Mycobacterium abscessus infections are increasing worldwide necessitating the development of new antibiotics and treatment regimens. The utility of third-generation tetracycline antibiotics was reestablished; their anti-M. abscessus activity needs further study. The activities of omadacycline (OMC), eravacycline (ERC), tigecycline (TGC), and sarecycline (SAC) were tested against two reference strains and 193 clinical M. abscessus isolates at different temperatures (30°C and 37°C). The minimum bactericidal concentrations (MBCs) of the four drugs were determined to distinguish between their bactericidal and bacteriostatic activities. The MICs of OMC, ERC, and TGC for the reference strains and clinical isolates were summarized and compared. OMC, ERC, and TGC exhibited a high level of bacteriostatic activity against M. abscessus. The MICs of OMC and ERC for M. abscess remained stable, while the MICs of TGC for the isolates/strains increased with increasing temperature. Notably, the MICs of OMC for M. abscessus isolates obtained in the United States are lower than for those obtained in China.IMPORTANCE The antimicrobial activities of four third-generation tetracycline-class drugs, omadacycline (OMC), eravacycline (ERC), tigecycline (TGC), and sarecycline (SAC), were determined for 193 M. abscessus isolates. The activities of the four drugs at two different temperatures (30°C and 37°C) were also tested. OMC, ERC, and TGC exhibited significant activity against M. abscessus. The anti-M. abscessus activity of TGC increased when the temperature was increased from 30°C to 37°C; the activities of OMC and ERC, on the other hand, remained the same. We found that in vitro MICs of OMC against Chinese and American isolates were distinct. Evaluations in in vivo models of M. abscessus disease or in the clinical setting will provide more accurate insight into potency of OMC against distinct isolates. Mycobacterium abscessus complex, consisting of three subspecies, abscessus, massiliense, and bolletii, constitutes the main pathogenic rapidly growing mycobacteria (RGM) , which include both rapidly and slowly growing organisms, are becoming more common worldwide . The Mycia (RGM) , 3. In the world . The incto treat . M. absciectasis –8. Accorcrolides . Usuallycrolides . While tcrolides , 12. TheM. abscessus activity in vitro and in vivo; ERC exhibits activity in vitro , eravacycline (ERC), and sarecycline (SAC) received US Food and Drug Administration (FDA) approval in recent years. Compared to the chemical structures of first- and second-generation tetracycline-class antibiotics, third-generation antibiotics possess more complex groups on the D ring . TGC andin vitro –16. Notae reason , 16, 17.M. abscessus reference strains (ATCC 19977 and CIP 108297) and 193 clinical isolates were collected for antibiotic susceptibility testing. Detailed test results for the activity of four tetracycline analogs  against 193 clinical isolates cultured at 30°C are presented in 50 was 1 μg/mL and the MIC90 was 4 μg/mL for subsp. abscessus; the MIC50 was 2 μg/mL and the MIC90 was 4 μg/mL for massiliense. The MICs of ERC ranged from 0.063 to 8 μg/mL: the MIC50 was 1 μg/mL and the MIC90 was 4 μg/mL for both subspecies abscessus and massiliense. The MICs of TGC ranged from 0.063 to 4 μg/mL: the MIC50 was 0.5 μg/mL and the MIC90 was 2 μg/mL for subsp. abscessus; the MIC50 was 1 μg/mL and the MIC90 was 2 μg/mL for subsp. massiliense. The MICs of SAC ranged from 4 to >8 μg/mL; both the MIC50 and MIC90 were >8 μg/mL, exhibiting significantly less activity than OMC, ERC, and TGC toward M. abscessus isolates. To summarize, TGC, ERC, and OMC were active against M. abscessus. TGC exhibited the greatest activity; OMC and ERC also expressed high anti-M. abscessus; SAC was inactive.Two The MBCs of OMC, ERC, and TGC were tested on the two reference strains and 18 randomly selected isolates following MIC testing, and the MBC/MIC ratios were determined . SAC was excluded since the MICs were >8 μg/mL. OMC, ERC, and TGC exhibited bacteriostatic activity with MBC/MIC ratios >4 or >8 against all strains tested at both 30°C and 37°C.M. abscessus isolates selected at random from 193 isolates were determined after 3 days incubation at 30°C and 37°C. A detailed comparison of the MICs for the 29 strains is shown in Table S3. Paired Wilcoxon tests were performed. SAC was excluded from statistical analysis since all the MICs were >8 μg/mL. The statistical results and MIC distribution are shown in M. abscessus isolates were higher when the incubation temperature was 37°C, compared to 30°C (P < 0.05); increasing the incubation temperature to 37°C, however, did not impact the MICs of OMC or ERC.The MICs of the 4 tetracycline analogs for 29 clinical M. abscessus isolates obtained in the current study and in 5 studies reported previously were collected and compared . OMC, ERC, and TGC were active toward the M. abscessus isolates collected in all the studies: the MIC50 of OMC ranged from 0.12 to 2 μg/mL, and the MIC90 ranged from 0.25 to 8 μg/mL; the MIC50 of ERC ranged from 0.12 to 1 μg/mL, and the MIC90 ranged from 0.25 to 8 μg/mL; the MIC50 of TGC ranged from 0.12 to 1 μg/mL, and MIC90 ranged from 0.25 to 8 μg/mL. Kaushik et al. (90 of OMC (4 μg/mL) and TGC (2 μg/mL) for subsp. abscessus, which were higher than the rest. Zhang et al. and Chew et al. reported the same MIC50 and MIC90 for ERC and TGC, which were the lowest among all the studies . Bax et al. .The anti-sistance . SAC hasM. abscessus infections  disturbances in 62.5% of treated patients, and the availability of an intravenous formulation only  of those infections were of pulmonary origin , purchased from the American Type Culture Collection , and M. abscessus subsp. massiliense (CIP 108297), purchased from the Biological Resource Center of the Institute Pasteur . One hundred ninety-three clinical isolates were obtained from sputum and bronchoalveolar lavage fluid specimens of patients diagnosed with NTM lung infections in the Shanghai Pulmonary Hospital. All genome sequences were published and are available at DDBJ/ENA/GenBank . All strains were grown at 37°C in Middlebrook 7H9 broth supplemented with 10% oleic acidalbumin-dextrose-catalase (OADC) or on Middlebrook 7H10 agar plates supplemented with 10% OADC.Two reference strains were used: OMC, ERC, TGC, and SAC (>98% purity) were purchased from MedChemExpress , solubilized in DMSO, aliquoted, stored at −80°C, and serially diluted just prior to each experiment.M. abscessus reference strains and 193 clinical M. abscessus isolates using the broth microdilution method according to Clinical and Laboratory Standards Institute (CLSI) document M24-A2 (5 CFU/mL in cation-adjusted Mueller-Hinton broth (CAMHB). A 100-μL suspension of bacteria and 100 μL of antibiotic serially diluted 2-fold in sterile saline were suspended evenly in each well of 96-well microtiter plates; control wells contained 100 μL of suspended bacteria and 100 μL of CAMHB only. The final working concentration of the four antibiotics ranged from 0.008 to 8.0 μg/mL. Plates were sealed and incubated at 30°C or 37°C for 3 days. MICs were defined as the minimum concentration at which no visible bacterial growth was observed.Antimicrobial susceptibility testing was performed on the 2 t M24-A2 . BrieflyThe MBC was determined following completion of MIC testing. The contents of each microtiter well containing drug concentrations greater than the MIC were suspended, and 100-μL aliquots were spread onto Middlebrook 7H10 agar plates supplemented with 0.2% glycerol and OADC. The plates were incubated for 5 days at 37°C then the CFU were counted. The MBC value was defined as the minimum drug concentration that yielded 0 CFU/mL. An MBC/MIC ratio ≤4 or >4 was used to classify an antimicrobial agent as bactericidal or bacteriostatic, respectively.M. abscessus isolates cultured at two different temperatures were compared by the Wilcoxon rank sum tests using GraphPad Prism 8 . P values <0.05 were considered statistically significant.The MICs of antibiotics determined for PRJNA488058, PRJNA448987, and PRJNA398137).All genome sequences were published and are available at DDBJ/ENA/GenBank (BioProject


Analysis of the composition of yogurt alternatives (YAs) during fermentation provides critical information for evaluating its quality and nutritional attributes.We investigated the effects of homotypic (HO) and heterotypic (HE) lactic acid bacteria on the nutritional and mineral bioavailabilities of soybean YA (SYA) during fermentation.The acidic amino acid  and organic acid contents in HO-fermented YA were increased from 2.93, 1.71, and 7.43 mg/100 g to 3.23, 1.82, and 73.47 mg/100 g, respectively. Moreover, both HO and HE lactic acid bacteria fermentation enhanced mineral absorptivity. They altered the molecular speciation of minerals from a large molecular type  to a small molecular type , which was manifested in a time-dependent manner. Furthermore, YA substantially increased the bone mass in a zebrafish osteoporosis model, further highlighting the potential of lactic acid bacterial fermentation for mineral bioavailability.This study provides a foundation for understanding the effects of fermentation conditions on the composition and bioavailability of minerals in YA and can assist in its production. Daily iSoybean milk is a primary source of mineral elements that play essential roles in sustaining life and supporting fundamental body functions –10. For In recent years, yogurt alternatives fermented by lactic acid bacteria have effectively enhanced the number of bioactive ingredients and flavors in soy milk and have attracted increasing attention. In our preliminary experiments, we found that the absorption rate of calcium significantly increased in yogurt alternatives, which could be related to changes in the mineral element morphology during fermentation. However, it is not clear whether the absorptivity of other mineral elements besides calcium increases during fermentation. Furthermore, the relationship between bacterial strains, mineral morphology, and absorption remains unclear. Health problems are highly related to diet and nutritional habits . Therefoin vitro and in vivo. The molecular weight distribution and the speciation of minerals in the YA samples were characterized by protein structural changes, amino acid production of the strains, and the release of organic acids during fermentation.To test this speculation, the present study aimed to investigate the absorptivity and speciation of minerals in YAs produced using different strains of lactic acid bacteria. Here, we prepared two YA samples by fermenting soybean milk with two different lactic acid bacteria and analyzed their absorptivity 2.2.1.Lactiplantibacillus plantarum CICC 21022 and Enterococcus faecalis ATCC19433were screened in our laboratory. Briefly, 5 g of pickled Chinese cabbage was washed five times with sterile water. Serial decimal dilutions were obtained, plated on MRS agar supplemented with 2% calcium carbonate, and then incubated under aerobic and anaerobic conditions at 37°C for 48 h. Gram-positive, catalase-negative isolates were considered presumptive lactic acid bacteria. Subsequently, the 16S rDNA segments were amplified and sequenced to identify the structure of lactic acid bacteria. Soybean was purchased from the Chinese Academy of Agricultural Sciences. Fetal bovine serum (FBS), Dulbecco’s Modified Eagle’s medium (DMEM), and other reagents used for cell culture were purchased from Corning . All other reagents were either analytical grade or pure .Pure freeze-dried thermophilic yogurt cultures for the direct-to-vat set form, 2.2.Lactiplantibacillus plantarum CICC 21022 and Enterococcus faecalis ATCC19433 were cultured with 7.512 × 107 CFU/ml, following incubate at 37°C at facultative anaerobic conditions for 12 h, the SYA was collected and stored at −20°C for subsequent use.YA was successfully prepared using a previously reported method  with sli2.3.L. plantarum and E. faecalis populations was determined by plating the aliquots of the serial dilution on MRS agar, and incubating at 37°C for 48 h at facultative anaerobic conditions. The aliquots of the serial dilution was plated on MRS agar, and incubated at 37°C for 48 h under the conditions described above. The colony-forming units per ml (CFU/ml) of L. plantarum and E. faecalis populations was determined following the official microbiological methods.The amount [colony-forming units per ml (CFU/ml)] of 2.4.2.4.1.The total soluble protein content was determined using bicinchoninic acid (BCA) assay. All samples were combined with 6 M HCl and left to hydrolyze for 4 h at 110°C. An automatic amino acid analyzer  was used to determine the amino acid composition. Briefly, 1 g of soybean milk or YAs fermented by different lactic acid bacteria was dissolved in 25 ml ultrapure water. Subsequently, 0.5 ml potassium ferrocyanide (106 g/L) and zinc sulfate (300 g/L) each were added to the solution and allowed to flow statically for 1 h. The separation was performed using a CAPCELL PAK C18 column (150 mm × 4.6 mm × 3 μm) and sodium dihydrogen phosphate solution (20 mmol/L) mobile phase under the following conditions: flow rate, 0.9 ml/min; detection wavelength, 210 nm; column temperature, 30°C; sample injection volume, 10 μl. The sample values were determined based on a calibration curve developed using five standards .2.4.2.SDS-PAGE was performed following a previously reported method  to analy2.5.2.5.1.Changes in free oligosaccharide content were detected using ion chromatography. Briefly, 0.2 g soybean yogurt was dissolved in 10 ml water and diluted to 25 ml by water. The chromatographic separation was performed using a DIONEX ICS-3000 USA system fitted with a Carbo PacTMPA20 3*150 mm analytical column (Diane Corporation of the United States), a pulse ampere detector, and gold electrode under the following conditions: Eluent solution, 250 mM NaOH; flow rate, 0.5 ml/min; injection volume, 10 μl. The separation was carried out under gradient elution conditions using pure water (solvent A) and 250 mM NaOH (Solvent B) as follows: 0–20 min, 94% A, 6% B; 20–20.1 min, 20% A, 80% B; 20.1–30 min, 20% A, 80% B, 30.1–40 min, 94% A, 6% B.2.5.2.g and analyzed, individually.The organic acid content was determined using HPLC. The chromatography was performed using a CAPCELL PAK C18 (150 mm × 4.6 mm × 3 μm) LC column and sodium dihydrogen phosphate solution (20 mmol/L) at a flow rate of 0.9 ml/min as the mobile phase. A sample volume of 10 μl was injected, and the signals were detected using a UV detector at 215 and 210 nm. Oxalic, tartaric, lactic, acetic, citric, succinic, malic, and ascorbic acids were used as standard solutions to develop the calibration curve. Briefly, One gram sample was dissolved in 15 ml buffer containing 0.5 ml potassium ferrocyanide (106 g/L) and 0.5 ml zinc sulfate (300 g/L) and allowed to stand for 1 h at room temperture. Afterward, the sample was centrifuged at 7104 × 2.6.2.6.1.3 (3:1), and left to stand for 4 h. Then, the PTFE tank was placed into a stainless steel coat and heated for 4 h at 165°C. The Ca, Zn, Mg, Fe, and P contents of the samples were determined using inductively coupled atomic emission spectrometry . The experimental conditions, including the auxiliary and gas flow rates, cooling gas flow, and generator power, were 0.8 L/min, 0.8 L/min, 12 L/min, and 1.4 kW, respectively.Soybean milk or YA samples (0.20 g) were weighed (accurate to 0.0001 g), placed in a Teflon  tank, combined with concentrated HNO2.6.2.HPLC, in combination with inductively coupled plasma-mass spectrometry (ICP-MS), was used to detect mineral and trace element species in YA samples during fermentation. Samples were separated using an XBridge Protein BEH SEC 200 Å column (7.8 mm × 300 mm × 3.5 μm). Briefly, the samples (50 mg) were dissolved in 0.03 mol/L Tris–HCl buffer (pH 7.4) and filtered using a 0.45 m filter membrane. The sample (20 μl) was injected and separated using – mobile phase at a flow rate of 0.5 ml/min; the detection wavelength was 214 nm (UV detector). The analysis was performed with external calibration by developing the calibration curve derived from five standards: BSA (66.4 kDa), trypsin inhibitor (20.1 kDa), insulin (5.8 kDa), VB12 (1.35 kDa), and L-tyrosine (0.187 kDa). The molecular weight of the sample peaks was determined based on that of the standards.2.6.3.The human colon carcinoma cell line, Caco-2, was used as an intestinal barrier model. Transport assays were performed in Transwell systems using the method described by Wang et al. . Three tBasal indicates the calcium concentration at different time points on the basal interior side; VBasal denotes the volume corresponding to the time points on the inner basal side; CIntermediate and CApical represent the mineral concentrations on the middle and apical sides, respectively; VIntermediate and VApical denote the corresponding volumes at the middle and apical sides, respectively.C2.6.4.We obtained the data on mineral absorptivity, Bacterial strain, and mineral type were collected and subjected to orthogonal transformation analysis using SPSS 16 software. A group of variables speculated to be correlated was converted into linearly unrelated variables to reduce the dimension while maintaining the characteristics to which the variance contributes the most. Then, we performed principal component analysis (PCA) using the ‘vegan’ package based on the Bray–Curtis distance on the R platform to analyze the correlations and trends of the data. The outputs were obtained in a grid, and the color-synthesis results of the first and second principal components were previewed.2.7.2.7.1.2, and 0.33 mM MgSO4 and treated with 0.1% DMSO (vehicle), 25 μM prednisolone (model), or co-incubated 3.3–30 μM CBP, or 0.308 μM alendronate (positive control) after 3 pdf till 7 dpf. On pdf 7, larvae were collected for skeletal staining and behavioral analysis.All the procedures were performed in accordance with the National Institutes of Health Guidelines for the Care and Use of Laboratory Animals. Adult zebrafish AB strains were kept at 28.5°C under a normal light/dark cycle of 14 h/10 h at 28°C for natural maturation. Zebrafish embryos were cultured in 6-well plates. The collected embryos were placed in fish water containing 5.0 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl2.7.2.The fish at 7 dpf were vitally stained with 0.2% calcein solution (pH adjusted to 7.0–7.5) for 5 min in the dark. Next, fish were rinsed thrice for 5 min with the system water, anesthetized with 0.016% MS-222, and fixed in 3% methylcellulose. At the end of the treatment, the regenerated bone areas on the larvae were visualized using a fluorescence microscope , and images were captured using digital cameras . The relative fluorescence intensity (RFI) of skull bone mass per zebrafish was determined using ImageJ software (n = 6).2.8.p-value of <0.05 were considered statistically significant.All tests were performed at least in triplicates in six independent experiments. The results were subjected to statistical analysis using one-way ANOVA with Duncan’s posthoc test for comparison with the vehicle. All figures were drawn using GraphPad Prism 5.0  and Excel . Differences at a 3.3.1.L. plantarum and E. faecalis, which produce more lactic acid from sugar , α′(71 kDa), and β (45–50 kDa) subunits  than that of E. faecalis-fermented YA (21%). A similar trend of increased absorption in L. plantarum-fermented YA compared to that in E. faecalis-fermented YA was observed in absorption of ferrum, zinc, magnesium, and phosphorus, which declined up to 2 h, and increased thereafter.Changes in the total mineral absorption ratio during fermentation are shown in 3.3.2.Different stains Absorptivity clustering analysis of minerals in soy yogurt fermented by different strains is shown in 3.3.3.L. plantarum are shown in The molecular weight distribution and morphology distribution of minerals in soybean milk during fermentation by E. faecalis exhibited a similar trend.Next, to understand the effect of the form of the mineral on bioavailability, we evaluated the possible mineral forms using HPLC-ICP-MS. As shown in 3.4.L. plantarum) treatments, which reversed (Glucocorticoid Induced Osteoporosis)GIOP-induced bone loss, as shown in Zebrafish have become an attractive animal model for studying mineral bioavailability because of their small size, short developmental cycle, and transparent embryos. Calcium is essential for the prevention of osteoporosis, and adequate intakes of zinc, magnesium, magnesium, and phosphorus are also presumed to play important roles in osteoporosis prevention . To inve4.Yogurt, a fermented milk product obtained by lactic acid bacteria-mediated acidification of milk, is consumed widely in developing and developed countries due to its functional activities . Howeverin vitro and a zebrafish model in vivo. Consumer attitudes toward and acceptability of YAs were also investigated. We found that the content of soluble substances and the absorption rate of minerals increased with decreasing pH during fermentation. The heterolactic lactic acid bacteria performed better than the heterolactic lactic acid bacteria in terms of the absorptivity of the minerals in YA they produced.In this study, the effect of different types of lactic acid bacteria on the absorption rate of minerals during fermentation was analyzed using a Caco-2 cell model L. plantarum uses the carbon source in soybean milk to produce organic acids during fermentation, which reduces the pH, leading to charge neutralization or a shielding effect on the protein surface. Most negatively charged proteins are converted to positively charged  and dipeptide and tripeptide transporter (DtpT), which can transport extracellular peptides into cells; meanwhile, many peptidases in cells can further degrade peptides , which was much better than soybean milk. Similarly, mineral and trace elements significantly improve the nervous system, enhancing swimming distance in zebrafish.The absorbed calcium ions can improve bone density after entering the body. To better understand mineral absorption 5.Enterococcus faecalis) and heterolactic (Lactiplantibacillus plantarum CICC 21022) lactic acid bacteria fermentation on the nutritional and mineral bioavailability of SYA was investigated. The Caco-2 cell model showed the potential of fermentation to improve the acidic amino acid and organic acid contents as well as mineral absorptivity in SYA. The fermentation altered the speciation of minerals from a large molecular type to a small one. Furthermore, the chemical composition changes revealed that peptide calcium might be the main mineral form resulting from fermentation. YA substantially increased the bone mass in a zebrafish osteoporosis model. The analyses in the zebrafish model showed that YA could effectively improve the bioavailability of minerals and trace elements, further highlighting the potential of lactic acid bacterial fermentation for mineral bioavailability. Overall, this study provides evidence that lactic acid bacteria regulates the form and bioavailability of minerals and trace elements in soybean milk during fermentation. The results provide a foundation for understanding the influence of processing conditions on the composition and function of YAs and can assist in the production and sales of SYA.In this study, the effects of homotypic  and the Natural Science Foundation of Shandong (ZR2022MC217) granted its approval to conduct this research. In addition, the Center for Mitochondria and Healthy Aging, College of Life Sciences, Yantai University partly provided support.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Pituitrin injection solution is an indispensable hemostatic utilized in clinical practice and is widely used in myomectomy. However, there have been reports of adverse reactions leading to gastrointestinal injury, hyponatremia and hypokalemia, anaphylaxis, cardiac arrest, etc. Thus, the safety of pituitrin should be taken seriously.In the present study, three cases of cardiac arrest caused by pituitrin injection during laparoscopic myomectomy, who were successfully resuscitated in our hospital, are reported.The clinical data and surgical procedures in the patient should be analyzed to find the causes of cardiac arrest. Medication and resuscitation should be summarized to ensure the safety of the patient. Pituitrin injection solution is a sterile solution extracted from the posterior pituitary gland of animals and contains two active ingredients: oxytocin and vasopressin. These can cause strong contractions of the uterine smooth muscle when injected locally into the uterine body. One study found that the application of pituitrin injection solution into the uterine body during laparoscopic myomectomy was found to promote contractions and reduce hemorrhage . TherefoCase No. 1: a female aged 37. Height: 160 cm, bodyweight: 45 kg. The preoperative diagnosis was cervical fibroids. The patient’s general condition was good. The B-ultrasonography revealed that the uterus was morphologically abnormal, approximately 5.9 × 4.3 cm in size. Only a third of the cervix was visible, while the rest was approximately 7.5 × 5.3 cm with hypo-solid echogenicity. The auxiliary examinations were as follows. There was no abnormality in the electrocardiogram (ECG), chest x-ray, or biochemical assays. The vital signs at the entrance of the operation room were as follows: temperature (T): 36.7℃; respirations (R): 20 times/minute; blood pressure (BP): 123/80 mmHg; heart rate (HR): 90 beats/minute. The laparoscopic myomectomy was conducted under general anesthesia at 19:40. The patient was grade I, based on the American Society of Anesthesiologists. The conventional monitoring results were as follows: BP: 128/85 mmHg and HR: 88 beats/minutes. The ECG showed sinus rhythm. At 21:00, the patient was injected with 6 U of pituitrin injection solution , diluted to 6 ml with 0.9% normal saline. One minute later, an ST-segment downward shift was demonstrated in the ECG, followed by an HR of approximately 20 beats/minute. The operation was stopped immediately, and after treatment with intravenous injection of 0.5 mg of atropine and chest compressions, the patient resumed autonomic rhythm at 21:05 with an HR of 115 beats/minute and normal ECG. The surgery was completed at 23:15, and the patient was safely returned to the intensive care unit (ICU) with intensive monitoring. There were no abnormalities in respiratory, circulatory, or cognitive function at the 3-day postoperative follow-up.Case No. 2: a female aged 32. The preoperative diagnosis was uterine cavity occupancy: residual pregnancy tissue with multiple uterine fibroids. There was no previous medical history. At admission, the patient’s vitals were as follows: T: 36.5℃, P: 76 beats/minute, R: 20 times/minute, BP: 118/80 mmHg, HR: 76 beats/minute. Her height was 158 cm, and her bodyweight was 52 kg. The preoperative examinations were completed, and the results of the B-ultrasonography showed an inhomogeneous and slightly strong echogenicity of 3.0 × 1.8 × 2.6 cm in size, a weak echogenic mass of 4.5 × 3.5 × 4.5 cm in size in the subplasma of the anterior wall, and a weak echogenic mass of 3.6 × 2.9 × 3.4 cm in size and 1.6 cm in diameter in the subplasma of the right wall in the uterine cavity. Sinus rhythm was shown in the ECG with no deviation of the electric axis and no ECG abnormality. A laparoscopic single-site myomectomy was conducted under general anesthesia at 13:00. At 14:13, the patient was intraoperatively injected with 6 U of pituitrin injection solution , diluted to 6 ml with 0.9% of normal saline and injected into the uterine wall. Cardiac arrest occurred in the patient approximately three minutes later. The operation was stopped, and chest compressions were started immediately. Meanwhile, 20 ug of epinephrine and 0.3 mg of atropine were administered intravenously. Sinus rhythm resumed approximately five seconds later with an HR of approximately 45 beats/minute. Then 0.3 mg of atropine was administered intravenously with the continuation of the surgery. The operation ended at 15:25. Vital signs were stable in the patient. There were no abnormalities in respiratory, circulatory, or cognitive function at the 3-day postoperative follow-up.2 infusion and a partial expulsion of CO2 to reduce the abdominal pressure. Chest compressions were conducted, and 40 ug of epinephrine and 0.5 mg of atropine were administered intravenously. The heartbeat resumed after five seconds, and sinus rhythm was restored after 30 s with a BP of 113/74 mmHg. The operation was completed at 13:20, and ICU monitoring continued for 24 h after extubation until the vital signs were stable. The patient was transferred to the general ward with clear consciousness. There were no abnormalities in respiratory, circulatory, or cognitive function at the 3-day postoperative follow-up.Case No. 3: a female aged 40 with a preoperative diagnosis of uterine fibroids, scarred uterus, and mild anemia. The past medical history was as follows. Hemorrhoid surgery was conducted more than five years ago, and she had a cesarean section in 2006. Her general condition at admission was as follows: T: 37.2℃, P: 89 beats/minute, R: 20 times/minute, BP: 126/87 mmHg, HR: 89 beats/minute, height: 155 cm, and bodyweight: 62 kg. The preoperative examinations were conducted. The results of the B-ultrasonography showed two to three weakly echogenic masses with a size of 4.6 × 3.0 × 4.1 cm and clear borders between the muscular walls and subplasma. The ECG results showed sinus rhythm, no deviation of the electric axis, and no ECG abnormality. A laparoscopic myomectomy was conducted under general anesthesia at 11:00. At 11:38, the patient was injected with 6 U of pituitrin injection solution intraoperatively , diluted to 6 ml with 0.9% of normal saline. The pituitrin was injected into the uterine wall and avoid undesirable systemic effects of inadvertent intravascular injection. Approximately two minutes later, the ECG waveform disappeared in the monitor. The surgical operation was immediately stopped with a pause of COThe changes in BP, HR, and T should be closely observed intraoperatively. Because pituitrin can affect HR, BP, and body temperature  in the cIntraoperative dosing and injection of pituitrin should be improved. Clinically, the local injection of pituitrin is sufficient to enhance uterine contractions and reduce hemorrhage, but different doses of pituitrin may have different effects on the circulatory system  . InconsThe puncture needle for intraoperative administration of pituitrin should be improved. Most of the pituitrin puncture needles adopted in our hospital’s department were laparoscopic puncture needles, which were too long and had too thick a diameter. Thus, it was not easy to observe the blood return when puncturing the vessels, and the dose might not be accurate enough and affect the efficacy of the drug. A lumbar puncture needle or a common syringe (5–10 ml in size) could be used to puncture at 2–3 cm above the median mons pubis and reduce the pneumoperitoneal pressure before puncture to facilitate puncture and observation of blood return.Adequate emergency drugs and apparatus should be prepared before surgery. The commonly used cardiovascular emergency drugs such as epinephrine and atropine should be routinely prepared before surgery and diluted in proportion in advance so that these drugs can quickly be administered intravenously in the event of an accident. Apparatus such as defibrillators should be checked in advance for performance and be in a standby condition. For the three cases in the present study, the resuscitation progressed well and successfully due to the well-prepared drugs and apparatus.In conclusion, administering pituitrin in patients undergoing myomectomy might be a double-edged sword, possibly leading to decreased HR or even cardiac arrest in severe cases. These conditions might progress rapidly, making them critical. During the operation, close attention should be paid to the vital signs to identify changes in BP, HR, and the waveforms in the ECG.


Laser interstitial thermal therapy (LITT) at 1064 nm is widely used to treat epilepsy and brain tumors; however, no numerical model exists that can predict the ablation region with careful in vivo validation.In this study, we proposed a model with a system of finite element methods simulating heat transfer inside the brain tissue, radiative transfer from the applicator into the brain tissue, and a model for tissue damage.To speed up the computation for practical applications, we also validated P1-approximation as an efficient and fast method for calculating radiative transfer by comparing it with Monte Carlo simulation. Finally, we validated the proposed numerical model in vivo on six healthy canines and eight human patients with epilepsy and found strong agreement between the predicted temperature profile and ablation area and the magnetic resonance imaging-measured results.Our results demonstrate the feasibility and reliability of the model in predicting the ablation area of 1,064 nm LITT, which is important for presurgical planning when using LITT. AlthougIn vivo experimental evidence is preferred as tissue properties differ significantly between the ex vivo and in vivo stages.Numerical modeling of temperature profiles and ablation boundaries is necessary in presurgical planning and plays a key role in the development and application of LITT . In suchin vivo validation. Development of this model is highly desired for further use of 1,064 nm LITT in treating brain lesions.The 1,064 nm LITT is widely used to treat brain epilepsy and tumors owing to its advantages in deeper penetration and larger ablation zone , 27–30. in vivo experimental validation of the 1,064 nm LITT in brain tissue. The proposed numerical model is a system of finite element methods simulating heat transfer inside the brain tissue, radiative transfer from the applicator into the brain tissue, and a model for tissue damage. To quicken the computation for practical applications, we also validated P1-approximation as an efficient and fast method of calculating radiative transfer by comparing it with Monte Carlo simulation. Finally, we validated the proposed numerical model in vivo on six healthy canines, six human patients with epilepsy lesions who underwent 1,064 nm LITT and two human patients who underwent 980 nm LITT.We conducted this study to provide a numerical model with fast computing speed and 2.2.1.2.1.1.As shown in 2.1.2.To calculate the temperature distribution in brain tissue, we used the Pennes bio-heat transfer model , which i−3), c is the heat capacity of the brain tissue [J kg−1°C−1], k is the thermal conductivity of the tissue (W m−1°C−1), T is the tissue temperature (°C), t is the time (s), ρb is the blood density (kg m−3), cb is the specific heat capacity of the blood , Tb is the arterial blood temperature, which was 37°C in this study, Qr is internal heat source term due to photon absorption (W m−3), Qm is the metabolic heat generation per unit volume, and ∇. is the divergence operator.where ρ is the tissue density (kg cmm); the fourth term reflects heat generation owing to the absorption of laser photons (Qr). However, in LITT, the metabolic activity of the tissue is often ignored  is the fluence rate at position r along direction s, μa(r) is the absorption coefficient, μs(r) is the scattering coefficient, and P is the probability of a photon being scattered from position r’ to r. The integral in the equation represents the scattering of photons from all directions at position r’ to position r.To solve this equation using Monte Carlo simulation, transport of many photons through the tissue is simulated. At each step, the photon is either absorbed or scattered according to the probability distribution functions of absorption and scattering. The position and direction of the photon are updated after each scattering or absorption event until the photon either escapes the tissue or reaches a predefined depth.2.1.3.2.Compared with Monte Carlo simulation, P1-approximation is a simplified and computationally efficient method for solving laser diffusion simulation problems , 33. MonThe diffusion coefficient D [m] is then defined by−1] and μs [m−1], scattering anisotropy in the Henyey-Greenstein phase function (g). They are eigenfunctions of the Laplace operator in spherical coordinates.The absorption and scattering coefficients are denoted by μa  for the brain, E is the activation energy [J mol−1 K−1] for the brain, R is the universal gas constant [J mol−1 K−1], T is the Kelvin temperature [K−1], and t is the total heating time [s−1]. The tissue is assumed to be irreversibly damaged when where A is a pre-exponential factor [solecules , 35, 37.2.1.5.2.2.2.2.1.To evaluate the accuracy of the simulation model for the 1,064 nm LITT, we first tested the 1,064 nm LITT  on six canine brains with real-time MRI temperature monitoring. All animal studies were conducted in accordance with protocols approved by the Zhejiang University Animal Care and Use Committee (Ethics code: ZJU 20210094). Six canines underwent the procedure. A water-cooled diffusing tip laser applicator, laser power levels between 10 and 15 W, and irradiation times of 90–180 s were used.2.2.2.To further evaluate the accuracy of the LITT simulation model in humans, we tested the 1,064 nm LITT  on six patients and 980 nm LITT on two patients with focal epilepsy.2.2.3.via PRF-MRI into temperature changes, a special conversion formula is used:To monitor real-time temperature changes during LITT ablation, MRI was performed using the temperature-dependent proton resonance frequency (PRF) method, which has an accuracy of ±2°C in several tissue types , 39–41. where 0 ppm/°C , γ is th2, matrix size: 144 × 144, slice thickness: 5 mm, For the canine MRI experiments, the PRF-MRI was performed on a 3.0 T Skyra MRI  with the following imaging parameters: field of view: 210 mm2, matrix size: 160 × 160, slice thickness: 5 mm, and For the human MRI experiments, the PRF-MRI was performed on a 3.0 T 750 W scanner  with the following imaging parameters: field of view: 220 mm2.2.4.2, matrix size: 512 × 512, slice thickness: 0.6 mm, and effective TE: 410 ms. For the human experiments, CE T1-weighted MRI was acquired with a gradient echo sequence with field of view: 230 mm2, matrix size: 512 × 512, slice thickness: 0.6 mm, and time of repetition (TR)/TE: 8.2 ms/3.1 ms.Immediately after the ablation treatment, T2-weighted images or contrast-enhanced (CE) T1-weighted images  were acqTwo neurosurgeons with >15 years of experience in neurosurgery and neuroradiology assessed and measured all post-surgery MRI images to quantify the final ablation area induced by LITT. On the CE MRI, the maximum cross-section of the ablation volume was selected, and the ablation diameters were determined by the two neurosurgeons who were blinded to the simulation results to verify the results .2.3.p < 0.05 was considered significant.Paired comparisons, including those between the simulation and experimental results and between the P1-approximation and Monte Carlo simulations, were performed using paired Student’s t-tests to evaluate group differences or Pearson correlation analysis. 3.3.1.r = 0.989, p < 0.0001). Likewise, the 10-mm diffusion tip with 15 W power yielded a highly significant correlation . Paired Student’s t-tests revealed no statistically significant differences for either the 4-mm diffusion tip with 10 W power (p = 0.6590) or the 10-mm diffusion tip with 15 W power (p = 0.0864). The computation time for the Monte Carlo simulation and P1-approximation for one case were approximately 805.2 ± 19.7 s  and 3–4 s, respectively. Thus, P1-approximation well represented the Monte Carlo simulations of radiative transfer with a 230-fold reduction in computation time. In the following sections, all simulations were performed using P1-approximation.We first tested whether P1-approximation could well represent the Monte Carlo simulations of radiative transfer of 1,064 nm LITT, which can increase the computation speed in modeling radiative transfer. To compare the results from the two models, we plotted the light intensity values obtained from both models at different distances from the outer edge of the tube wall along the radial direction. P1-approximation showed great agreement with the Monte Carlo simulation at two different setups with different diffuser tip lengths (L1) and laser power . Further3.2.Next, we evaluated the accuracy of the temperature distribution models by comparing them with the experimental results obtained from MRT. Because the diffusion tip length and duration and power of the laser can be adjusted in clinical practice to better cover the targeted lesion, the numerical simulations were also set to mimic the experimental settings in via MRT, especially at higher temperature regions . We performed 180 s of ablation during the course of the experiment on canine 6. However, a brief interruption occurred at 120 s; therefore, we analyzed only the data for up to 120 s for canine 6. At 5.6 mm, the standard deviation and median of the temperature differences between the simulated and MRT-measured temperature time-course profiles were 1.26°C and 0.19°C (canine 1), 2.79°C and 1.39°C (canine 2), 5.2°C and − 3.69°C (canine 3), 2.26°C and 0.54°C (canine 4), 1.93°C and 0.39°C (canine 5), 1.97°C and − 0.59°C (canine 6), respectively. At 7.0 mm, these values were 1.33°C and − 0.71°C (canine 1), 3.23°C and − 0.74°C (canine 2), 2.54°C and − 0.79°C (canine 3), 1.11°C and 0.42°C (canine 4), 1.45°C and − 0.26°C (canine 5), and 1.54°C and − 1.18°C (canine 6). At 8.4 mm, these values were 1.15°C and 0.66°C (canine 1), 0.68°C and 0.33°C (canine 2), 1.03°C and 0.04°C (canine 3), 0.73°C and − 0.32°C (canine 4), 1.18°C and − 0.38°C (canine 5), and 1.21°C and − 0.61°C (canine 6). At 7.0 mm, our proposed model slightly underestimated the temperature compared with that of the MRT results in the first 60 s with the laser on, but such bias was generally small (<3°C in the six canine experiments).Next, we quantitatively compared the time-course profile of the temperature at different locations obtained from our proposed model and MRT measurements. For better visualization and generalization testing, we selected three locations at 5.6, 7.0, and 8.4 mm from the center of the fiber tip. The simulated temperature profile well approximated the measured ones 3.3.The ablation region predicted by our proposed simulation model was evaluated by comparing it with the ablation region obtained from post-LITT MRI. Here, the T2-weighted images were acquired immediately after LITT (~10 min after the laser was turned off) and used to determine the ablation region. p = 0.987). Additionally, in our proposed simulation model, the 980 nm laser always resulted in a smaller ablation diameter than did the 1,064 nm laser.via a human trial that included six epileptic patients were treated with 1,064 nm LITT (p = 0.648). In other two cases, patients were treated with 980 nm LITT, the prediction ablation boundary also showed good agreement with those drawn by the experienced neurosurgeons on the CE MRI. Furthermore, the variance in ablation area diameter between our proposed model and the measurements obtained from CE MRI was minimal, with a maximum difference of 1.56 mm  method, it has become the most widely used approach for monitoring water-based tissue temperatures , 45, 46.We also simulated the ablation region of the 980 nm laser and compared it with the results for the 1,064 nm laser. Compared with the 1,064 nm laser, the 980 nm laser ablation had a smaller but clear ablation range, partly due to the different absorption properties of laser wavelengths for water-based proteins and other tissues, resulting in an inconsistent temperature spread range and distribution pattern. The core temperature of the 980 nm laser ablation at the same power was higher, and the energy was more concentrated, resulting in sharper ablation boundaries. However, in the complex structure and high water inclusion area, based on the characteristics of 980 nm laser energy concentration, it will have better ablation form preservation and heating properties.These results have several implications for future use of the proposed numerical model. This approach allows adjusting the laser power and duration, thereby achieving the desired ablation size while minimizing associated risks. Among the critical factors to consider, selecting the laser wavelength is particularly important. Different laser wavelengths, such as 1,064 nm and 980 nm, exhibit distinct energy absorption properties within tissues, consequently resulting in variations in their respective thermal effects. For the ablation radius, the 980 nm laser is more suitable for target areas smaller than 15 mm, where precise ablation contour boundaries are required. Conversely, for lesions with ablation diameters >15 mm and necessitating stable control over ablation boundary diffusion, the 1,064 nm laser offers notable advantages.ex vivo experiments and in vivo animal studies is required to confirm the applicability of the model to different organs.As for the transfer of the ablation simulation model to other human organs, it is essential to recognize that different human organs possess different anatomical structures and characteristics. Therefore, the initial step involves a clear investigation of the specific anatomical structure of the target organ. Secondly, understanding the thermal properties of the target organ, such as its thermal conductivity and heat capacity, is crucial. This includes recognizing variations in vascular distribution among different tissues and disparities in blood perfusion. Next, optical properties also exhibit significant differences among various tissues, necessitating modifications in material properties. Finally, validation through in vivo validation is needed. First, changes in blood perfusion can significantly affect temperature distribution, and we assumed a constant blood perfusion in this study. Blood perfusion dissipates the heat generated during ablation, but changes in blood perfusion over time can alter the thermal properties of tissue, leading to inaccurate temperature predictions (in vivo experiments (This study had several limitations, and future work on the proposed numerical model and dictions . Second,dictions . Third, dictions . Fourth,dictions . After pdictions , 54. Fiferiments .5.in vivo validation. The proposed model uses a system of finite element methods to model heat transfer, radiative transfer and damage within brain tissue. To improve the computational efficiency, P1-approximation was validated and applied to radiative transfer calculations, saving significant time compared with that of Monte Carlo simulations. The numerical model was validated on six healthy canines and eight human patients with epilepsy, demonstrating excellent agreement between the predicted temperature profile and ablation region and MRI measurements. Furthermore, we compared the ablation regions of 980 nm and 1,064 nm lasers. The results revealed differences in diameter range, temperature spread, and ablation boundary sharpness between the two wavelengths. These findings highlight the importance of choosing an appropriate laser wavelength based on the desired ablation size and boundary accuracy. Future research should address the limitations identified herein and further validate the model through in vivo experiments.Here, we presented a numerical model for predicting the ablation region of 1,064 nm LITT, addressing the lack of a comprehensive model with zjm135@zju.edu.cn.Anonymized data not published within this article will be made available by request from any qualified investigator. Requests to access the datasets should be directed to The studies involving humans were approved by Human Research Ethics Committee, the Second Affiliated Hospital of Zhejiang University School of Medicine. The studies were conducted in accordance with the local legislation and institutional requirements. The participants provided their written informed consent to participate in this study.JiZ, PC, and RB conceived of the presented idea. PC, JuZ, and DS developed the theory and performed the computations. DS, ZZ, DL, and RB verified the analytical methods. All authors contributed to the article and approved the submitted version.


J Strength Cond Res 37(9): 1815–1820, 2023—The functional threshold power (FTP) field test is appealing for junior cyclists, but it was never investigated in this age category, and even in adults, there are few data on FTP collected in field conditions. Nine male junior road cyclists (16.9 ± 0.8 years) performed laboratory determination of maximal aerobic power (MAP), 4-mM lactate threshold (P4mM), critical power (CP), and the curvature constant (W′), plus a field determination of FTP as 95% of the average power output during a 20-minute time trial in an uphill road. The level of significance was set at p < 0.05. Outdoor FTP (269 ± 34 W) was significantly higher than CP (236 ± 24 W) and P4mM (233 ± 23 W). The −1·min−1) was significantly higher than the −1·min−1). Functional threshold power was correlated, in descending order, with MAP (r = 0.95), P4mM (r = 0.94), outdoor and indoor r = 0.93 and 0.93, respectively), CP (r = 0.84), and W′ (r = 0.66). It follows that in junior road cyclists, the FTP field test was feasible and related primarily to aerobic endurance parameters and secondarily, but notably, to W′. However, the FTP field test significantly exceeded all laboratory performance tests. When translating laboratory results to outdoor uphill conditions, coaches and sport scientists should consider this discrepancy, which may be particularly enhanced in this cycling age category.Vinetti, G, Rossi, H, Bruseghini, P, Corti, M, Ferretti, G, Piva, S, Taboni, A, and Fagoni, N. The functional threshold power field test exceeds laboratory performance in junior road cyclists.  All tests were conducted at least 24 hours apart and concluded within 15 days.A junior road cycling team performed (a) an indoor incremental test for the determination of PNine male junior road cyclists  were calibrated with ambient air, with the use of a CO2 scrubber sodium and calcium hydroxides and with a mixture of known gases  and the turbine flowmeter by means of a 3-L syringe as per manufacturer's guidelines. Capillary blood lactate concentration ([La−]) was assessed by an electroenzymatic method  in 0.3 μL samples taken from the earlobe.In all tests, subjects were on their own road bicycle equipped with a crank-based power meter , wearing a portable metabolic cart  and a heart rate (HR) monitor . The 30-second rolling average of oxygen consumption  between 2 and 6 hickness  were recnd [La−] , whereaslim between 12 and 20 minutes  with an average gradient of 6.7% Figure , on anotlim and PO for biological and technological reasons, respectively  was used.Variables and parameters were compared by means of Bland-Altman plots and reduced major axis linear regression analysis. Values are reported as mean ± standard deviation. The power-duration relationship was fitted by means of geometric mean nonlinear regression method to account for the presence of heteroscedastic random error both in Tectively ,39. Repery large .The leve. Incremental test peak values of PO, HR, and [La−] were 309 ± 30 W, 198 ± 5 beats·min−1, and 11.0 ± 1.7 mM, respectively. −1·min−1), and extrapolated MAP was 276 ± 28 W. Peak p = 0.48 and 0.50, respectively). There was a significant effect of the laboratory test on peak HR and [La−] . Peak HR of the 3 shorter constant-power trials was moderately lower than the incremental test (d = 0.8–1.2), and peak [La−] of the 2 longest constant-power trials was largely lower than the incremental test and the shorter constant-power trial (d = 1.6 for both). Post–warm-up [La−] was 1.6 ± 0.7 mM. Critical power and W′ estimates were 236 ± 24 W and 20.7 ± 4.7 kJ . P4mM was 233 ± 23 W, not significantly different from CP  and in fair agreement with it . Critical power and P4mM were in good correlation with MAP  and with r = 0.90 for both).A graphical summary of laboratory and field tests is presented in Figure 2−] was 2.0 ± 0.6 mM (p = 0.11 vs laboratory). Subjects adopted a positive pacing strategy, with a decrease of −2 ± 1 W·min−1 in PO throughout the TT. Mean PO of the TT (283 ± 35 W) significantly outperformed the prediction calculated from the indoor power-duration relationship , and FTP resulted 269 ± 34 W . Correlations and agreements are shown in Figure 4mM (p < 0.001 and d = 1.2 for both) although in good correlation with them (r = 0.84 and 0.94 respectively). Functional threshold power was not significantly different from MAP  and highly correlated with MAP (r = 0.95) and indoor and outdoor r = 0.92 and 0.93, respectively). Moreover, FTP was modestly correlated with W′ (r = 0.66).Post–warm-up  higher than in laboratory conditions, although the HR response may have been attenuated by the lower air temperature in the field by a reduced cardiovascular drift (Strikingly, contrary to laboratory studies in adults, the outdoor 20-minute TT outperformed laboratory tests. The fact that field FTP was essentially equal to laboratory MAP Figure  is to bear drift . Cyclingar drift , despitekinetics ,12. Thers indoor  and in us indoor ,34, in ps indoor ,6,25. Res indoor , althougV˙O2peak  and its boratory . These fW′. However, absolute performance was significantly higher in outdoor uphill conditions with respect to the laboratory.W′. However, outdoor uphill conditions where the FTP test took place were responsible for a significant improvement in PO with respect to laboratory predictions. Specifically, outdoor 20-minute mean PO was 12% higher than that predicted from the indoor power-duration relationship, and the resulting FTP was 14–15% higher than indoor CP and P4mM. When translating laboratory results to outdoor training-intensity prescription or performance predictions, coaches and sport scientists should consider these discrepancies, which may be particularly enhanced in this cycling age category.In junior road cyclists, FTP was closely related to aerobic endurance parameters and partially also to In conclusion, in junior road cyclists, the FTP test resulted feasible and related primarily to aerobic endurance parameters and, secondarily but notably, to The authors are grateful to Marco Sbragi of Gobat Srl, Foiano della Chiana, Arezzo, Italy, for his technical assistance. The authors have no conflicts of interest or founding sources to disclose.


In the current study, none of the tested quinolones (QNs) at any of the examined concentrations were lethal for common duckweed plants. However, at the highest concentration (12.8 mg × L−1), LVF increased Ir and Iy values by 82% on average and increased the values of NAL, PEF and MOXI by 62% on average. All tested QNs led to the loss of assimilation pigments. In consequence, all QNs, except for LVF, induced changes in chlorophyll fluorescence (Fv/Fm), without any effect on phaeophytinization quotient (PQ) values. The uptake of NAL, MOXI, LVF by Lemna minor during the 7-day chronic toxicity test was directly proportional to drug concentrations in the growth medium. Nalidixic acid was absorbed in the largest quantities, whereas in the group of fluoroquinolones (FQNs), MOXI, LVF and PEF were less effectively absorbed by common duckweed. This study demonstrated that biosorption by L. minor occurs regardless of the plants’ condition. These findings indicate that L. minor can be used as an effective biological method to remove QNs from wastewater and water and that biosorption should be a mandatory process in conventional water and wastewater treatment.Plant growth and the development of morphological traits in plants are inhibited under exposure to pharmaceuticals that are present in soil and water. The present study revealed that moxifloxacin (MOXI), nalidixic acid (NAL), levofloxacin (LVF) and pefloxacin (PEF) at concentrations of >0.29, >0.48, >0.62 and >1.45 mg × L Pharmac−1 ,23. Amon−1 [QNs are used in the treatment of infectious diseases in human medicine ,25. Nali−1 . Wastewa−1 . Only su−1 . Due to −1 . One of −1 . In gene−1 .The following quinolone antibiotics were analyzed in this study:First generation QNs—nalidixic acid (NAL)—naphthyridine derivative;Second generation QNs—pefloxacin (PEF)—fluoroquinolone (FQN);Third generation QNs—levofloxacin (LVF)—FQN;Fourth generation QNs—moxifloxacin (MOXI)—FQN.L. minor), which is an important link in the food chain in freshwater environments . The current assessment of toxicity is based mainly on the analysis of common duckweed growth parameters: growth rate, yield and weight. However, they do not include research on the assessment of plant productivity at the biochemical level, after exposure to toxic substances, as well as their specific response mechanisms [The aim of this study was to determine the effects of exposure to four antibiotics: NAL, PEF, LVF and MOXI, popular drugs in common duckweed  plants used in the study were obtained from the collection of the Department of Chemistry of the University of Warmia and Mazury in Olsztyn, Poland.Common duckweed  in a 16 h light/8 h dark cycle (mean maximum temperature of 20 °C during daytime and 16 °C during night time) for 7 days. The biotest was validated in accordance with OECD guidelines: in the temperature 24 ± 2 °C and pH at 6.5 ± 0.2, the doubling time of frond number in the control was less than 2.5 days (60 h), corresponding to approximately a seven-fold increase in seven days and an average specific growth rate of 0.275 d−1 [−1) were determined based on the percent inhibition of growth rate (Ir), percent reduction in yield (Iy), percent reduction in dry weight (IDW), Chl a and Chl b, TCC, PQ of tissues, Fv/Fm, and the fresh (FM) and dry matter content (DM) of 100 duckweed plants. Frond area was measured using the Lucia 5.0 program . The following formulas were applied to calculate the values of Ir and Iy based on the number of duckweed fronds and frond area according to OECD guidelines [DW:i–j—average specific growth rate in time i to j;μi—measurement variable in the test or control vessel at time i;Nj—measurement variable in the test or control vessel at time j;Nt—time from i to j;c—mean value of µ in the control group;μT—mean value of µ in the treatment group;μc—final number of duckweed fronds and frond area minus the initial number of duckweed fronds and frond area in the control group;bT—final number of duckweed fronds and frond area minus the initial number of duckweed fronds and frond area in the treatment group.bDW—percent reduction in dry weight;%Ic—dry weight of duckweed fronds in the control group (mg);DWt—dry weight of duckweed fronds in the treatment group (mg).DWCommon duckweed was grown in 50 mL of OECD medium for testing chemicals  in a pla.275 d−1 . All solidelines , and thea, Chl b and TCC were determined in leaf extracts prepared with 96% (v/v) aqueous ethanol and centrifuged at 12,000× g for 15 min. The supernatant was separated; the pigments were quantified spectrophotometrically  according to [a and Chl b, TCC and PQ:The contents of Chl rding to , and therding to . The fola, Chl b—chlorophyll content in plant material (µg × mg−1);Chl e—volume of ethanol extract (mL);Vx—dilution coefficient;m—sample weight (mg);Chl a—concentration of Chl a in the extract (µg × mL−1);CChl b—concentration of Chl b in the extract (µg × mL−1);Ci—solution absorbance at ith wavelength.A−2 × s−1), and minimum chlorophyll fluorescence (Fo), maximum chlorophyll fluorescence (Fm) and variable chlorophyll fluorescence (Fv = Fm − Fo) were determined. The Fv/Fm of PSII was determined based on the chlorophyll fluorescence kinetics of common duckweed.The Fv/Fm of PSII (Photosystem II) was measured with the HandyPEA chlorophyll fluorescence system . Common duckweed leaves were placed in a leaf clip and stored in the dark for 30 min to quench chlorophyll fluorescence. After dark adaptation, chlorophyll was excited at light intensity of 2500 (µmol × mE) of adsorbate solutions was measured with the Hitachi UV/VIS U-1800 spectrophotometer . The absorption spectra of QNs were set at 191–800 mm to determine the maximum absorption (maxλ) of the analyzed solutions with a concentration of 1.6 mg × L−1 each. The maxλ values for quantitative measurements were determined at: 210 and 333 nm for NAL, 274 nm for PEF, 290 nm for LVF and 287 nm for MOXI.The extinction (E = f(c) were plotted for each adsorbate solution with the use of the previously determined maxλ values. The Bs process was analyzed by placing 100 duckweed plants in 50 cm3 of adsorbate solution and stirring flask contents manually only once. The extinction of the analyzed solutions was determined before placing the plants in the solution (E0) and after 168 h (7E). After the measurement, 2 mL of the adsorbate solution was returned to the flask. The difference (C0 − C7) was the adsorption of a given adsorbate on the analyzed adsorbent.The calibration curves of absorption vs. concentration Biosorption was calculated with the following formula:−1);Bs—biosorption ;C7—concentration of adsorbate after 168 h (mg × L−1);CV—solution volume (mL);m—adsorbent mass (mg).p < 0.01. The results of the experiment  of common duckweed were processed in the STATISTICA 13.3 statistical package . The number and total area of fronds were used as the endpoints for assessing phytotoxicity [x) were analyzed with a selected regression model to calculate the concentrations at 20% and 50% response levels.The experiment was conducted in six replicates. The results were expressed as means ± standard deviation (SD). Data were processed statistically by two-way analysis of variance—ANOVA (F test) . The exptoxicity ,43,44. E−1) in the growth medium on the Ir, Iy, IDW, FM and DM, Chl a, Chl b, TCC, Fv/Fm and the PQ of common duckweed tissues was evaluated.The impact of increasing NAL, PEF, LVF and MOXI concentrations  of NAL, PEF, LVF and MOXI, the Ir of duckweed plants was determined at 0.45, −18.81, 5.96 and 21.06%, respectively. The highest concentration of the tested drugs (12.8 mg × L−1) significantly increased Ir values by 58.47 and 81.57% in plants exposed to PEF and LVF, and insignificantly by 62.54 and 59.41% for NAL and MOXI , the most toxic was MOXI (Iy = 27.81%), whereas at the highest concentration (12.8 mg × L−1), it was LVF (Iy = 81.78%)  B.20 = 0.48 mg × L−1, EC20 = 1.45 mg × L−1, EC20 = 0.62 mg × L−1 and EC20 = 0.29 mg × L−1, respectively. NAL, PEF, LVF and MOXI applied at concentrations of 3.27 mg × L−1, 5.99 mg × L−1, 2.67 mg × L−1 and 3.98 mg × L−1 (EC50), respectively, increased the Ir and Iy of common duckweed by 50%   of NAL, whereas in plants exposed to PEF, the same (the highest for PEF) effect was observed only at a concentration of 3.2 mg × L−1. In plants treated with the highest concentration (12.8 mg × L−1) of NAL, IDW values significantly reached 24.07%. Other drugs were less phytotoxic. Plants exposed to PEF, LVF and MOXI were characterized by insignificant IDW of 8.97, 5.09 and 3.92%, respectively  . Nalidixectively C. Nalidimg × L−1 .−1), excluding plants that were grown in the presence of LVF, where FM was somewhat higher at 324.33 mg.Fresh mass (100 duckweed plants) was analyzed after 7 days of exposure to QNs. The type and concentrations of antibiotic significantly influenced the FM of duckweed . The FM −1) concentration, respectively (20) when applied at a concentration of 1.09 mg × L−1, which was not observed under exposure to the remaining antibiotics . The lowest concentrations of NAL, PEF and MOXI were somewhat less toxic for photosynthetic pigments. When the tested drugs were applied at the highest concentration (12.8 mg × L−1), the greatest significant reduction in the content of Chl a, Chl b and TCC  was observed under the influence of MOXI. Nalidixic acid was least toxic by 14% on average for Chl a, Chl b and TCC and the highest concentration of NAL (12.8 mg × L−1) (a levels in plant tissues were reduced by 20% (EC20) in the presence of 0.63, 0.55, 0.61 and 0.58 mg × L−1 of NAL, PEF, LVF and MOXI, respectively, in the growth medium, as well as for the content of Chl b at 0.56, 0.50, 0.49, 0.57 mg × L−1, respectively. Moxifloxacin was the only drug that reduced TCC by 20% (EC20) at a concentration of 0.90 mg × L−1 (The leaves of control plants (grown in a medium without antibiotics) contained 1.26 mg × g−1 of FM A–C. The −1 of FM . The cong × L−1) A–C. Chlomg × L−1 . None ofmg × L−1 D.−1), the Fv/Fm ratio was 6%, 10%, 4.5% and 7% insignificantly lower, respectively, than in the control  at the lowest concentration of this drug (0.4 mg × L−1), and it reached 5.19 mg × g−1 DM at its highest concentration (12.8 mg × L−1). The biosorption of MOXI was somewhat lower, and it was determined at 0.16  and 4.55 mg × g−1 DM  in plants exposed to 0.4 and 12.8 mg × L−1 of MOXI, respectively. Levofloxacin was not absorbed when supplied at the lowest concentration (0.4 mg × L−1).The uptake of NAL, MOXI, LVF by common duckweed during the 7-day chronic toxicity test was directly proportional to drug concentrations in the growth medium. Nalidixic acid was the most readily absorbed compound. Its content in common duckweed tissues was 0.72 mg × g−1), and its uptake significantly increased in concentrations of LVF ≥ 3.2 mg × L−1. The LVF content of duckweed plants peaked at 1.31 mg × g−1 DM under exposure to 6.4 mg × L−1 LVF. In contrast, rising concentrations of PEF almost did not affect its biosorption, which was determined at 0.67 mg × g−1 DM on average in all samples (The uptake of this drug began when LVF concentration was doubled (0.8 mg × L samples .a) and chlorophyll b (Chl b) content, total carotenoid content (TCC), phaeophytinization quotient (PQ), maximum quantum efficiency (Fv/Fm) and quinolone biosorption (Bs) by duckweed plants. The lowest concentration of PEF (0.4 mg × L−1) stimulated (by 17% on average) the Ir and Iy of common duckweed plants, whereas NAL, LVF and MOXI applied at this concentration were phytotoxic. Moxifloxacin was the strongest inhibitor of plant growth and yield, and it increased Ir and Iy values by 24% (−1), LVF significantly increased Ir and Iy values by 82% on average, and NAL, PEF and MOXI by 62% on average (90) by 40 µM of tetracycline, which is equivalent to 18 mg × L−1 of tetracycline [−1) of MOXI, a synthetic antibiotic, induced the greatest significant decrease (36% on average) in the content of assimilation pigments , which is widely used as a model organism in biological research, the application of tetracycline, a natural antibiotic, did not induce a chloroplast-specific stress response because the expression of chloroplast-specific chaperones (PsaB and PsbA) was unaltered. Tetracycline is phytotoxic, as it induces the expression of mitochondrial stress genes, impairs mitochondrial translation and inhibits the function of mitochondria, which generate most of the cell’s supply of adenosine triphosphate (ATP) [−1) induced changes in chlorophyll fluorescence. Exposure to PEF led to the greatest decrease in the Fv/Fm ratio (7.5% on average) relative to control. Moxifloxacin and NAL were somewhat less toxic, and they decreased the Fv/Fm ratio by 6.5% on average. Common duckweed plants responded completely differently to LVF. The average Fv/Fm ratio of plants grown in the presence of LVF was nearly identical to that noted in control plants (0.78) (Pinus sylvestris L.) needles and European beech (Fagus sylvatica L.) leaves decreased in soil contaminated with diesel oil. Scots pines adapt (grow more rapidly and produce higher biomass) to long-term soil contamination with diesel [−1). However, their growth was inhibited by 20% (EC20) in response to a PEF concentration of 1.45 mg × L−1  to a different degree. Chlortetracycline exerted a bi-directional effect on plant growth, where low concentrations of the drug promoted growth, whereas high concentrations inhibited growth and photosynthesis. While the common duckweed accumulates tetracycline, it accumulates biogenic amines such as: tyramine, putrescine, cadaverine, spermidine and spermine. The amines spermine and tyramine were most sensitive when the plants were treated with this drug [Lemna minor and Lemna minuta [Plant growth and the development of morphological traits in plants are inhibited under exposure to pharmaceuticals that are present in soil and water ,46,47,48s by 24% A,B. Low  average A,B. Baseacycline . Similaracycline ,51, and acycline . A glyphpigments A–C. Meante (ATP) . In thiss (0.78) . Chloroph diesel . In the mg × L−1 . The lowectively . Accordihis drug . In addihis drug , and expa minuta .−1 DM at the lowest concentration (0.4 mg × L−1) and 5.19 mg × g−1 DM at the highest concentration (12.8 mg × L−1) (DW and FM by 20% (EC20) at concentration of 1.02 mg × L−1 on average, which was not observed under exposure to the remaining antibiotics  . Nalidixibiotics . Howeveribiotics A,C. Accoibiotics . Howeverta algae , whereastibiotic . Therefotibiotic . Common tibiotic ,43.The results of this study confirm that the Lemna test is a useful analytical tool for evaluating the toxicity of QNs and for predicting the consequences of the chemical contamination of freshwater bodies based on the calculated values of toxicity indicators. The presence of the tested QNs in water triggered negative morphological and physiological responses in duckweed, which suggests that these compounds not only decrease its populations but also impact other trophic levels in the aquatic food webs.−1), LVF increased Ir and Iy values by 82% on average, and NAL, PEF and MOXI by 62% on average. The uptake of NAL, MOXI, LVF by common duckweed during the 7-day chronic toxicity test was directly proportional to drug concentrations in the growth medium. Nalidixic acid was absorbed in the largest quantities, whereas in the group of FQNs, MOXI, LVF and PEF were less effectively absorbed by common duckweed. Common duckweed removes QNs from the solution by active rather than by passive biosorption (dead biomass), and it can transform the accumulated pollutants by internal processes, thus reducing their concentrations in the environment.In the current study, none of the tested QNs at any of the examined concentrations were lethal for common duckweed plants. However, at the highest concentration (12.8 mg × LThe research demonstrated that biosorption by common duckweed occurs regardless of the plants’ condition. In addition, plants that absorb contaminating substances can be easily removed from the solution and neutralized. Even compromised plants removed QNs from the growth medium to a varied extent. These findings indicate that common duckweed can be used as an effective biological method to remove QNs from wastewater and water, and that biosorption should be a mandatory process in conventional water and wastewater treatment.


Helix aspersa Müller snail flesh. H. aspersa Müller snail flesh exhibits a high nutritional content, a good ω3/ω6 ratio and higher levels of unsaturated fatty acids. Various minerals have been found in the flesh of H. aspersa Müller. Around 76.91 kcal, or 3.84% of the energy of a daily meal of 2000 kcal, are present in 100 g of this flesh. The evaluation of the antioxidant capacity indicated that the flesh’s extracts contained a large quantity of antioxidant biomolecules. Administration of the aqueous extract of H. aspersa Müller flesh didn’t cause death in laboratory rats, indicating that the lethal dose 50 is greater than 2000 mg·kg−1 body weight. The consumption of the flesh of H. aspersa Müller is highly recommended for human consumption due to its high concentration of nutrients and essential elements, as well as unsaturated fats, and due to its safety.Humans consume snail flesh as part of their diet. To assess its nutritional value and toxicity, chemical analyses were conducted to confirm the presence of protein, total and reduced carbohydrates, fat, fatty acid composition and mineral components. Furthermore, an acute toxicity study was carried out to determine the safety of  Inexpensive natural resources are used for their medicinal virtues in the prevention or treatment of a variety of pathologies. Most people believe that natural products are non-harmful and safe . NaturalMollusks are the second-largest branch of the animal kingdom, with around 100,000 species , a , a 34], H. aspersa Müller flesh would be required for calcium, 199.69 g for magnesium, 129.87 g for zinc, 105.26 g for copper, 145.99 g for iron, 185.19 g for manganese, and 121.25 g for phosphorus. Thus, 100 g/d of dry H. aspersa Müller flesh offers 77% of the RDI for zinc, 54% of the RDI for calcium, 50% of the RDI for magnesium, 68% of the RDI for iron, 59% of the RDI for copper, and 82% of the RDI for phosphorus.To reach the RDI, 184.98 g of dried H. aspersa Müller is shown in H. aspersa Müller. The unsaturated fatty acid content was 50.78%, and this ratio was higher than those of saturated fatty acids. The major fatty acids were arachidonic acid (10.67%), palmitic acid (7.11%), stearic acid (16.86%), oleic acid (7.03%), linolenic acid (5.56%), 11.14-Eicosadienoic acid (5.42%), linoleic acid (14.16%), and 34% was composed of saturated fatty acid.The fatty-acid profile of 18:0), followed by palmitic acid (C16:0) which made up 49.59 and 20.91% of the total saturated fatty acid content, respectively. Oleic acid (C18:1) was determined to be the primary monounsaturated fatty acid , while linoleic acid (C18:2) was determined to be the primary polyunsaturated fatty acid .However, 10.33 and 40.45%, respectively, of the total acids were MUFA and PUFA. The main saturated fatty acid was stearic acid (CH. pomatia were C16:0 (6%), C18:0 (11%), C18:1 (14%), C18:2 (20%), and C20:2 (11%), while the main fatty acids of H. memoralis were C16:0 (6%), C18:0 (13%), C18:1 (15%), C18:2 (19%), and C20:2 (13%). Özogul et al. [H. pomatia 21 fatty acids with 12% C16:0, 19% C18:0, 7% C22:0, 17% C18:1, 16% C18:2 and 10% C20:2.According to published research in the literature, Milinsk et al.  reportedl et al.  discoverH. aspersa Müller is nevertheless high.A helpful metric for assessing the relative nutritional benefits of dietary oils has been proposed, namely, the ω3/ω6 ratio. Numerous studies have linked a greater ratio of ω3/ω6 to improved nutritional value . The daiH. aspersa Müller, in which Özogul et al. [The ω6/ω3 ratio was reported to be, at most, 4.0 ; this ral et al.  observedThe potential advantages of MUFAs in reducing the risk of cardiovascular disease have been studied by many researchers . In thisFrom these results, we can conclude that PUFAs are essential to human physiology and should be included in the diet, perhaps in the form of snail flesh, which is a particularly good source of these fatty acids.H. aspersa Müller revealed interesting information on its composition. The analysis showed that reducing sugars were present at a concentration of 2.92 ± 0.04 mg·g−1. Phenolic compounds, known for their potential health benefits, were present in average quantities of around 1.467 ± 0.077 mg·g−1, while the flavonoid content was 0.132 ± 0.002 mg·g−1.The aqueous extract has an extraction yield of approximately 6.38%. Comprehensive biochemical analysis of the aqueous extract of H. aspersa was shown to contain 0.132 mg·mL−1 of polyphenols [On the other hand, these results are consistent with previous research. In one particular study, the crude extract of yphenols . This cr−1 DM, indicating that the flesh of H. aspersa could be a natural source of antioxidants. Other results have been reported by Gogas et al. [H. aspersa Müller (0.587–0.715) fed with different protein sources under intensive rearing conditions.The results showed that the total antioxidant activity of HAAE was 9.628 ± 0.010 mg AAE·gs et al.  in H. asH. aspersa Müller could be useful for pharmaceutical applications, and further study of the antioxidant response compounds and their characterization is needed.The total antioxidant activity was evaluated by creating a green phosphate complex with an acidic ionic nature; this indicates that HAAE is a good source of natural antioxidants. Indeed, −1) was found to be between 36.07 ± 0.5 and 89.35 ± 0.52%. The maximum inhibition was observed at the highest concentration of 1000 μg·mL−1 of HAAE, while the scavenging activity of L-ascorbic acid was 89.46% at a concentration of 200 μg·mL−1. The reduction in absorbance of the DPPH radical is induced by antioxidants because of the reaction between the antioxidant and the radicals, which results in the recovery of the radical by the donation of hydroxyl.The DPPH radical scavenging activity was evaluated by quantifying the inhibition of the DPPH radical by the extract. The effect of HAAE on hydroxyl-radical-induced oxidative damage at different concentrations   . Althoug µg·mL−1 .H. aspersa Müller strongly depends on the presence of reducers, which have antioxidant potential by disrupting free radical chains to generate hydrogen atoms. Higher concentrations of antioxidants become pro-oxidants and lead to higher antioxidant activity, so dose dependence can be very significant in the application of antioxidants. However, even at low concentrations, the DPPH radical scavenging capacity of ascorbic acid was superior to that of the tested samples.According to Zou et al. , the redIn this study, HAAE had potential activity in scavenging hydroxyl radicals at different concentrations. Natural polyphenols generally play a crucial role in natural medicines, and may be responsible for increased free-radical scavenging activity by increasing the levels of phenolic and polyphenolic compounds .3+ to Fe2+. The quantity of Fe2+ can then be determined by detecting the development of Perl’s Prussian blue at 700 nm. The HAAE demonstrated good ferrous-reducing activity in this research, with an IC50 value of 42.58 ± 1.64 µg·mL−1; however, the reducing power rises with the increasing concentration of the HAAE. The reduction activity was compared to ascorbic acid as a reference antioxidant (53.253 ± 2.277 µg·mL−1), and there was a significant difference (p ≤ 0.05) between the analyzed extract and ascorbic acid.The antioxidant activity of the HAAE was measured by quantifying its capacity to transfer an electron to convert FeH. aspersa Müller’s water extract may work similarly, providing electrons and interacting with radicals to transform them into more stable products and terminate the chain reaction of free radicals.The existence of reductions, which have antioxidant potential by disrupting the chain of free radicals by giving a hydrogen atom, determines the ferrous reduction capacity . The chePhyllanthus species, in which the lowering ability was proportionate to the amounts. Subavathy and Janet [−1 concentrations of 73.35, 95.36, and 87.5% in T. brunneus, C. annulus, and B. spirata, respectively, and minimum activity at 100 µg·mL−1 concentrations of 52.07, 67.21, and 57.51% in T. brunneus, C. annulus, and B. spirata, respectively. At the maximum quantity (100 µg·mL−1) of O. macrocera, a greater reducing capability was found [Kumaran and Karunakaran  found sind Janet  discoveras found .H. aspersa Müller flesh is presented in 50 is greater than 2000 mg·kg−1. Consequently, no signs of toxicity were observed during the first four hours following administration, with no decreased sensitivity to stimuli (pain and noise), decreased mobility, softening of feces, alteration of behavior, loss of body weight, or death registered for 14 days.The research we conducted on the toxicity of HAAE is classified as a fifth-category substance according to the OECD Globally Harmonized System of Classification and Labelling of Chemicals (GHS), rated as non-toxic by the oral route . AccordiThe effects associated with daily oral administration of repeated doses of HAAE were closely scrutinized. However, evaluation of behavioral parameters and weight gain showed no changes in the aforementioned parameters.Observation of behavior throughout the study period showed that regardless of the dose administered, no behavioral changes were observed during the experimental period. As shown in H. aspersa maxima, H. aspersa Müller, and H. pomatia appeared non-toxic, and that the mice remained healthy throughout the experiment. Kouachi et al. [H. aspersa crude extract was not toxic.In agreement with the other studies, Danilova  found thi et al.  found thCaetano et al.  suggest −1 for lead, mercury, and cadmium, respectively. It is important to note that the tests conducted in our study adhered to a meticulously established methodology. The snails used in the experiments were obtained under controlled laboratory conditions, thereby ensuring the lack of any external contamination. An interesting opportunity for further exploration is in the examination of the potential toxicity shown by indigenous snail species inhabiting regions characterized by industrial or agricultural activities.Several scientific research efforts have shown that land snails exhibit sensitivity towards the accumulation of heavy metals and other contaminants, making these substances suitable candidates for bio-indicators ,33,54. TH. aspersa Müller snails were gathered during the spring season in the Moulay Bousselham region of Morocco. These were immediately moved to the laboratory, where healthy individuals were housed in rectangular plastic boxes (24 × 32 × 12 cm) with a sponge, moist soil, and food . Crates holding 50 snails were sprinkled with water every day to keep them wet. This experiment included shells weighing 10.44 ± 3.11 g and measuring 3.35 ± 0.48 cm in length.Uninfected We should mention that the snails were treated as part of a scientific experiment that followed animal care guidelines.150 snails were cleaned with running water, and then stripped of their shells; the visceral masses were eliminated. The feet (with the head) were recovered and cleared of slime with a NaCl solution (1.2%) and were washed with distilled water and dried on absorbent paper. The samples were dried in an oven (60 °C for 48 h), then ground and sieved for recovery.The method used for moisture determination was based on the loss of sample mass until a constant mass was obtained at 105 °C .The determination of the percentage of dry matter (DM) involves placing the sample in an oven at a temperature of 70 °C until a variation of at least 3 mg is obtained between two consecutive weighings performed separately with a 2-h interval .The pH measurement was determined using the method described in . In addiConsistent with ISO 936, total ash from snail samples was evaluated using the gravimetric technique at 550 °C. The results were represented as a percentage  .H. aspersa Müller flesh [The AOAC method (928.08) was used to estimate the average total protein in er flesh . Results−1 wet weight [The Soxhlet technique was used to calculate the mean lipid content, which was expressed in g·100 gt weight .H. aspersa Müller was calculated by deducting the amounts of fat, protein, and ash from the dry-matter value [Following Equation (1), the amount of carbohydrates in the flesh of er value .Carbs  and filtered. This extract was used for mineral determination. The determination of the trace elements studied was carried out by inductively coupled argon plasma atomic emission spectrometry  8 L/min, auxiliary gas flow rate (Ar) 0.2 L/min, axial view size, copying and playback time 45 min, and copying time 15 min) were used to determine the trace elements studied. H. aspersa Müller flesh was determined colorimetrically.Because phosphorus is found in organic molecules, oxidizing the organic material to liberate the phosphorus requires a digestion or calcination procedure. Therefore, using AOAC Method 965.17 , the tot−1), the mixture was violently agitated. The top layer was transferred to bottles once the layers had been separated.A modified version of the transmethylation process from Milinsk et al.  was used−1 for 18 min until it reached 260 °C. The BR-5ns FS capillary column (30 m × 0.25 mm ID × 0.25 m) was utilized. In undivided mode, the injection volume of helium is 1.0 mL·min−1. The whole analysis took 100 min. The mass spectrometry detector (MSD) was set to electronic impact ionization mode, with an ionizing energy of 70 eV and an m/z scan range of 50 to 500.The fatty-acid methyl esters were analyzed by gas chromatography–mass spectrometry under the following conditions: the injector port temperature was 250 °C., the starting oven temperature was 40 °C, and the temperature was gradually increased at 8 °C·minThe temperature of the ion source was 230 °C, and it then quadrupled to 150 °C. The electron multiplier voltage (EM voltage) was kept at 1100 V above the self-regulatory level, with a 3 min solvent delay.Ten grams of snail flesh flour were mixed with 100 mL of distilled water and subjected to maceration for 48 h. The resulting mixture was filtered using filter paper, and the filtrate volume was evaporated using a rotary evaporator and dried at 60 °C until the weight stabilized. The obtained crystals were crushed using a mortar, and the resulting powder was stored in a sterile glass jar hermetically sealed and kept refrigerated at 4 °C. The extraction yield (EY) was calculated relative to the weight of the dry material using the following equation, Equation (2):3) method [The total phenol content of HAAE was determined by the Folin–Ciocalteu method reported by Zargoosh et al. , with so) method .The quantification of reducing sugars was based on the use of Di-Nitro-Salicylic Acid as a reagent according to the method of Negrulescu et al. .Total antioxidant capacity (TAC)−1 DM) [In this assay, 50 µL of HAAE is mixed with 2000 µL of reagent . The reaction mixture was then incubated at 95 °C for 90 min, cooled to room temperature and the absorbance at 695 nm was recorded against a blank. L-ascorbic acid was used as a reference antioxidant. The total antioxidant capacity of the extract was expressed as ascorbic-acid equivalent per gram of dry matter (mg EAA·g−1 DM) .Free radical scavenging activity (DPPH)The absorption of DPPH radicals at characteristic wavelengths is followed by a decrease in optical density. From a series of tubes, 500 µL of ethanol solutions containing different concentrations of extracts were taken, and 2500 µL of DPPH solution (0.2 mM) was added to all tubes. The contents were shaken well and allowed to react at room temperature for 30 min. The absorbance of the samples was recorded at 517 nm against the control. A negative control was made by mixing 500 µL of methanol and 2500 µL of DPPH solution, and the methanol was used as a blank in parallel with the test sample .Ech: absorbance of the sample and Abs Cn: absorbance of negative control.The percentage inhibition was calculated as follows in Equation (3):Ferrous-ion-reducing antioxidant power assay (FRAP)w/v), were allowed to react at 50 °C for 20 min. Then, 2500 µL of TCA  were added and the mixture was centrifuged . 2500 µL supernatant, 500 µL ferric chloride  and 2500 µL distilled water were mixed. The absorbance was determined at 700 nm relative to negative control. The same procedure was followed for the standard solution of L-ascorbic acid. The reduction of activity can be calculated using the following analysis, Equation (4):The formation of the ferrous form of the iron/ferricyanide complex was measured by the formation of pearly Prussian blue at 700 nm . 200 µL H. aspersa Müller at 500, 1000, and 2000 mg·kg−1, respectively, while the control group received distilled water at 10 mL·kg−1. Behavioral observation was conducted 1 h after administration.The ethical institutional committee for the Faculty of Sciences, University Ibn Tofail, Kenitra, Morocco, approved the protocol. All of the experimental proceedings using laboratory animals followed the Organization for Economic Cooperation and Development (OECD) guidelines 423 . The expDuring the experiment, the rats were provided with hydration and daily food in equal quantities. Additionally, signs of toxicity, such as changes in coat, motility, tremors, changes in body weight (mass), food intake, grooming, respiration, stool appearance, mobility, and respiration, as well as convulsions and deaths, were noted during the few hours after gavage and then daily for 14 days.n = 3). One-way analysis of variance (ANOVA) was used to analyze the obtained data, followed by Duncan’s multiple distribution test. α = 5% was considered to describe the significant level (SPSS Version 20).The analyses’ results were presented as mean and standard deviation (SD). All experimental data collected were realized in triplicate (H. aspersa Müller meat has a very high nutritional value, which is originated in its substantial protein content. The notable presence of unsaturated fatty acids in this flesh highlights its nutritional value. A harmonious balance between ω6 and ω3 fatty acids, combined with a favorable alliance between polyunsaturated and monounsaturated fatty acids, lends eminent importance to this remarkable composition. In addition, the abundance of essential minerals such as potassium, calcium, and iron confirms the vital importance of this food resource. Its potential as a rich source of bioactive molecules is undeniably implied by the considerable presence of antioxidants and the diverse range of chemicals detected in its aqueous extract. However, it is essential to note that oral administration of this extract showed no signs of acute toxicity in terms of behavioral effects, and it is equally remarkable that no noticeable disruption of the rats’ body weights was observed following this administration. Thus, using OECD standards, our study allows flesh from H. aspersa Müller to be included in the fifth group. It should be stressed, nonetheless, that these findings require more research through extended studies of chronic oral toxicity, cytotoxicity, and histological examinations of certain organs in rats. Such comprehensive studies are imperative for a more profound understanding of the potential benefits and safety aspects associated with the consumption of H. aspersa Müller flesh and that of any other species utilized locally.The results clearly show that