Buserelin 100micrograms/dose nasal spray

Buserelin acetate Quality by design Chromatography BA-Loaded polymeric nanoparticles Validation A B S T R A C TBuserelin acetate (BA) is the first gonadotropin hormone to reduce the level of estrogen for the treatment of breast cancer.In the present study, RP-HPLC study has been developed and validated subsequently using the analytical quality by design (AQbD) approach.Initially, an analytical target profile (ATP) was defined that outlined the performance of the established method.The risk identification and its assessment were performed using the Ishikawa fishbone diagram and risk assessment method (RAM) to identify critical method parameters (CMPs) having influence on critical analytical attributes (CAAs).The flow rate and pH of buffer were identified as CMPs and retention time (Rt) and peak area (Pa) were recognized as CAAs.The optimization of the method was determined by response surface methodology based on central composite design (CCD).The chromatographic separation was achieved by mobile phase (water: acetonitrile, 80:20 %, v/v) and pH was adjusted using orthophosphoric acid with Zorbax Eclipse plus C18 (4.6 mm 150 mm 5 m) column.Elution was monitored at 220 nm using a photodiode array (PDA) detector.The calibration curve showed the linearity (regression coefficient, R 2 = 0.9991) over the concentration range of 10-60 g/mL.The limit of detection (LOD) and limit of quantitation (LOQ) were found to be 0.051 g/mLand 0.254 g/mL respectively.The method for analysis of BA was accurate using recovery ranging from 100.55 0.93 to 103.45 0.32 whereas the method was precise with % RSD for all parameters of chromatographic system was found to be not more than 1.0 %.Further, the method was robust based on intentionally changing the chromatographic conditions according to the recommended ICH Q2 (R1).Furthermore, poly D, L-lactic-co-glycolic (PLGA-Resomer RG505 and Resomer RG750) (50:50) based nanoparticles were prepared to encapsulate BA and understand the release of BA over the period of 48 h with Korsmeyer-Peppas release kinetics model.The stability of the stock solution was assessed over the 8th day and found to be stable for a longer duration of time.The method has been successfully applied for the analysis of BA in polymeric nanoparticles.

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The use of exogenous hormones has long been of interest for improving reproductive performance in swine production. Enhancing litter size directly impacts the economic efficiency of pig production. Various strategies, including nutritional, genetic, and hormonal approaches, have been explored with varying degrees of success. Administering a gonadotropin-releasing hormone (GnRH) agonist, such as buserelin, at the onset of estrus can induce ovulation and reduce the variation in ovulation timing among sows. This study assessed the impact of GnRH agonist supplementation in boar semen doses on the litter size of inseminated gilts. The research was conducted on a commercial swine herd in northern Thailand. A total of 231 Landrace × Yorkshire crossbred gilts, aged 224.5 ± 16.2 days at the onset of estrus synchronization, participated in the experiment. The gilts’ estrus was synchronized with oral altrenogest supplementation at a dosage of 20 mg/day for 18 days. After exhibiting standing estrus, the gilts were randomly divided into three groups. Control group: gilts were inseminated at 0 and 12 h post standing estrus onset with a conventional semen dose (n = 94). Treatment 1: similar to the control group, but with an added 5 µg (1.25 mL) of buserelin acetate to the boar semen dose during the first insemination (n = 71). Treatment 2: similar to the control group, but with 10 µg (2.5 mL) of buserelin acetate added to the boar semen dose during the first insemination (n = 66). All gilts were inseminated twice during their standing estrus using the intrauterine artificial insemination method. Each semen dose contained 3.0 × 109 motile sperm in 80 mL. The farrowing rate averaged 78.8% and did not significantly differ between the groups (p = 0.141). The total number of piglets born per litter in the treatment 2 group was greater than in the control group (14.0 ± 0.3 vs. 13.2 ± 0.3, respectively, p = 0.049), but was not significantly different from the treatment 1 group (13.3 ± 0.3, p = 0.154). Similarly, the number of live-born piglets in the treatment 2 group was greater than in the control and treatment 1 groups (13.2 ± 0.4 vs. 12.3 ± 0.3 and 12.0 ± 0.4, respectively, p < 0.05). Moreover, the live-born piglets’ litter birth weight in the treatment 2 group was greater than in the control group (17.0 ± 0.4 vs. 15.6 ± 0.3 kg, respectively, p = 0.008) and the treatment 1 group (15.7 ± 0.4 kg, p = 0.025). In conclusion, adding a GnRH agonist to boar semen appears to enhance the litter size of gilts. Further research should focus on understanding the underlying mechanisms and determining the optimal dose and timing for GnRH agonist supplementation.

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The subject of polycystic ovary syndrome (PCOS) has been extensively covered in the literature; however, there is a paucity of data regarding eumenorrheic women with hyperandrogenism and/or hyperandrogenemia without ultrasound evidence of PCO morphology (EuHyperA), and even less data on the comparison between PCOS and EuHyperA subjects. It has previously been shown that around half of PCOS women exhibit a hyper-response of serum 17-hydroxy-progesterone (17-OHP) to the stimulation by GnRH-agonists, also indicated as functional ovarian hyperandrogenism (FOH). Often, this stimulation test is preceded by suppression of the adrenal steroidogenesis with oral dexamethasone (Dex). FOH has been associated with an increase of the P450c17 activity in the ovaries driven by elevated insulin levels. Interestingly, treatment of women with PCOS with Dex suppression and GnRH-agonist stimulation (buserelin) highlighted the possible existence of two clusters of patients: hyper-responders (HR) and normal responders (NR). In this retrospective study, we included 15 hyper-responders (HR) EuHyperA, 34 normal responders (NR) EuHyperA, 62 HR-PCOS and 45 NR-PCOS. The demographic characteristics, glucose-metabolism indices, and the hormonal response to Dex or buserelin were analyzed, with both intra-group and inter-group comparisons performed. The rate of FOH was significantly greater in PCOS than EuHyperA women. Compared to HR-PCOS, HR-EuHyperA had [i.] significantly greater age at observation; [ii.] lower cortisol, 17-OHP, Δ4-androstenedione (Δ4-ASD), total testosterone (TT), LH, and buserelin-stimulated whole curve of dehydroepiandrosterone sulfate (DHEAS), 17-OHP, Δ4-ASD and TT. Compared to NR-PCOS, NR-EuHyperA had [i.] significantly greater FSH, and buserelin-stimulated whole curve of DHEAS; [ii.] significantly lower post-HD Dex Δ4-ASD, TT, buserelin-stimulated whole curve of 17-OHP, Δ4-ASD and TT. Compared to NR-PCOS, HR-PCOS had [i.] significantly greater body mass index (BMI), homeostasis model assessment for insulin resistance (HOMA-IR), cortisol, DHEAS, Δ4-ASD, TT, FT, FAI, E2, and insulin AUC 0-120min (area under the curve) at oral glucose tolerance test (OGTT); [ii] higher levels of post-LD and post-HD Dex 17-OHP, Δ4-ASD, TT, post-HD Dex DHEAS (with greater levels indicating weaker adrenal suppression), whole curve of DHEAS, 17-OHP, Δ4-ASD, TT and LH; [iii] significantly lower sex-hormone binding globulin (SHBG). Even if most of the parameters evaluated were statistically similar in the two sets of comparisons, interesting differences were observed. Women with PCOS exhibit higher androgen levels at baseline, after adrenal suppression and at the buserelin test, further to a higher ovarian volume. Of note, the percentage of women with HOMA-IR≥2.5 and serum insulin levels were greater in PCOS group compared to EuHyperA women. Moreover, within women with PCOS, the HR subgroup has higher insulin levels compared to the NR subgroup, when OGTT is performed. The alteration of the glucose-insulin balance and elevation of circulating androgens were more pronounced in PCOS, thus indicating that [i.] metabolic alterations might be crucial in the onset of PCOS itself and, [ii] EuHyperA might represent a milder form of PCOS.

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Purpose: In this study, we prepared inhalable buserelin microparticles using the spray freeze-drying (SFD) method for pulmonary drug delivery. Raffinose as a cryoprotectant carrier was combined with two levels of five different cyclodextrins (CDs) and then processed by SFD. Methods: Dry powder diameters were evaluated by laser light scattering and morphology was determined by scanning electron microscopy (SEM). Differential scanning calorimetry (DSC) and X-ray diffraction (XRD) analysis were utilized for the determination of crystalline structures. The aerodynamic properties of the spray freeze-dried powders were evaluated by twin stage impinger (TSI) and the stability of prepared samples was assessed under normal and accelerated conditions. Results: The prepared powders were mostly porous spheres and the size of microparticles ranged from 9.08 to 13.53 μm, which are suitable as spray-freeze dried particles. All formulations showed amorphous structure confirmed by DSC and XRD. The aerosolization performance of the formulation containing buserelin, raffinose and 5% beta-cyclodextrin (β-CD), was the highest and its fine particle fraction (FPF) was 69.38%. The more circular and separated structures were observed in higher concentrations of CDs, which were compatible with FPFs. The highest stability was obtained in the formulation containing hydroxypropyl beta-cyclodextrin (HP-β-16. CD) 5%. On the contrary, sulfobutylether beta-cyclodextrin (SBE-β-CD) 5% bearing particles showed the least stability. Conclusion: By adjusting the type and ratio of CDs in the presence of raffinose, the prepared formulations could effectively enhance the aerosolization and stability of buserelin. Therefore, they can be proposed as a suitable career for lung drug delivery.

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The present study compared 2 strategies to initiate a progesterone (P4)-based timed artificial insemination (TAI) protocol for lactating dairy cows: only GnRH or estradiol benzoate (EB) plus GnRH (EB+GnRH). Lactating Holstein cows (n = 487; 184 primiparous and 303 multiparous) from 2 commercial dairy herds were used for their second or greater services postpartum. Each week, cows that were nonpregnant at the pregnancy diagnosis 32 d after a previous AI were randomly assigned to 1 of 2 experimental groups that differed only in the strategy to initiate (d 0) the TAI protocol. On d 0, every cow received a 2.0-g P4 implant; in the EB+GnRH group, cows were treated with 2.0 mg i.m. of EB and 16.8 µg i.m. of the GnRH analog buserelin acetate, whereas in the GnRH group, cows received only 16.8 µg i.m. of GnRH. On d 7 after the initial treatment, 0.530 mg i.m. of cloprostenol sodium (PGF) was administered in all cows, followed by a second dose on d 8, concomitant with 1.0 mg i.m. of estradiol cypionate and P4 implant withdrawal. The TAI was performed on d 10 (48 h after P4 device withdrawal) in both experimental groups. Only conventional Holstein semen was used throughout the study. The percentage of cows with corpus luteum (CL) on d 0 (73%) and overall ovulation rate after d 0 (54%) did not differ between groups. The CL regression between d 0 and the first PGF treatment was greater in the EB+GnRH group than the GnRH group (42% vs. 31%). Consequently, the proportion of cows with CL at PGF was greater when only GnRH was used on d 0 compared with EB+GnRH (86% vs. 82%), and the mean number of CL at PGF was greater (1.23 vs. 1.11). The expression of estrus near TAI was greater in GnRH group (84% vs. 77%), and cows showing estrus had greater (44% vs. 10%) pregnancy per AI (P/AI) on d 32 for both treatments. We found no effect of the presence of CL on d 0 or at PGF, nor of ovulation after d 0 or CL regression between d 0 and d 7 on fertility. However, fertility was critically impaired when cows did not have CL at both times, d 0 and at PGF treatment. We did not observe any interaction between treatment and other variables, and the P/AI was similar in cows receiving EB+GnRH or only GnRH on d 0 (37.8% vs. 36.6%). In summary, although there was no detectable difference in P/AI between treatments, this study demonstrated potential negative physiological outcomes caused by EB treatment on d 0 (greater incidence of luteolysis after d 0 and fewer cows with CL at PGF treatment). Overall, we found no benefit of adding EB at the initiation of a P4-based TAI protocol on fertility compared with using GnRH alone, despite differences in ovarian dynamics and expression of estrus.

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Rabbit is a reflexively ovulating species. Accordingly, in the practice of artificial insemination (AI) ovulation must be induced via exogenous GnRH (Gonadotropin-Releasing Hormone) administration, which may be performed intramuscularly, subcutaneously, or intravaginally. Unfortunately, the bioavailability of the GnRH analogue when added to the extender is lower due to the proteolytic activity in the seminal plasma and the poor permeability of the vaginal mucosa. The aim of the study was to refine the practice of AI practice in rabbits by replacing parenteral GnRH analogue administration (subcutaneous, intravenous, or intramuscular injection) with intravaginal application, while reducing its concentration in the diluent. Extenders containing the buserelin acetate in chitosan-dextran sulphate and chitosan-alginate nanoparticles were designed and 356 females were inseminated. Reproductive performance of females inseminated with the two experimental extenders, receiving 4 μg of buserelin acetate intravaginally per doe, was compared with that in the control group, the does of which were inseminated with the extender without the GnRH analogue and induced to ovulate with 1 μg of buserelin acetate administered intramuscularly. The entrapment efficiency of the chitosan-dextran sulphate complex was higher than that of chitosan-alginate. However, females inseminated with both systems showed similar reproductive performance. We conclude that both nanoencapsulation systems are an efficient way of intravaginal ovulation induction, allowing a reduction in the level of the GnRH analogue normally used in seminal doses from 15-25 μg to 4 μg.

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Modern reproductive technologies require artificial insemination (AI) protocols that are safe for the health of animals and do not violate their welfare. Stress factors arising in the reproduction process of industrial rabbit breeding negatively affect the sexual function and reproductive performance of rabbits and their offspring. In recent years, researchers have focused on alternative methods of ovulation stimulation, including intravaginal introduction of hormonal means of its induction. Therefore, the aim of our study was a comparative evaluation of the effectiveness of the intramuscular method of administration of the gonadotropin-releasing control (GnRH) analogue and its addition to the sperm dose for AI on Hyplus rabbit (n = 30), which were randomly divided into three groups. For this, buserelin acetate was administered intramuscularly at a dose of 1 μg per rabbit immediately after artificial insemination (control group). Experimental animals were inseminated with sperm doses with the addition of a GnRH analogue in the amount of 25 μg (experimental group 1) and 15 μg (experimental group 2). The influence of the method of administration of GnRH analogue on reproductive function was evaluated by indicators of the effectiveness of AI, fertility, and the dynamics of the progesterone level during pregnancy. A decrease in fecundity was observed in females; for example, in experimental group 1, the effectiveness of AI in terms of the number of rabbits was 64.3 %; in animals of experimental group 2 – 41.2 %, and the number of inseminations in this group of rabbits was 41.7% higher compared to control data. The fertility of rabbits of all groups was high, reaching the maximum value in the control group (10.42 ± 0.52 heads/female). Fertility according to live newborns in experimental group 1 was the highest among all groups of rabbits and was 9.67 ± 0.49 heads/female, which showed an increasing trend compared to the control. Also, in the animals of experimental group 1, an increase in the level of progesterone in blood serum was observed (on the 7th day of pregnancy by 13.2 %, on the 14th day by 14.3 %, on the 21st day by 16.4 %). In contrast, in experimental group 2, a significant decrease in the progesterone level was noted. Thus, 25 μg of GnRH analogue added to the extender is optimal. It is worth noting that there were no advantages of intravaginal administration over intramuscular administration in terms of reproductive performance. However, the number of live births and progesterone levels were improved.

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Fixed-timed artificial insemination (FTAI) protocols combining estradiol (E2) and progesterone (P4) are commonly used in the tropics, with adequate pregnancy rates throughout the year in buffaloes. However, the application of E2 and its related esters in food-producing animals is banned in the European Union and by the FDA-USA. This study aimed to evaluate the pregnancy rate (PR) after FTAI in buffalo cows (Bubalus bubalis) treated during the breeding (September-January) and non-breeding season (February-June) using a protocol based on P4 intravaginal devices (IVD) combined with GnRH and PGF2α. The trial was performed on a commercial buffalo farm in Zulia State, Venezuela. Two hundred forty-seven buffalo cows (1-14 calvings) with a body condition score (BCS) of 3.15 ± 0.39 were selected. Cows were synchronized on a random day of the estrus cycle. To synchronize the ovulation on day 0, a dose of 10.5 μg GnRH-analog (Buserelin benzoate acetate, Gestar®) was injected intramuscularly (IM) plus the insertion of an IVD maintained for nine days [TRIU B®: 1.0 gr P4, (n=139); CIDR®: 1.38 gr P4 (n=77); or PREGNAHEAT-E®: 250 mg of medroxyprogesterone acetate (MPA), (n= 18)]. On day 9, IVD was removed, and 0.50 mg of Cloprostenol (Estrumate®, Intervet, MSD Animal Health) plus 500 IU of eCG (Folligon®, Intervet, MSD Animal Health) were given IM. On day 11, a dose of 10.5 μg GnRH-analog IM was used, and 8-12 hours later, they were artificially inseminated using frozen-thawed semen of bulls with proven fertility. The pregnancy rate (PR) was determined 30-32 days after AI by ultrasound. The data was analyzed by logistic regression through the Proc Logistic, and continuous and linear character variables were analyzed through the Proc GLM of the SAS® package. The overall PR was 52.2% (n = 129/247). Evaluation of independent variables, such as the number of calving and BCS, did not affect the PR (p>0.05). The PR was similar (p>0.05) amongst the reproductive and non-reproductive seasons [52.2%: (66/129) vs. 53.4% (63/118), respectively]. IVD-type did not affect the PR (p>0.05) [TRIU- B®: 52.6% (80/152), CIDR®: 53.3% (41/77) and PREGNAHEAT-E®: 44.4% (8/18)]. In conclusion, this FTAI protocol based on P4 and GnRH and PGF2α produced adequate pregnancy rates during breeding and non-breeding seasons. These findings suggest that GnRH could be considered to substitute E2 esters in the P4-based FTAI protocols in water buffaloes under tropical conditions.

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