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This experiment was designed to evaluate the effects of increasing doses of estradiol cypionate (ECP) on reproductive function of lactating dairy cows during the summer. In Exp. 1, 643 lactating Holstein cows were blocked by parity and assigned to receive 1) an intravaginal P4 device (1.9 g of P4) and 2.0 mg of estradiol benzoate on day -11, 25 mg (i.m.) of dinoprost tromethamine on day -4, 1.0 mg (i.m.) of estradiol cypionate and CIDR withdrawal on day -2, followed by TAI on day 0 (n = 326; ECP-1) or 2) the same synchronization protocol with 2.0 mg of ECP on day -2 (n = 317; ECP-2). In both treatments, cows were TAI on day 0 of the protocol, and cow rectal temperature was measured on days -2, 0, and 7. In Exp. 2, 608 lactating crossbred Holstein × Gir dairy cows were blocked by parity and enrolled to the same treatments as in Exp. 1, but on day 7, cows received one viable embryo into the uterine horn. In Exp. 1, a greater percentage of ECP-2 cows were detected on estrus (81.3 vs. 91.1%, respectively). A treatment × body condition score (BCS) interaction was observed on day 60 pregnancy per AI (P/AI), as ECP-2 cows with a BCS <2.75 had a greater P/AI vs. ECP-1, but an opposite result was observed in cows with a BCS ≥2.75. Regardless of treatment, there were effects of mean rectal temperature and heat stress events on P/AI. Treatment affected the diameter of the ovulatory follicle at TAI (ECP-1 = 15.3 mm vs. ECP-2 = 14.8 mm) and a greater percentage of ECP-1 cows had larger follicles (≥16.5 mm), but ECP-2 resulted in a greater incidence of early ovulatory cows (ovulating before day 0). Therefore, follicle diameter at TAI affected P/AI on day 60 in cows receiving ECP-2 and tended to affect ECP-1 cows. A treatment effect was observed on time to estrus following ECP treatments and a reduced proportion of ECP-2 cows showed estrus at TAI. Regardless of treatment, cows detected on estrus 48 h after ECP administration had a greater P/AI on day 60 vs. cows detected on estrus 24 h. In Exp. 2, a greater percentage of ECP-2 cows were detected on estrus (68.3 vs. 81.4%, respectively). In summary, a greater dose of ECP increased the percentage of animals expressing estrus, but it did not benefit the reproductive function of lactating dairy cows during the summer, regardless if animals were assigned to a TAI or timed-embryo transfer (TET) protocol., (Copyright © 2022 Elsevier Inc. All rights reserved.)

There are multiple factors that contribute to reduced fertility in lactating dairy cows. Recently, a reproductive tract size and position score (SPS) system was developed as a management tool to identify dairy cows with decreased fertility. The aim of this study was to evaluate the association between the SPS on fertility outcomes such as ovulation failure, pregnancy per artificial insemination (P/AI), concentration of pregnancy-associated glycoproteins (PAGs), and pregnancy loss in lactating dairy cows. Primiparous and multiparous lactating Holstein cows (n = 869) were enrolled at two locations. Location 1 (Loc. 1) in Minas Gerais, Brazil (n = 613) and location 2 (Loc. 2) in Agassiz, British Columbia, Canada (n = 256). At the time of AI (d 0), cows were classified as SPS (small [SPS1], medium [SPS2], or large [SPS3] sized reproductive tract) and ovulation failure was determined at 48 h and 7 d post-AI via ultrasonography (Loc. 2 only). Blood samples were collected on d 24 and 31 of gestation for quantification of PAGs and pregnancy diagnosis was performed via ultrasonography at d 31 and 60 post-AI (Loc. 1) and at d 31 ± 3 and 60 ± 3 post-AI (Loc. 2). Cows diagnosed pregnant at d 31 post-AI but not pregnant at d 60 were defined to have undergone late embryonic pregnancy loss. Parity was found to impact SPS (P < 0.01), as primiparous cows had a higher frequency of SPS1 and lower frequency of SPS3 when compared with multiparous cows (SPS1: 42.6 vs. 15.0%; SPS3: 7.0 vs. 22.0%, respectively). Cows classified as SPS3 had greater ovulation failure at 48 h (P = 0.04) and 7 d post-AI (P = 0.05). Cows classified as SPS1 had greater P/AI when compared to SPS2 and SPS3 (45.9 ± 3.3 vs. 37.4 ± 2.6 and 29.1 ± 3.5%, respectively; P = 0.004). There was no interaction between parity and SPS on P/AI. Pregnancy loss between 31 and 60 d post-AI was increased in cows classified as SPS3 compared to SPS2 and SPS1 (24.3 ± 0.05 vs. 11.6 ± 0.02 and 9.4 ± 0.02%, respectively; P = 0.04). Cows classified as SPS1 and SPS2 had greater concentrations of PAGs at 31 d post-AI when compared to SPS3 at both Loc.1 (P < 0.01) and Loc. 2 (P < 0.01). There was no interaction between SPS and pregnancy loss on PAGs at 24 and 31 d post- AI for either Loc. 1 (P = 0.75 and P = 0.76, respectively) or Loc. 2 (P = 0.61 and P = 0.81, respectively). In conclusion, cows that were classified as SPS3 had greater ovulation failure, reduced P/AI, similar concentrations of PAG on d 24, but decreased on d 31, and a greater incidence of pregnancy loss. Thus, size and position of the reproductive tract is associated with fertility and this scoring system could be used to make reproductive management decisions on dairy operations., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The objective of this study was to determine the association of beta-carotene concentration in plasma at the moment of artificial insemination (AI) on pregnancy/AI in lactating Holstein cows. A total of 399 events from 364 lactating Holstein cows were enrolled in the trial (143 primiparous and 221 multiparous). All cows were assigned to a timed AI protocol based on estradiol and progesterone. Blood samples were collected at the moment of AI and at 24 and 31d post-AI (samples on 31 d post-AI were collected only from cows that were diagnosed pregnant). The BCS were recorded at the time of AI. Plasma beta-carotene was quantified from blood samples taken at the time of AI using a single step denaturation and extraction into a solvent, followed by measurement using a portable spectrophotometer. Pregnancy-associated glycoproteins (PAG) were analyzed in blood samples taken at 24 and 31 d post-AI of pregnant cows. Milk production was collected for the entire experimental period. Pregnancy diagnosis was performed by ultrasound 31 and 60 d post-AI. Data was analyzed using the MIXED and GLIMMIX procedures of SAS. Cows classified as thin (<2.75) tended to have lower concentration of beta-carotene at AI when compared with those classified as Moderate (≥3.00; 3.8 ± 0.1 vs. 4.3 ± 0.1 μg/mL; P = 0.09). Concentration of beta-carotene were greater in multiparous compared with primiparous (P < 0.01). There was no correlation between concentration of beta-carotene and milk production (r = 0.04; P = 0.10). When plasma beta-carotene was categorized in quartiles, cows in the 1st quartile had lower pregnancy/AI and higher pregnancy losses when compared with cows that were in the 2nd, 3rd and 4th quartile (pregnancy/AI = 19.2 ± 4.5, 33.7 ± 4.7, 36.9 ± 5.0 and 39.8 ± 5.4%, respectively; P = 0.05; pregnancy losses = 41.9 ± 4.8, 20.4 ± 3.7, 22.1 ± 4.1, and 15.7 ± 4.2%, respectively; P < 0.05). There was no association between concentrations of beta-carotene at AI and PAG at 24 d post-AI (P = 0.60). Cows with greater concentrations of beta-carotene at AI were more likely to have greater concentrations of PAG at 31 d post-AI (P < 0.01). In conclusion, the concentration of beta-carotene at AI was affected by BCS and parity. Cows with higher concentrations of plasma beta-carotene at AI had greater pregnancy/AI, lower pregnancy losses and greater concentrations of PAG at d 31 post-AI, suggesting it may be associated with placental function in lactating dairy cows., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Expression of estrus (EST) near the time of fixed-time artificial insemination (FTAI) increases pregnancy success in beef females. This outcome has been associated with improved pregnancy establishment and maintenance, although research is still warranted to validate this theory. Hence, this experiment compared ovarian, uterine and conceptus factors associated with pregnancy establishment in Bos indicus beef cows according to estrous expression during a FTAI protocol. One hundred lactating multiparous Nelore cows received a 2 mg injection of estradiol benzoate and an intravaginal progesterone (P4) releasing device on day -11, a 12.5 mg injection of prostaglandin F2α on day -4, P4 device removal in addition to 0.6 mg injection of estradiol cypionate and 300 IU injection of equine chorionic gonadotropin on day -2, and FTAI on day 0. An estrous detection patch was attached to the tailhead of each cow on day -2, and estrous expression was defined as removal of >50% of the rub-off coating from the patch at FTAI. Overall, 39 cows expressed EST, 55 did not express EST (NOEST), and six cows lost their patch and were discarded from the experiment. Ovarian ultrasonography was performed at FTAI, and on days 7 and 15 of the experiment. Blood samples were also collected on days 7 and 15. Only cows without a corpus luteum (CL) on day 0, and with a CL on days 7 and 15 remained in the experiment (EST, n=36; NOEST, n=48). On day 15, cows were randomly selected within each group (EST, n=29; NOEST, n=30) for conceptus collection via transcervical flushing, followed by endometrial biopsy in the uterine horn ipsilateral to the CL. Within cows not assigned to conceptus collection, blood samples were collected for whole blood RNA extraction (day 20) and pregnancy status was verified by transrectal ultrasonography (day 30). Diameter of dominant follicle on day 0 and plasma P4 concentrations on day 7 were greater (P⩽0.02) in EST v. NOEST cows. Conceptus length and messenger RNA (mRNA) expression of prostaglandin E synthase and interferon-tau were greater (P⩽0.04) in EST v. NOEST cows. Moreover, EST cows diagnosed as pregnant on day 30 had greater (P<0.01) blood mRNA expression of myxovirus resistance 2 on day 20 compared with NOEST. In summary, estrous expression near the time of FTAI enhanced pregnancy establishment factors in B. indicus cows, including conceptus development and mRNA expression of interferon-tau.

Bioluminescence, the emission of visible light in a living organism, is an intriguing phenomenon observed in different species and environments. In terrestrial organisms, the bioluminescence is observed mainly in beetles of the Elateroidea superfamily (Coleoptera). Several phylogenetic studies have been used different strategies to propose a scenario for the origin and evolution of the bioluminescence within this group, however some of them showed incongruences, mainly about the relationship of the bioluminescent families. In order to increase the number of molecular markers available for Elateroidea species and to propose a more accurate phylogeny, with high supported topology, we employed the Next-Generation Sequencing (NGS) methodology to perform the RNA-Seq analysis of luminescent (Elateridae, Phengodidae, Rhagophthalmidae, and Lampyridae) and non-luminescent (Cantharidae) species of Neotropical beetles. We used the RNA-Seq data to construct a calibrated phylogeny of Elateroidea superfamily using a large number of nuclear molecular markers. The results indicate Lampyridae and Phengodidae/Rhagophthalmidae as sister-groups, suggesting that the bioluminescence evolved later in Elateridae than other families (Lampyridae, Phengodidae, and Rhagophthalmidae), and indicating the Upper Cretaceous as the period for the main diversification of Elateroidea bioluminescent species., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The objective of this study was to assess the impact of mastitis-causing bacteria and somatic cell count (SCC) on pregnancy per embryo transfer (P/ET) in Holstein-Gir crossbred (Girolando) lactating dairy cows. Cows (n = 1397) were subjected to a timed-embryo transfer protocol. Milk samples were collected two days before embryo transfer for SCC and bacteriological culture analyses. Pregnancy diagnosis was performed on days 31 and 66 after timed-embryo transfer. The animals were grouped according to the National Mastitis Council recommendations: Gram-positive environmental (EV+), Gram-negative environmental (EV-), Gram-positive contagious (C+), coagulase-negative staphylococci (CNS) and control (no bacterial growth). Additional analysis was made by categorizing bacteria based on degree of pathogenicity (Major or Minor). Bacterial growth reduced P/ET (P < .01) at both 31 and 66 days of gestation. The P/ET was lower (P < .05) at 31 days in EV- (30.1%) and EV+ (29.9%) groups and tended (P = .09) to be lower in the C+ group (36.6%) than the control group (44.0%). The P/ET from the Major group at 31 days of gestation was lower (P = .03) compared with the Minor and control groups (32.1 vs 41.1 vs 43.2%, respectively). Cows with SCC > 400,000 cells/mL had lower P/ET (P < .01) than animals with SCC < 200,000 cells/mL at both 31 (30.4% vs 40.8%) and 66 days (24.7% vs 32.2%) of gestation. Pregnancy loss was not different between bacterial isolates and SCC categories. Elevated SCC significantly reduced P/ET, whereas environmental agents and those with Major pathogenicity yielded the greatest reduction in P/ET., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The aim of this study was to test the effect of expression of estrus at artificial insemination (AI) on endometrium, conceptus, and CL gene expression of beef cows. Thirty-six multiparous nonlactating Nelore cows were enrolled on an estradiol- and progesterone (P4)-based timed AI protocol (AI = Day 0) and then slaughtered for the endometrium, CL, and conceptus collection on Day 19. The animals were retrospectively grouped on the basis of cows that (1) showed signs of estrus near AI (n = 19; estrus) and (2) did not show any signs of estrus (n = 17; nonestrus). Body condition score, blood sampling, and ultrasound examination were performed on Days 0, 7, and 18 of the experiment followed by messenger RNA extraction and quantitative reverse transcription polymerase chain reaction analysis of 58 target genes. Data were checked for normality and analyzed by ANOVA for repeated measures using proc GLM, MIXED, and UNIVARIATE of SAS. Only pregnant cows were included in the analyses (n = 12; nonestrus, n = 11). Estrous expression had no correlation with parameters such as body condition score, preovulatory follicle and CL diameter, P4 concentration in plasma on Days 7 and 18 after AI, and interferon-tau concentration in the uterine flushing (P > 0.15); however, a significant increase was observed in conceptus size from cows that expressed estrus (P = 0.02; 38.3 ± 2.8 vs. 28.2 ± 2.9 mm). The majority of transcripts affected by estrous expression in the endometrium belong to the immune system and adhesion molecule family (MX1, MX2, MYL12A, MMP19, CXCL10, IGLL1, and SLPI; P ≤ 0.05), as well as those related with prostaglandin synthesis (OTR and COX-2; P ≤ 0.05). Genes related to apoptosis, P4 synthesis, and prostaglandin receptor were downregulated (CYP11A, BAX, and FPr; P < 0.05) in the CL tissue of cows that expressed estrus. In addition, four genes were identified as differentially expressed in the 19-day-old conceptus from cows that expressed estrus (ISG15, PLAU, BMP15, and EEF1A1; P < 0.05). There was also a significant effect of Day 7 concentration of P4 mainly affecting the immune system, adhesion molecules, and wnt signaling pathway of the endometrium (IGLL1, MX2, SLPI, TRD, APC, WNT2, GLYCAM1, and MYL12A; P < 0.05). A significant interaction between estrous expression and P4 concentration on Day 7 was more pronounced in immune system genes (MX1, MX2, TRD, SLPI, and IGLL1; P < 0.05). This study reported that estrous expression at the time of AI favorably altered the gene expression profile in reproductive tissues during the preimplantation phase toward a more receptive state to the elongating conceptus. These effects seem to be more evident in the endometrium during the time of dynamic remodeling for embryo implantation., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Holstein heifers (n = 57) were monitored using accelerometers and video observations with the objective of better understanding the behavioral expression of estrus, the variation within and between the heifers, and the possible sources of variation. IceTags recorded walking activity from 7 to 13 months of age. Activity peaks (n = 282) were obtained from a rolling sum of steps within 24-hour periods and validated to be estrus by ovarian ultrasonography. Behavior around activity peak of one estrus for each of 12 heifers was described in detail from video recordings. Baseline behavior was monitored in a corresponding interval 1 week before. Estrus and baseline total steps and steps per hour, estrus relative increase in activity, duration, and interval between episodes were analyzed by descriptive statistics and Spearman rank correlations. Effects of category of baseline walking activity, estrus order (pubertal vs. second and greater episodes), season, hour of estrus onset, and number of heifers simultaneously in estrus were evaluated with proc MIXED. Behavioral changes from baseline to estrus were evaluated by a signed-rank test. Estrus total steps varied greatly (4743 ± 1740; range: 837-10,070), as well as the relative increase in activity (290 ± 160%; range: 30%-1190%). Duration of estrus was 14 ± 4 hours, ranging from 4 to 26 hours. The interval between episodes was the trait that varied the least. Pubertal estrus was shorter and had a smaller relative increase in activity than second and greater episodes (P < 0.05). The number of steps during estrus was greater for heifers of high baseline activity (P < 0.01). Estrus episodes occurring in the winter and starting between 4 PM and 3 AM had the greatest relative increase in activity (P < 0.05). The number of heifers simultaneously in estrus did not influence estrus expression (P > 0.05). The behaviors with greatest change from baseline to estrus were chin rest, sniff, back mount, crossover, accept chin rest, and follow, but variation was large. Overall, estrus was apparent in behavioral changes with large variation within and between the heifers. Estrus order, onset hour, season, and baseline walking activity are important factors affecting estrus activity. Therefore, estrus detection tools should account for potential sources of variation. The visual and automated measurements of estrus expression reported in this study reveal possibilities for improved on-farm estrus detection technologies and potential genetic selection for estrus expression., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The objectives were to evaluate pregnancy per AI (P/AI) of dairy cows subjected to the 5-day timed AI protocol under various synchronization and luteolytic treatments. Cows were either presynchronized or received supplemental progesterone during the synchronization protocol, and received a double luteolytic dose of PGF2α, either as one or two injections. In Experiment 1, dairy cows (n=737; Holstein=250, Jersey=80, and crossbred=407) in two seasonal grazing dairy farms were randomly assigned to one of four treatments in a 2×2 factorial arrangement. The day of AI was considered study Day 0. Half of the cows were presynchronized (G6G: PGF2α on Day -16 and GnRH on Day -14) and received the 5-day timed AI protocol using 1 mg of cloprostenol, either as a single injection (G6G-S: GnRH on Day -8, PGF2α on Day -3, and GnRH+AI on Day 0) or divided into two injections of 0.5 mg each (G6G-T: GnRH on Day -8, PGF2α on Day -3 and -2, and GnRH+AI on Day 0). The remaining cows were not presynchronized and received a controlled internal drug-release (CIDR) insert containing progesterone from GnRH to the first PGF2α injection of the 5-day timed AI protocol, and 1 mg of cloprostenol either as a single injection on Day -3 (CIDR-S) or divided into two injections of 0.5 mg each on Days -3 and -2 (CIDR-T). Ovaries were examined by ultrasonography on Days -8 and -3 and plasma progesterone concentrations were determined on Days -3 and 0. In Experiment 2, 655 high-producing Holstein cows had their estrous cycle presynchronized with PGF2α at 46±3 and 60±3 days postpartum and were randomly assigned to receive 50 mg of dinoprost during the 5-day timed AI protocol, either as a single injection or divided into two injections of 25 mg each. Pregnancies per AI were determined on Days 35 and 64 after AI in both experiments. In Experiment 1, presynchronization with G6G increased the proportion of cows with a CL on Day -8 (80.6 vs. 58.8%), ovulation to the first GnRH of the protocol (64.2 vs. 50.2%), and the presence (95.6 vs. 88.4%) and number (1.79 vs. 1.30) of CL at PGF(2α) compared with CIDR cows. Luteolysis was greater for two injections compared to a single PGF2α injection (two PGF2α=95.9 vs. single PGF2α=72.2%), especially in presynchronized cows (G6G-T=96.2 vs. G6G-S=61.7%). For cows not presynchronized, two PGF2α injections had no effect on P/AI (CIDR-S=30.2 vs. CIDR-T=34.3%), whereas for presynchronized cows, it improved P/AI (G6G-S=28.7 vs. G6G-T=45.4%). In Experiment 2, the two-PGF2α injection increased P/AI on Days 35 (two PGF2α=44.5 vs. single PGF2α=36.4%) and 64 (two PGF2α=40.3% vs. single PGF2α=32.6%) after AI. Presynchronization and dividing the dose of PGF2α (either cloprostenol or dinoprost) into two injections increased P/AI in lactating dairy cows subjected to the 5-day timed AI protocol., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Lactating Holstein cows (n=135) were randomly assigned to one of the two sources of supplemental selenium (Se), sodium selenite (SS) or Se yeast (SY), fed at 0.3mg/kg diet dry matter from 25 d before calving to 70 d in milk (DIM), in diets not suboptimal in basal Se concentrations. Cows were evaluated for health daily in the first 10 DIM, and uterine cytology of the previously gravid uterine horn was assessed at 30 DIM. The Ovsynch protocol was initiated at 42 DIM; ovarian responses to hormonal treatments were evaluated by ultrasonography. The uteri of cows were flushed 6d after timed AI for collection of embryos and oocytes. Plasma concentrations of Se and progesterone were measured throughout the postpartum period and during the reproductive protocol, respectively, and plasma glutathione peroxidase activity was determined 6d after AI. Concentrations of Se in pre- and postpartum diets ranged from 0.43 to 0.56 mg/kg of dry matter. Incidence of retained placenta, fever, ketosis, mastitis, acute puerperal metritis, clinical endometritis, and subclinical endometritis were not significantly different between treatments. There were no differences between groups in concentrations of Se and progesterone or glutathione peroxidase activity in plasma. Treatment did not influence ovarian responses to the synchronization protocol, fertilization rate, number of blastomeres and live blastomeres, or proportions of grades 1 and 2, degenerated, and degenerated-unfertilized embryos/oocytes. Odds of subclinical endometritis on Day 30 postpartum more than doubled in cows with fever of unknown origin or acute puerperal metritis in the first 10 DIM. Fertilization rate tended to be reduced in cows with subclinical endometritis. In summary, replacing SS with an organic source of Se in diets not suboptimal in basal Se concentrations did not improve Se status, uterine health, fertilization, or embryo quality in early lactation dairy cows.

Nutrition of the donor cow can influence oocyte and embryo quality, which can affect the success of embryo transfer. Severe undernutrition compromised ovarian follicular development, with implications for superovulatory response and embryo quality. In postpartum lactating cows, undernutrition or inability to consume sufficient nutrients delayed resumption of ovulation, reduced the number of follicles, and compromised oocyte quality. Moderate undernutrition of nonlactating cows was unlikely to affect embryo quality; conversely, nonlactating animals on maintenance diets usually had better superovulatory responses and improved oocyte competence and embryo quality. The negative effects of overfeeding are thought to be mediated by alterations in endocrine cues, such as hyperinsulinemia and increased glucose and IGF-I, which may interfere with glucose transport in the embryo and increase apoptosis. Manipulating energy sources such as carbohydrates and fatty acids (FA) may influence embryo viability, but the effects of FA were not consistent in vitro; increasing concentrations of unsaturated FA in follicular and embryonic cells usually improved embryo viability and resistance to cryopreservation. Excess protein intake and increased urea and ammonia in body fluids can be toxic to embryos, impairing their development; these effects seemed to be associated with alterations in uterine pH and granulosa cell function. Likewise, toxins in feeds (e.g. gossypol), reduced embryo development and increased pregnancy losses. Diet of donor cows should be formulated to optimize the supply of nutrients to meet needs; however, manipulating energy intake, source of FA and protein content of donor diets, particularly moderate underfeeding in nonlactating cows, may further optimize responses.

In recent years, the frequent appearance of phenethylamine designer drugs on the illicit drug market has been a matter of concern for all authorities involved. New phenethylamine drugs are being introduced because these compounds are not covered by existing legislation. Therefore, the new drugs cannot be considered illicit drugs until their names are officially recognized. This paper describes a method to screen for and quantify four 2,5-methylenedioxy-derivatives of 4-thio-phenethylamine (2C-T-series) in human plasma, using capillary electrophoresis coupled with electrospray ionisation-mass spectrometry (CE-ESI-MS). Prior to CE-MS analysis, a simple liquid extraction was used for sample cleanup. The method was validated according to international guidelines.

In recent years, a number of newer designer drugs have entered the illicit drug market. The methylenedioxy-derivates of amphetamine represent the largest group of designer drugs. This paper describes a method for screening for and quantification of ten 2,5-methylenedioxy-derivates of amphetamine and phenylethylamine in human urine, using capillary electrophoresis coupled to electrospray ionisation-mass spectrometry (CE-ESI-MS). Prior to CE-MS analysis, a simple solid-phase extraction (SPE) was used for sample cleanup. The method was validates according to international guidelines.

This study examined the influence of a GnRH agonist containing either 450 or 750 microg of deslorelin in an implant form or a gonadorelin injection (control) to induce ovulation in the Ovsynch protocol on pregnancy rates (PR), embryonic loss, and ovarian function in 593 lactating Holstein cows. Cows were given two injections of PGF2alpha 14 days apart, followed 14 days later by the Ovsynch protocol, and were timed artificially inseminated (TAI) at 68 +/- 3 days postpartum. Blood samples for determination of plasma progesterone concentrations were collected at 24 and 10 days prior to and 11 days after TAI. Pregnancy was diagnosed on Day 27 and reconfirmed on Day 41 after TAI. Non-pregnant, not re-inseminated cows at Day 27 had their ovaries examined by ultrasonography, and the number and size of follicles and presence of luteal tissue were determined. Simultaneously, these cows were re-synchronized with the Ovsynch protocol. Pregnancy during the re-synchronization period was determined between 35 and 41 days after insemination. On Day 27, PR were higher for control (39.0%) and deslorelin 450 microg (DESLORELIN 450) implant (41.3%) than for those receiving the deslorelin 750 microg (DESLORELIN 750) implant (27.5%; P<0.05). Pregnancy losses tended to decrease for DESLORELIN 450 compared with control (5.0% versus 12.7%; P<0.13). Plasma progesterone concentrations did not differ significantly among treatments. Deslorelin suppressed ovarian activity and decreased PR during the re-synchronization period compared with control. The percentage of non-pregnant animals that were re-inseminated by Day 27 was less for deslorelin compared with control. In conclusion, incorporation of an implant of the GnRH agonist deslorelin to induce ovulation in the Ovsynch protocol has the potential to reduce pregnancy losses, but the response was dependent upon implant concentration. Evaluation of lower doses to minimize the negative effects on subsequent fertility is warranted.

Objective: To evaluate in a multicenter retrospective study, the clinical and laboratory characteristics and the outcome of patients with acute myeloid leukemia (sAML) previously diagnosed with breast cancer (BC) among an adult acute leukemia population., Patients and Methods: Between June 1992 and July 1996, 3934 new cases of adults with acute leukemia were recorded in GIMEMA Archive of Adult Acute Leukemia (2964 AML, 901 ALL, 69 acute leukemia expressing both myeloid and lymphoid surface markers)., Results: Two hundred patients (5.1%) presented with a history of previous malignancy (21 of them were affected by ALL and 179 by AML). Among sAML, 37 patients (29%) had a previous breast cancer. They consisted of 36 females and 1 male, median age 56 years, range 34-87. The median latency between the 2 malignancies was 54 months (range 5-379). Twenty-seven patients received chemo- and/or radiotherapy for breast cancer (7 only chemotherapy, 6 only radiotherapy, and 14 combined treatment). All patients were surgically treated but in 10 patients surgical debridement was the sole therapy for breast cancer. The drugs most frequently employed were alkylating agents (18 patients), topoisomerase II inhibitors (9 patients), antimetabolites (20 patients) (CMF, CEF and MMM combinations). At onset of sAML the median WBC count was 7.7 x 10(9)/l (0.8-153) and the median platelet count was 33.5 x 10(9)/l (3-305). Considering morphological features, FAB subtypes were 4 M0, 5 M1, 11 M2, 5 M3, 8 M4, 3 M5, and 1 M6. Cytogenetic study was performed on 28 patients and 12 of them presented abnormalities. It is noteworthy that chromosome 5 or 7 abnormalities (typically observed in those patients treated with alkylating agents) were present only in three cases. Thirty-four patients received chemotherapy for sAML, and twenty-five of them achieved a CR (74%), with a median duration of twenty-eight weeks (5-280+). The overall survival was 8 months (1-80+)., Discussion: The high number of sAML we observed in patients with a previous breast cancer, may be due to the fact that this malignancy is the most frequent neoplasm in women and by the high probability of cure with a consequent long disease-free survival. Our results suggest that the risk of sAML after recovery from breast cancer is increasing due to the rise in the number of patients cured from breast cancer, and in the future could be a relevant problem for haematologists.

Seventeen (un)substituted N-[4-(propyl)cyclohexyl]-amides (6a-h, 7a-h and 8) were synthesized and tested as anti-inflammatory and analgesic agents. The substituents on the aromatic ring were chosen in order to study the influence of electron-withdrawing or electron-donating residues, that change the electronic density on the aromatic moiety. The pharmacological results allow drawing some preliminary considerations on structure-activity relationships.

Two series of N-[4-(alkyl)cyclohexyl]-substituted benzamides, i.e. a series of N-[4-(tert-butyl)cyclohexyl]-substituted benzamides and a series of N-[4-(ethyl)cyclohexyl]-substituted benzamides, were synthesised and evaluated for their anti-inflammatory and analgesic potencies, and gastrointestinal irritation liability.

Two series of 1-methyl-4-(N-aroyl)-piperidinamides were synthesized and evaluated for their anti-inflammatory and analgesic properties, as well as for their gastrointestinal irritation liability. A non-aromatic derivative, 1-methyl-4-(N-cyclohexanoyl)-piperidinamide, was synthesized and evaluated in order to obtain a more exhaustive knowledge of the structure-activity relationship.

Two hundred fifty-three patients with newly diagnosed acute promyelocytic leukemia (APL) were eligible to enter the multicentric GIMEMA-AIEOP "AIDA" trial during the period July 1993 to February 1996. As a mandatory prerequisite for eligibility, all patients had genetic evidence of the specific t(15;17) lesion in their leukemic cells confirmed by karyotyping or by reverse transcription-polymerase chain reaction (RT-PCR) of the PML/RAR alpha fusion gene (the latter available in 247 cases). Median age was 37.8 years (range, 2.2 to 73.9). Induction treatment consisted of oral all-trans retinoic acid (ATRA), 45 mg/m2/d until complete remission (CR), given with intravenous Idarubicin, 12 mg/m2/d on days 2, 4, 6, and 8. Three polychemotherapy cycles were given as consolidation. Hematologic and molecular response by RT-PCR was assessed after induction and after consolidation. At the time of analysis, 240 of the 253 eligible patients were evaluable for induction. Of these, 11 (5%) died of early complications and 229 (95%) achieved hematologic remission. No cases of resistant leukemia were observed. Of 139 cases studied by RT-PCR after induction, 84 (60.5%) were PCR-negative and 55 (39.5%) PCR-positive. One hundred sixty-two patients were evaluable by RT-PCR at the end of consolidation. Of these, 159 (98%) tested PCR-negative and 3 (2%), PCR-positive. After a median follow up of 12 months (range, 0 to 33), the estimated actuarial event-free survival for the whole series of 253 eligible patients was 83% +/- 2.6% and 79% +/- 3.2% at 1 and 2 years, respectively. This study indicates that the AIDA protocol is a well-tolerated regimen that induces molecular remission in almost all patients with PML/RAR alpha-positive APL. Preliminary survival data suggest that a remarkable cure rate can be obtained with this treatment.

A series of N-Aroyl-cyclohexyl- and cyclohexenylamides 3- or 4-methylsubstituted were synthesized and evaluated for their anti-inflammatory and analgesic potencies, and gastrointestinal irritation liability. One compound, N-benzoyl-4-methyl-cyclohexylamide 6a, possessed an anti-inflammatory activity comparable to that of indomethacin.

The antimycotic activity of 16 o-nitrophenylhydrazones against strains of Hansenula anomala, Saccharomyces cerevisiae, Candida parapsylosis, and Cryptococcus albidus was tested. All 16 compounds inhibited growth of the yeast strains. The inhibitory activity of the 4 methyl-derivatives substituted on the aromatic nucleus was particularly significant.

The glycoside fraction of fresh rhizomes from Magydaris pastinacea (Lam.) Paol. was separated from the alcoholic extract using the method of Kobayashi et al. The fraction was found to have six main constituents, the most abundant of which had previously been isolated and identified as 7-O-beta-D-glucopyranosyl-8-(2',3'-dihydroxy-3-methyl-butylcoumarin++ +). The present paper describes the separation and characterization of the other constituents, all with coumarin or furancoumarin structures. Pharmacological experiments to test these compounds as platelet antiaggregants are also reported.

Forty amides, formamidines and trifluoromethylsulfonylamides bearing on the nitrogen a cyclohexyl residue, eventually 2-substituted, were prepared and tested for analgesic activity against a chemical stimulus. Good activity was exhibited by amides 9, 11 and 28, by formamidine 34, as well as by triflyamide 40. Eleven additional compounds exhibited a moderate activity.

Pursuing our investigations on N-aryl or N-cycloalkyl substituted amides and formamidines with antiinflammatory and/or analgesic activity, two set of cycloalkyl substituted amides and formamidines were prepared and tested for analgesic activity against a chemical stimulus. Some compounds, particularly the amides 7, 11, 17 and the formamidine 35 proved to be endowed with this activity.

Pursuing our investigations on 7-amino-2,3-polymethyleneindole derivatives, a set of 7-(dimethylaminomethylene)-amino-2,3-polymethyleneindoles, together with some other aryl or cycloalkyl substituted formamidines, were prepared and tested for analgesic and antiinflammatory activities. Several compounds resulted endowed with one or both of these activities; the indole derivatives 1 and 2 exhibited a good degree of both of them.