulphur (as sodium sulphate) 0.19% of DMI + SRB 0.5 ml/kg BW (T6). Duration of study was 150 days and goats were fed as per ICAR (2013) feeding standard. Methane (CH4) production (l/kg DMI) was reduced by 11.8% (P = 0.052) in T6 where sulphur (0.19% DMI) was supplemented along with SRB4 (at the rate 0.5 ml/kg BW) as compared to T1 (un-supplemented group). However, the dry matter intake (DM), total weight gain (TG), average daily gain (ADG), feed conversion ratio (FCR), excretion of purine derivatives (allantoin, uric acid, xanthine and hypoxanthine) and digestibility of organic matter (OM), dry matter (DM), ether extract (EE), crude protein (CP), acid detergent fibre (ADF) and neutral detergent fibre (NDF) were similar (P > 0.05) among all the groups. The experimental data revealed that feeding of SRB as a microbial feed additive along with sulphur (as sodium sulphate) is capable of reducing enteric CH4 emission without any adverse effect on rumen fermentation and digestibility of the nutrients.

Effects of riboflavin supplementation on performance, nutrient digestion, rumen microbiota composition and activities of Holstein bulls.

To investigate the influences of dietary riboflavin (RF) addition on nutrient digestion and rumen fermentation, eight rumen cannulated Holstein bulls were randomly allocated into four treatments in a repeated 4 × 4 Latin square design. Daily addition level of RF for each bull in control, low RF, medium RF and high RF was 0, 300, 600 and 900 mg, respectively. Increasing the addition level of RF, DM intake was not affected, average daily gain tended to be increased linearly and feed conversion ratio decreased linearly. Total tract digestibilities of DM, organic matter, crude protein (CP) and neutral-detergent fibre (NDF) increased linearly. Rumen pH decreased quadratically, and total volatile fatty acids (VFA) increased quadratically. Acetate molar percentage and acetate:propionate ratio increased linearly, but propionate molar percentage and ammonia-N content decreased linearly. Rumen effective degradability of DM increased linearly, NDF increased quadratically but CP was unaltered. Activity of cellulase and populations of total bacteria, protozoa, fungi, dominant cellulolytic bacteria, Prevotella ruminicola and Ruminobacter amylophilus increased linearly. Linear increase was observed for urinary total purine derivatives excretion. The data suggested that dietary RF addition was essential for rumen microbial growth, and no further increase in performance and rumen total VFA concentration was observed when increasing RF level from 600 to 900 mg/d in dairy bulls.

Synthesis, In Vitro and In Silico Anticancer Activity of New 4-Methylbenzamide Derivatives Containing 2,6-Substituted Purines as Potential Protein Kinases Inhibitors.

A novel class of potential protein kinase inhibitors 7-16 was synthesized in high yields using various substituted purines. The most promising compounds, 7 and 10, exhibited inhibitory activity against seven cancer cell lines. The IC50 values for compounds 7 and 10 were 2.27 and 2.53 µM for K562 cells, 1.42 and 1.52 µM for HL-60 cells, and 4.56 and 24.77 µM for OKP-GS cells, respectively. In addition, compounds 7 and 10 dose-dependently induced the apoptosis and cell cycle arrest at G2/M phase, preventing the cell division of OKP-GS cells. Compounds 7, 9, and 10 showed 36-45% inhibitory activity against PDGFRα and PDGFRß at the concentration of 1 µM. Molecular modeling experiments showed that obtained compounds could bind to PDGFRα as either type 1 (compound 7, ATP-competitive) or type 2 (compound 10, allosteric) inhibitors, depending on the substituent in the amide part of the molecule.

A New Smoothened Antagonist Bearing the Purine Scaffold Shows Antitumour Activity In Vitro and In Vivo.

The Smoothened (SMO) receptor is the most druggable target in the Hedgehog (HH) pathway for anticancer compounds. However, SMO antagonists such as vismodegib rapidly develop drug resistance. In this study, new SMO antagonists having the versatile purine ring as a scaffold were designed, synthesised, and biologically tested to provide an insight to their mechanism of action. Compound 4s was the most active and the best inhibitor of cell growth and selectively cytotoxic to cancer cells. 4s induced cell cycle arrest, apoptosis, a reduction in colony formation and downregulation of PTCH and GLI1 expression. BODIPY-cyclopamine displacement assays confirmed 4s is a SMO antagonist. In vivo, 4s strongly inhibited tumour relapse and metastasis of melanoma cells in mice. In vitro, 4s was more efficient than vismodegib to induce apoptosis in human cancer cells and that might be attributed to its dual ability to function as a SMO antagonist and apoptosis inducer.

Valorization of dietary edible mushrooms waste: chemical and physical properties, nutrient digestibility, microbial protein synthesis and nitrogen balance in sheep.

BACKGROUND: The optimal use of feed resources must be considered by most livestock farmers. The use of low-cost agricultural by-products and the processing of these materials is one possible solution in this respect. One such compound is edible button mushroom waste (EM), a large amount of which is produced annually in the mushroom production cycle worldwide. RESULTS: Bulk density 100 of EM was smaller than the other groups. These changes also applied to alfalfa for bulk density, which was higher than the replaced waste. The dry matter solubility of EM was higher than that of alfalfa hay, whereas the ash solubility rate for EM was greater compared to alfalfa. Replacing up to 210 g kg-1 alfalfa with EM did not affect the production of purine derivatives, microbial protein, nitrogen excreted in urine and feces, and retained nitrogen, although the organic matter digestibility (OMD) increased, whereas the crude protein digestibility and neutral detergent fiber (NDF) decreased (P < 0.05). Fermentation potential, gas production rate, metabolizable energy and short-chain fatty acids were increased. On replacing up to 210 g kg-1 alfalfa with EM, the diet OMD increased, whereas the crude protein and NDF digestibility decreased (P < 0.05). CONCLUSION: EM usage in the experimental diets did not affect the production of purine derivatives, microbial protein, nitrogen excreted in urine and feces, and retained nitrogen. The physical properties, chemical composition and nutritional value of EM, as well as its low cost, show that it can be used as an alternative part of the diet forage in the ruminant's diet. © 2021 Society of Chemical Industry.

Influence of chitosan sources on intake, digestibility, rumen fermentation, and milk production in tropical lactating dairy cows.

The aim of study was to compare the influence of chitosan sources (commercial chitosan vs chitosan extract) on rumen fermentation, methane (CH4) emission, and milk production in tropical lactating dairy cows. Six lactating Holstein-Friesian crossbreeds (410 ± 5 kg, 120 ± 21 day-in-milk) were arranged in a 3 × 3 replicated Latin square design. In addition to control, a 2% chitosan extract supplement and a 2% commercial chitosan supplement of dry matter intake were the treatments. The results denoted that no significant differences on daily dry matter, nutrients, or estimated energy intake were noted when cows received different sources of chitosan. Nutrient digestibility was not influenced differently by extraction-based or commercial chitosan supplements. The pH, temperature, ammonia nitrogen, blood urea, and microbial count were similar among treatments. The different sources of chitosan supplements did not change the totals of volatile fatty acids, acetate, and butyrate; in contrast, different chitosan sources influenced (P<0.05) propionate content. The ruminal acetate to propionate ratio was markedly (P<0.05) reduced with chitosan supplement, but no change appeared between sources of chitosan. At 4 h after feeding, the methane estimation significantly decreased with the addition of chitosan supplementation (P<0.05) compared to the control group. The purine derivatives and microbial protein synthesis were not altered by the treatments. No significant differences existed on milk yield, milk composition, or milk urea nitrogen when cows received different sources of chitosan (P>0.05). In summary, supplementing extracted chitosan showed more potential than did the commercial chitosan for enhancing economic efficiency and recycling shrimp residues, therefore, reducing environmental waste.

New aromatic 6-substituted 2'-deoxy-9-(ß)-d-ribofuranosylpurine derivatives as potential plant growth regulators.

Cytokinins are naturally occurring substances that act as plant growth regulators promoting plant growth and development, including shoot initiation and branching, and also affecting apical dominance and leaf senescence. Aromatic cytokinin 6-benzylaminopurine (BAP) has been widely used in micropropagation systems and biotechnology. However, its 9-glucoside (BAP9G) accumulates in explants, causing root inhibition and growth heterogenity. To overcome BAP disadvantages, a series of ring-substituted 2'-deoxy-9-(ß)-d-ribofuranosylpurine derivatives was prepared and examined in different classical cytokinin bioassays. Amaranthus, senescence and tobacco callus bioassays were employed to provide details of cytokinin activity of 2'-deoxy-9-(ß)-d-ribosides compared to their respective free bases and ribosides. The prepared derivatives were also tested for their recognition by cytokinin receptors of Arabidopsis thaliana AHK3 and CRE1/AHK4. The ability of aromatic N6-substituted adenine-2'-deoxy-9-(ß)-d-ribosides to promote plant growth and delay senescence was increased considerably and, in contrast to BAP, no loss of cytokinin activity at higher concentrations was observed. The presence of a 2'-deoxyribosyl moiety at the N9-position led to an increase in cytokinin activities in comparison to the free bases and ribosides. The antioxidant capacity, cytotoxicity and effect on the MHV-68 gammaherpesvirus strain were also examined.

New 2,6,9-trisubstituted purine derivatives as Bcr-Abl and Btk inhibitors and as promising agents against leukemia.

Bcr-Abl and Btk kinases are among the targets that have been considered for the treatment of leukemia. Therefore, several strategies have focused on the use of inhibitors as chemotherapeutic tools to treat these types of leukemia, such as imatinib (for Bcr-Abl) or ibrutinib (for Btk). However, the efficacy of these drugs has been reduced due to resistance mechanisms, which have motivated the development of new and more effective compounds. In this study, we designed, synthesized and evaluated 2,6,9-trisubstituted purine derivatives as novel Bcr-Abl and Btk inhibitors. We identified 5c and 5d as potent inhibitors of both kinases (IC50 values of 40 nM and 0.58/0.66 µM for Abl and Btk, respectively). From docking and QSAR analyses, we concluded that fluorination of the arylpiperazine system is detrimental to the activity against two kinases, and we also validated our hypothesis that the substitution on the 6-phenylamino ring is important for the inhibition of both kinases. In addition, our studies indicated that most compounds could suppress the proliferation of leukemia and lymphoma cells (HL60, MV4-11, CEM, K562 and Ramos cells) at low micromolar concentrations in vitro. Finally, we preliminarily demonstrated that 5c inhibited the downstream signaling of both kinases in the respective cell models. Therefore, 5c or 5d possessed potency to be further optimized as anti-leukemia drugs by simultaneously inhibiting the Bcr-Abl and Btk kinases.

Design, synthesis, antitumor activity and theoretical calculation of novel PI3Ka inhibitors.

PI3Kα has been identified as an ideal target to treat with PIK3CA gene mutation disease, including drugs such as Alpelisib and Copanlisib. Five purine analogues and four thiazole analogues were designed and synthesized. Their enzymaticactivity against PI3Ka/ß/γ/δ were tested, respectively. All compounds showed excellent selectivity in modulating PI3Ka activity, and parts of the compounds showed good inhibition. Meanwhile, we used Autodock 4.2 to explore the binding mode of the most potential compound Tg with the target protein. In addition, DFT was used to calculate the HOMO-LUMO maps of the compounds Tf, Tg and positive control. This paper will provide some useful information for further drug design of PI3Kα inhibitors.

Milk production responses, rumen fermentation, and blood metabolites of dairy cows fed increasing concentrations of forage rape (Brassica napus ssp. Biennis).

The aim of this study was to determine animal performance, rumen fermentation, and health-related blood metabolites of dairy cows in mid lactation fed with increasing levels (30 and 45%) of forage rape (FR) in the diet. Twelve pregnant multiparous lactating Holstein-Friesian dairy cows were randomly allocated to 1 of 3 dietary treatments in a replicated 3 × 3 Latin square design. The experiment was divided into three 21-d periods. For the control diet, 13.0 kg (dry matter, DM) of grass silage, 3.0 kg DM of commercial concentrate, 2.7 kg of DM cold-pressed extracted canola meal, and 0.45 kg DM of solvent-extracted soybean meal were offered daily. For the other two treatments, 30 and 45% of the DM from silage, canola meal, and commercial concentrate were replaced in equal proportions with FR. Data were analyzed individually using linear and quadratic orthogonal polynomials. Ingestive behavior was altered by the inclusion of FR. We observed a linear increase in eating time at the expense of rumination time. Nevertheless, total DM intake was not affected by dietary treatments, averaging 19.5 ± 0.24 kg of DM/d. Milk yield increased linearly with increasing concentration of FR in the diet. Thus, feed efficiency of cows (kg of milk/kg of DM intake) increased linearly with the percentage of FR in the diet. Inclusion of FR in the diet had no effect on milk composition or milk sensory characteristics. Mean rumen pH of cows decreased linearly from the control to the 45% FR diet; however, dietary treatments had no effect on the daily amount of time that rumen pH was below 5.8 (252 ± 71.4), indicating no risk of subacute ruminal acidosis. Concentrations of total volatile fatty acids in the rumen and molar proportions of acetate and butyrate were increased with FR inclusion, whereas the proportion of propionate was linearly reduced. Excretion of uric acid and total purine derivatives tended to be greater for cows fed FR, which resulted in a trend toward a linear increase in estimated microbial N flow. However, N use efficiency was not affected by FR inclusion. Although differences for some hematological measures (increased white blood cell and neutrophils counts) and a quadratic response for glutamate dehydrogenase for cows fed FR in the diet (decreased with inclusion of 30% and increased with 45% in the diet) were observed, all values were within appropriate ranges for dairy cows. These results indicated that including FR to dairy cow diets, up to 45% of diet DM, improved milk production due to changes in volatile fatty acids and predicted microbial N flow and had no negative effects on dairy cow health or sensory characteristics of milk.

Promising 2,6,9-Trisubstituted Purine Derivatives for Anticancer Compounds: Synthesis, 3D-QSAR, and Preliminary Biological Assays.

We designed, synthesized, and evaluated novel 2,6,9-trisubstituted purine derivatives for their prospective role as antitumor compounds. Using simple and efficient methodologies, 31 compounds were obtained. We tested these compounds in vitro to draw conclusions about their cell toxicity on seven cancer cells lines and one non-neoplastic cell line. Structural requirements for antitumor activity on two different cancer cell lines were analyzed with SAR and 3D-QSAR. The 3D-QSAR models showed that steric properties could better explain the cytotoxicity of compounds than electronic properties (70% and 30% of contribution, respectively). From this analysis, we concluded that an arylpiperazinyl system connected at position 6 of the purine ring is beneficial for cytotoxic activity, while the use of bulky systems at position C-2 of the purine is not favorable. Compound 7h was found to be an effective potential agent when compared with a currently marketed drug, cisplatin, in four out of the seven cancer cell lines tested. Compound 7h showed the highest potency, unprecedented selectivity, and complied with all the Lipinski rules. Finally, it was demonstrated that 7h induced apoptosis and caused cell cycle arrest at the S-phase on HL-60 cells. Our study suggests that substitution in the purine core by arylpiperidine moiety is essential to obtain derivatives with potential anticancer activity.

Effects of feed intake level on efficiency of microbial protein synthesis and nitrogen balance in Boran steers consuming tropical poor-quality forage.

This study aimed at evaluating the effects of feed intake level on the efficiency of rumen microbial protein synthesis (EMPS), nitrogen (N) excretion, and N balance in twelve 18-months old Boran (Bos indicus) steers with initial average liveweight of 183 kg (standard deviation (SD) 15.2). The experiment followed a 4 × 4 complete Latin Square design with four dietary treatments tested in four periods. Each period ran for 5 weeks with 3 weeks of adaptation and 2 weeks of sample collection; separated by 2 weeks of re-feeding. Steers were fed at 100%, 80%, 60%, and 40% of their metabolisable energy requirement for maintenance (MER, referred to as MER100, MER80, MER60, and MER40, respectively). Steers receiving MER80, MER60, and MER40 were only fed Rhodes grass hay. MER100 steers were offered Rhodes grass hay at 80% of their MER and cottonseed meal and sugarcane molasses at each 10% of MER. Mean daily dry matter intake differed between treatments (p < 0.001) and ranged between 2.1 kg/animal (SD 0.13) in MER40 and 4.5 kg/animal (SD 0.31) in MER100. Urinary N excretion and N balance did not differ between MER80, MER60, and MER40. According to contrast test, declining feed intake level from MER80 to MER40 reduced duodenal microbial crude protein flow (p < 0.001), but did not alter the EMPS (g microbial N/kg digestible organic matter intake). Yet, if scaled to N intake, EMPS increased (p < 0.049), whereas total N and faecal N excretions decreased linearly with declining intake level (p < 0.001 for both variables). At similar grass hay intake, duodenal microbial crude protein flow was 41% higher in MER100 than in MER80 steers (p < 0.001). In cattle offered poor-quality tropical forage below their MER, the very low EMPS and thus microbial protein supply aggravate the negative effects of low dietary nutrient and energy intakes in periods of feed shortage.