Herein, we present magnetic proxy data that indicate a 4-fold increase of dust between ~ 2.73 and ~ 2.72 Ma, with subsequent increases at the start of glacials thereafter, indicating a strengthening of the mid-latitude westerlies. Moreover, a permanent shift in dust composition after 2.72 Ma is observed, consistent with drier conditions in the source region and/or the incorporation of material which could not have been transported via the weaker Pliocene winds. The sudden increase in our dust proxy data, a coeval rapid rise in dust recorded by proxy dust data in the North Atlantic (Site U1313), and the Site 1208 shift in dust composition, suggest that the iNHG represents a permanent crossing of a climate threshold toward global cooling and ice sheet growth, ultimately driven by lower atmospheric CO The amount and composition of North Pacific dust tracked by rock magnetism suggests that the intensification of North Hemisphere Glaciation ca. 2.7 million years ago marked the permanent crossing of a climate threshold. This relatively warm and humid Pliocene world deteriorated into a drier, cooler climate, with Northern Hemisphere glaciation commencing at ~3.6 Ma as inferred from δ18O data3. However, attention has focused on intensification of Northern Hemisphere glaciation, hereafter the iNHG, at ~2.73 Ma, recorded by decreases in biogenic opal and alkenone mass accumulation rate (MAR)5. The iNHG trigger remains unresolved. Proposals include orographic changes6, closure of the Panama gateway7, changes in orbital forcing7, and a reduction in atmospheric CO28. The processes associated with these events are not mutually exclusive, but there is also a lack of agreement on whether the iNHG was gradual3 or abrupt9. While the cause of the iNHG is still debated, the event itself is well-documented. It is marked by pronounced increases in IRD at ~2.75 to 2.7 Ma in the North Pacific10, and in the North Atlantic11 and Nordic Seas12, with more than an order of magnitude increase recorded after ~2.72 Ma in the latter. Distinct climatological and palaeoceanographic changes are also associated with the iNHG15.The Pliocene Epoch (5.3–2.6 Ma) is considered an analog for future climate given current anthropogenic warming because temperatures were 2–4 °C higher18. Dust deposition can be controlled by, or itself modify changes in climate. The former reflects shifts in atmospheric circulation. The latter means dust also has a direct influence, through scattering and absorption of solar radiation19, or indirect influence by serving as condensation nuclei20. It can also strengthen the biological pump by supplying limiting nutrients like iron which can directly influence atmospheric CO221. Here we apply environmental magnetism to trace dust deposition across the iNHG at ODP Leg 198 Site 1208 of the North Pacific , with a water depth of 3346 m, is located on the Central High of Shatsky Rise. The site lies within the western, north-flowing side of the North Pacific subtropical gyre, known as the Kuroshio Current Extension, and is sufficiently far from the Asian continent to preclude hemipelagic inputs. The topographic height and high sedimentation rate22 make this site well-suited to record eolian inputs and paleoclimate. Barium records for Site 120816 suggest that sulfate  and its “hard isothermal remanent magnetization" . An assessment of this parameter using a lower reference magnetic field , supports the interpretation that changes in antiferromagnetic  concentration and not grain size controls HIRM  over ~15 kyr  grains are in general finer than pseudosingle domain (PSD) grains, which are in turn finer than multidomain (MD) grains. But for grain populations of mixed composition, as is indicated for the Site 1208 samples from our other magnetic measurements, interpretations of grain size and domain state from magnetic hysteresis data are nonunique. Nevertheless, the magnetic hystereses parameters define a nominal pseudosingle domain (PSD) state, and trend toward values consistent with larger grain sizes with time. In particular, the post-iNHG magnetic hysteresis data are shifted from the clumped pre-iNHG data , and isothermal remanent magnetization (IRM) curves , ii. middle  and iii. high , magnetic coercivity components. We also note an outlier, where B1/2 reaches ~1 T, that may record volcanic ash. On the basis of their coercivity values, we interpret these components as i. PSD–MD magnetite, ii. SD magnetite, and iii. hematite. The IRM component importance varies across the iNHG as follows: i. 9% to 15%, ii. 77% to 75%, and iii. 15% to 10%. This analysis reveals the possibility of a nearly invariant component (ii) of relatively fine-grained magnetite. We note that similar IRM components have been found in studies of Shatsky Rise sediments younger than ~1 Ma, and the middle-coercivity component has been interpreted to reflect biogenic magnetite31. This might account for some of the nominally unvarying component across the iNHG in our Site 1208 IRM data. Biogenic magnetite might be less well-expressed in our other magnetic data  because of abundant eolian material in an overlapping grain size range.Data defining the acquisition of an IRM can be decomposed into three components that provide another view of magnetic domain state Supplementary Figure 300), and grain size (χARM/SIRM) proxies support this interpretation, as do the magnetic hysteresis and FORC analyses.In summary, our magnetic data indicate that Site 1208 witnessed a pulse of hematite-rich eolian material at the iNHG, best recorded by our dust proxy (HIRM flux). This was followed by successive depositional events in glacials, consisting of a greater abundance of hematite-rich material with larger magnetic grain sizes. Concentration and flux , composition 18, and the Fe mass accumulation rate reported for South Atlantic Site 1090 17. These sites provide a synoptic global view because they are at similar latitudes representing the Pacific, North Atlantic, and South Atlantic realms. We also include North Pacific Site 885/886 33, which is slightly poleward of the other sites, for comparison,  that are problematic in terms of the high-resolution assignment. The difference in dust increase between Site 1208 and 885/886 could imply an equatorial shift16 of the westerlies. But we note that a recent restudy of Site 885/88638 has failed to confirm the original magnetostratigraphy, pointing to uncertainties in age so large that they presently preclude the use of these sediments in high-resolution paleoclimate studies. However, a shift in dust source, from the more northern Gobi Desert to the more southerly Taklimakan Desert, is consistent with our magnetic parameters and implies an equatorial migration of the westerlies around this time34 at North Pacific ODP Site 885/886, which instead seems to record an order of magnitude increase of dust MAR starting at a nominal age of 3.6 Ma, close to the age of initial buildup of North Hemisphere ice. Environmental magnetic studies of Site 885/886 and similar sites39, and the possibility of hiatuses, also limit further comparisons with our Site 1208 record, but the age resolution at North Atlantic Site U1313 does afford the possibility of further comparisons 42 or Indian Monsoon (blocked by Tibetan Plateau)44. Thus, shifts in hematite formation—likely via ferrihydrite dehydration45—would explain this trend and the pre-iNHG muted HIRM signal.Sources of the hematite increases observed at the iNHG and subsequent glacials are reasonably explained. The Taklimakan may have experienced an expansion and/or drying event at 2.8–2.7 Ma17, they were unable to transport large quantities of hematite to Site 1208, and those magnetic grains that were transported were finer. These pre-iNHG dust events are recorded by the synchronous peaks amongst our concentration proxies. Other peaks, decoupled from HIRM, are likely the result of events not associated with atmospheric circulation shifts . However, given the relatively low resolution in this part of the record, we caution against overinterpretation of the data. With intensification of the westerlies at the iNHG, larger grain sizes, and thus higher concentrations of coarse-grained hematite could be carried aloft.As winds in the Pliocene are thought to have been weaker than after the iNHG16 and the Site U1313 n-Alkane flux18. However, HIRM flux dust peaks are not always coincident with these other peaks. However, these differences also highlight that any common signal seen amongst the proxies is likely a primary record of wind processes responsible for dust deposition.It is important to recognize that each of the proxies has benefits and limitations and should be expected to record slightly different aspects of the total eolian load. In particular, the benefit of magnetic measurements like HIRM flux is the ability to link to mineral type, but it might also be influenced by volcanic ash, irrespective of our efforts to avoid ash horizons during sampling. The Th-based dust flux relies on the assumption of constant influx of micro-meteoritic material, which is unknown on 10’s of kyr time scales. In addition, the n-Alkane proxy relies on the transport of plant waxes which could reflect source changes in ways different from the other proxies. As these records depend on completely independent sources, they are likely to be controlled on different time scales. There is some agreement in peak timing with those observed in the Site 1208 dust flux recordWind intensity is a function of meridional temperature gradients, with an increase resulting in stronger winds and vice-versa. In response to global cooling and ice volume increase, the high-latitudes would have cooled much greater than the tropical-subtropical regions. This would increase the meridional temperature gradient causing the winds to intensify and likely migrate equatorward.∘ further north of their pre-industrial locations46. Additionally, a weakening of the westerlies was observed in models focused on the last deglaciation47. Conversely, last glacial maximum (LGM) simulations in the Northern Hemisphere indicate equatorial migrations and strengthened westerlies48,49.Modeling results of the mid-Piacenzian Warm Period  have shown the Northern Hemisphere westerlies were located some ~42. Prior to the iNHG, CO2 levels remained above 300 ppm and as high as 440 ppm50,51, though there is significant scatter between these records. During this time, Site 846 δ18O records indicate at least two apparent arrested shifts toward glaciation52. However, modeling work suggests a 280 ppm threshold for Northern Hemisphere glaciation53 and it was not until marine isotope stage G6 (2.73 to 2.7 Ma) that this threshold was persistently crossed2. Thus, the climate threshold crossed at the iNHG implied by our observations of rapid changes of dust at the iNHG appears to be one of ice volume and atmospheric CO2 content in a cooling world.These climate changes are ultimately interwoven with changes in atmospheric CO56. Shifts in position are more robust. Historical period data indicates a poleward displacement57, a finding supported by the most recent IPCC report58. Climate simulations examining the future Northern Hemisphere jet stream behavior echo the historical findings61.It is interesting to note that historical records disagree on changes in Northern Hemisphere jet stream speeds62. This increase is thought to reflect an increase in Asian aridity, despite concentration increases observed for all grain sizes present and what this means for shifts in wind strength63. Similarly, HIRM values from Chaona section of the Chinese loss plateau (CLP) increase markedly at 2.72 Ma64. This part of the record is ascribed to variations in hematite/goethite alteration, even though the increase roughly doubles over the preceding 5 Myr average. However, using the CLP for paleoclimatic studies might be problematic65.Importantly, we can reassess the interpretations of environmental magnetic records from marine and terrestrial sites in the region around Site 1208. The concentration of the 8 μm grain size fraction increases at 2.73 Ma at ODP Site 882A300 compositional proxy and concentration proxy HIRM, and increases in ferrimagnetic grain size (recorded by our grain size proxy χARM/SIRM). The changes recorded by these magnetic proxy data younger than 2.68 Ma  recorded by our S66 leading to increased aridification. This cooling climate will eventually lead to a switch in the contribution of ferrimagnetic minerals over antiferromagnetic minerals because the latter are preferentially formed in warmer, drier conditions67. The decrease in our post-iNHG HIRM flux records highlights this gradual decrease of hematite in the source regions, which will eventually be replaced with PSD and MD magnetite as the dominant magnetic contributor, as is the case today69. Also, with increased westerly winds, newly eroded ferrimagnetic material might spend less time in the source region where it might otherwise be further oxidized. Finally, we also note that the increase in ferrimagnetic grain size (recorded by our grain size proxy χARM/SIRM) indicates that winds are still intense post-iNHG, but their dust compositional load is different.Global cooling would reduce the amount of precipitation received by central Asia2 in a cooling world. It is as yet uncertain whether a similar threshold could cause an acceleration of the weakening and poleward shift of the westerlies that is predicted by some models of future anthropogenic warming.Our dust proxy, grain size, and compositional records based on environmental magnetism indicate fundamental changes at the iNHG as viewed from the North Pacific ODP Site 1208, highlighted by a prominent increase in dust deposition. This change in dust deposition is seen in sites from the North and South Atlantic, indicating its global nature. After the iNHG, the dust magnetic mineralogy at Site 1208 is more antiferromagnetic  dominated, with a coarser grain size. These changes are consistent with colder, more arid conditions and/or the incorporation of material that could not have been transported via the weaker Pliocene winds. The age sequence of dust deposition changes at the iNHG are best recorded by Site 1208 and North Atlantic Site U1313. These sites both record a distinct, coeval onset age of increased dust deposition between ~2.74 and 2.73 Ma and a peak 15–30 kyr thereafter. This temporal character indicates that the westerlies rapidly intensified at this time. Nonetheless, it is necessary to couple the intensification demonstrated here with the equatorial migration proposed elsewhere. Such an integration reveals the complete scope of paleoclimatic changes in the North Pacific around the iNHG. Combined with our statistical analysis and the greater nuances available from our high-resolution record, we posit that the iNHG represents the permanent crossing of a climate threshold defined by ice volume and decreased atmospheric CO22 were avoided during sampling. Samples were collected in 10 × 75 mm borosilicate glass test tubes. A Parafilm strip was placed over the test-tube end following collection to prevent drying. These tubes were subsequently trimmed at the University of Rochester so the sediment was fully exposed to the laboratory-induced fields used for our environmental magnetic proxies.Samples were collected from the ODP Site 1208 working half cores at the Gulf Coast core repository. Ash layers documented in the ODP Site 1208 initial reports70 to construct a Site 1208 age model spanning 1.8 to 3.7 Ma70. These ‘recompressed’ depths (RD) were calculated asTo correct for sediment expansion after recovery, we applied a modified version of the equation used by Venti and Billupsδ18O record70, which had been previously tuned to the global benthic δ18O stack71. After adjusting the reported Site 1208 depths using our ‘recompressed’ equation (1), we used linear interpolation between depth-age tie point to assign ages to our samples. Age and depth tie points from ~3.7 Ma to 4.25 Ma are from astronomically tuned paleomagnetic datums72.We assigned ages on the basis of two previously determined age models for Site 1208. The section spanning ~2.45–3.7 Ma is based on the Site 1208 χ); 2. natural remanent magnetization (NRM); 3. alternating field (AF) demagnetization of the NRM; 4. acquisition of an anhysteretic remanent magnetization (ARM); 5. demagnetization of the ARM; 6. acquisition of a saturation isothermal remanent magnetization (SIRM) in an applied field of 1.5 T; 7. acquisition of a backfield of −100 mT; 8. acquisition of a backfield of -300 mT; 9. magnetic hysteresis parameter meaurements of coercivity (Hc), coercivity of remanence (Hcr), saturation magnetization (Ms) and saturation remanent magnetization (Mrs). 10. acquisition of isothermal remanent magnetizations (IRMs); 11. aquisition of First-order reversal curve data (FORCs). Measurements 1–8 were collected on test-tube sediment samples.All magnetic measurements were conducted in the paleomagnetic laboratories at the University of Rochester. Eleven types of magnetic data were collected in the following sequence: 1. Bulk mass-normalized magnetic susceptibility (χ measurements were made using a KLY-4 Agico Kappabridge in a 300 A/m field. NRM measurements were performed on a 2G Enterprises (Model 755) 3-component DC SQUID magnetometer with high-resolution sensing coils in a magnetically shielded room at the University of Rochester (background field <200 nT). AF demagnetization and ARM acquisition was done using a Magnon Alternating Field Demagnetizer 300. The samples were demagnetized in 10 mT steps up to 80 mT at a decay rate of 5 mT/s. ARMs were imparted at a peak field of 80 mT and a bias field of 50 μT. SIRMs and backfields were imparted in applied fields of 1.5 T, and −100, and −300 mT, respectively, with an ASC Scientific IM-10-30 Impulse Magnetizer. Magnetic hysteresis parameters, IRM acquisition curves, and FORCs were measured on representative samples with a Princeton Measurements Corporation MicroMag 2900 Series Alternating Gradient Force Magnetometer (AGFM).Bulk mass-normalized magnetic susceptibility 300)73 is defined as74 is defined as27 is defined asmT100 is calculated similarly to HIRMmT300 but with IRMmT−100.The S-Ratio  indicate the presence of high-coercivity material  while higher values (~1) indicate the dominance of low-coercivity material . The L-Ratio is a check to ensure that S300 and HIRM can be interpreted in this way27. Comparison of the environmental proxies , concentration proxies , and grain size proxy  allow for discernment of times with greater dust deposition into the ocean.S75. This software requires manual ‘tuning’ of the Mean Coercivity (B1/2), Dispersion Parameter (DP), Relative Proportion, and Skewness to achieve a model that best represents the fitted data, in order to minimize the Residual Sum Square (RSS) value. FORCs were processed using the VARIFORC protocol76 within FORCinel77.IRM acquisition curves were decomposed using the MAX UnMix software2/(cm2*kyr) was determined by multiplying HIRM (A m2/kg) concentration values by mass accumulation rates (MARs), which were in turn calculated by multiplying dry-bulk density ) data22 by corresponding sedimentation rate values. We note that our HIRM values are mass normalized, and differentiated from other efforts that have used volume normalization36. We urge use of mass normalization following the method described above to allow future comparisons between data sets.HIRM flux . The test was implemented in MATLAB using the signtest function with the null hypothesis that the data in a particular subgroup  had a median equivalent to that of the entire parameter’s population. We implemented one-tail tests (right or left) with an alpha level = 0.1. A one-tailed test is appropriate when you are only concerned if there is a difference between populations in a particular direction and you have an assumption of what that direction might be. The choice of using a right- or left-tailed was based on whether we believed the values in the subgroup should be larger (right-tailed) or smaller (left-tailed) than the population median. The null hypothesis was rejected if the p value was <0.5.The sign test78 is another nonparametric test that tests the null hypothesis that the medians of two populations are equal. A critical chi2 value of 2.706 was used, given degrees of freedom (df) = 1 and an alpha level = 0.1. The null hypothesis was rejected if the calculated chi2 value was greater than the critical chi2 value. This test was carried out using the mediantest function  which can be downloaded from MATLAB’s file exchange webpage.Moods median testSupplementary Information


The overall results showed that pan-roasting could be used to obtain relatively high health benefits from the antioxidant and inhibitory activities of sacha inchi seeds. The information obtained from this study may serve as the basis for the proper processing of sacha inchi seeds to optimize their functional food and nutraceutical applications.Roasted sacha inchi seeds are now commercialized as a health food product, but the influence of roasting methods on their proclaimed health effects has yet to be explored. This study investigated the total phenolic contents (TPCs), antioxidant potential, and inhibitory activities of raw and roasted sacha inchi seeds in vitro. Individual phenolics in raw seeds were also identified in an attempt to explain the bioactivities of the seeds. The results suggested that roasting in a cooking pan, vacuum oven, and tray dryer had distinct impact on TPC in sacha inchi seeds, and thus differentially altered their antioxidant and inhibitory properties. Seeds that underwent roasting exhibited 1.5–2.7-fold higher antioxidant potentials than raw seeds. Certain roasting methods provided the products with anti-α-amylase and anti-cholinesterase activities, while inhibitions of these enzymes were not detected in raw seeds. Roasted seeds also possessed superior anti-lipase and anti-glycation activities when compared with raw seeds . The inhibitory properties observed in the seed samples might be attributed to their  Plukenetia volubilis L.) or the Inca peanut is an important indigenous food crop in Latin America, and also a traditional remedy employed by local communities in the Amazon region [Sacha inchi  such as coronary heart disease and hypertension ,8. PositTraditionally, sacha inchi seeds are consumed as a gourmet oil or cooked by pan-roasting, steaming, boiling, and other conventional cooking methods . The proThis comparative study investigated the phenolic content, antioxidant activities, and inhibitory properties of sacha inchi seeds both raw and cooked using different roasting methods. Cooked seeds were prepared by roasting in a cooking pan, vacuum oven or tray dryer to mimic the processes generally used for producing snacks from whole nuts. Enzyme inhibitory assays were conducted using the key enzymes associated with common metabolic  and neurodegenerative (Alzheimer’s disease) disorders. Inhibition of non-enzymatic glycation was also observed in vitro. Furthermore, we identified phenolic compounds in raw sacha inchi seeds in an effort to explain the health properties of the seeds. The results obtained from the present study can support further research on the bioactivities of roasted sacha inchi seeds and also serve as the basis for the proper processing of sacha inchi seeds for functional food and nutraceutical applications.Sacha inchi seeds, with kernels separated from shells, were kindly provided by Thai Rubber Land and Plantation Co., Ltd., Muang, Chiang Rai, Thailand. We conducted all experiments within one year after obtaining the samples in March 2014. The proximate composition and appearance of the seeds are presented in v/v) aqueous ethanol in a 70 °C water bath shaker for 15 min. The supernatants were received after centrifugal solid–liquid separation  and then filtered using 0.45 µM PES membranes. The clear extracts were collected and kept at −20 °C until analysis. For further investigations, all reagents and chemicals were obtained from Sigma-Aldrich , unless stated otherwise, and were of the highest quality.Extraction of sacha inchi samples was conducted following Kittibunchakul et al. (2022) . BrieflyAnalysis of the phenolic profiles was performed following the method of Temviriyanukul et al. (2020) with no modifications . BrieflyTM HT, BioTek Instruments Inc., Winooski, VT, USA) with BioTek Gen5 software (version 2.09), as previously described by Hinkaew et al. (2020) with no modifications [v/v) Folin–Ciocalteu phenol reagent and 7.5% (w/v) sodium carbonate at 25 °C for 2 h. Subsequently, the absorbance value was determined as an end-pointed assay at 765 nm, and results were reported as mg gallic acid equivalent (GAE)/100 g dry weight (DW).Total phenolic contents (TPCs) were determined using a microplate reader (Model Synergyications . In brieex) and emission wavelength (λem) of 485 nm and 528 nm, respectively. The FRAP assay was conducted by incubating the extracted sample with the FRAP reagent at 25 °C for 8 min, and the absorbance was subsequently measured as an end-pointed assay at 600 nm. The DPPH radical scavenging assay was conducted by incubating the extracted sample with DPPH reagent at 25 °C for 30 min before the absorbance measurement as an end-pointed assay at 520 nm. For all the antioxidant activity assays, results were reported as µmol Trolox equivalent (TE)/100 g DW.Antioxidant activities were determined spectrophotometrically with a microplate reader utilizing the ORAC, FRAP and DPPH radical scavenging assays. All assays followed protocols previously described by Sripum et al. (2017) with no modifications . BrieflyA is the initial reaction rate of the control reaction with enzyme but without the sample extract, a is the initial reaction rate of the control reaction without enzyme and the sample extract, B is the initial reaction rate of the enzymatic reaction with the sample, and b is the initial reaction rate of the reaction with the sample but without enzyme.Inhibitions of the key enzymes associated with type II diabetes (α-amylase and α-glucosidase), obesity (lipase), and Alzheimer’s disease  were investigated in vitro following protocols previously described by Sirichai et al. (2022) with no modifications . The inhex 330 nm and λem 410 nm. The inhibitory activities reported as percentages of inhibition at a particular extract concentration were calculated using Equation (1).The inhibition of non-enzymatic protein glycation of bovine serum albumin (BSA) was investigated, as previously described by Kittibunchakul et al. (2022), with no modifications . In brien = 3), with results presented as mean ± standard deviation (SD). An ANOVA followed by Duncan’s multiple range test was used to compare significant differences between mean values. An unpaired t-test was used to compare significant differences between the values of two data sets. All statistical tests were conducted using SPSS software , and values were considered significantly different at p-values less than 0.05 (p < 0.05).All experiments were conducted in triplicate (p-coumaric acid (84.38 µg/100 g DW), ferulic acid (90.26 µg/100 g DW), and quercetin (71.65 µg/100 g DW) by HPLC analysis compared with 17 authentic phenolic standards, including 8 flavonoids  and 9 phenolic acids  (The major phenolic compounds in our raw seeds were identified as ic acid) . As showThe capacity to resist oxidation was explored in vitro using ORAC, FRAP and DPPH radical scavenging assays, with results shown in v/v) aqueous ethanol for comparison, as shown in The health-related properties of raw and roasted seeds of sacha inchi were explored in vitro by determining the inhibition of the key enzymes relating to type II diabetes (α-amylase and α-glucosidase), obesity (lipase), hypertension (ACE), and Alzheimer’s disease . Results were presented as percentages of inhibition at a final extract concentration of 0.4 mg of sacha inchi seeds per mL of 20% (50) of vacuum oven-roasted seeds on this enzyme was, thus, higher than 0.4 mg/mL.Inhibition of the carbohydrate-hydrolyzing enzymes, α-amylase and α-glucosidase, retarded starch degradation, slowed down the release of glucose, and thus reduced the glucose absorption and blood glucose levels. Therefore, this was adopted as a diabetes prevention strategy. The results indicated that only α-amylase inhibitory activity was detected using an extract concentration of 0.4 mg/mL, while the α-glucosidase inhibitory activity of all the samples was undetected using the same extract concentration. However, α-amylase inhibitory activity was only detected in sacha inchi seeds roasted using a vacuum oven (16.33% inhibition), with no activity observed in the other samples. The half-maximal inhibitory concentration  in the brain is also a hallmark of this disease. The formation of Aβ plaques is induced by the hydrolysis of amyloid precursor protein by BACE-1. BACE-1 inhibition has thus been shown to be a therapeutic approach to this neurodegenerative disorder. Among all studied sacha inchi seed samples, AChE inhibitory activity was only observed in seeds cooked via pan-roasting (12.25% inhibition) using an extract concentration of 0.4 mg/mL, indicating that its IC50 of the extracted samples was considered to be lower than 0.125 mg/mL. The inhibitory activities working against MG- and D-glucose-induced glycations followed the same trend, with seeds cooked via pan-roasting possessing the highest inhibitory activities, followed by seeds roasted using a vacuum oven and tray dyer, respectively; raw seeds exhibited the lowest value. Increases in inhibitory activities by 4.4–4.8-fold and 3.9–3.5-fold after roasting processes were observed in anti-glycation reactions induced by MG and D-glucose, respectively. Moreover, significantly stronger inhibitions in the reaction induced by D-glucose than MG were observed in raw seeds and pan-roasted seeds.In vitro inhibitions of non-enzymatic glycations by raw and roasted seeds of sacha inchi are reported in Sacha inchi seeds have gained increasing attention from both consumers and the industrial sector due to their attractive nutritional and functional properties. Products made from the seeds such as seasoned seeds, crunchy snacks, protein powder, and omega-3 rich oil supplements are now industrialized . Previoup-coumaric acid, ferulic acid, and quercetin, which were somewhat distinct from the individual phenolics detected in sacha inchi oil [The phenolic content (45.15 mg GAE/100 g DW) found in our raw sacha inchi seeds was different from the TPC values reported by Chirinos et al. 2013) (65–80 mg GAE/100 g) [  and microwaving (radiation), concurring with the study of Liao et al. (2012) . On the Sacha inchi seeds are nutritious and contain several phytochemicals with potential health-promoting effects. The seeds are usually processed into food products by roasting in a pan, as well as in an industrial oven or dryer for commercial purposes. Roasting affects the chemical composition of sacha inchi seeds, which in turn influences their TPC, antioxidant potentials, and in vitro inhibitory activities. The effects of roasting on the seeds were greatly dependent on the roasting method used. Roasting under the studied conditions potentially results in cooked seeds with improved health properties. The inhibitions of disease-associated enzymes and glycation reactions by the seed extracts suggested that phenolic compounds might be responsible for some of these inhibitory effects; however, other phytochemicals  might also play roles in these inhibitions. In this study, inhibitory activities were only reported using a particular extract concentration, making a comparison of our results in enzyme inhibitions rather limited. In addition, further optimization of the extraction process is still required for a detailed phytochemical analysis of sacha inchi seeds. Nevertheless, the information obtained from the present study can support further research on the health properties (including bioaccessibility and toxicity) of roasted sacha inchi seeds using human cell line models or in vivo investigations. Knowledge gained from this research may serve as the basis for the proper processing of sacha inchi seeds for optimal functional food and nutraceutical applications, using thermal conditions that maximize the retention of health-related activities.


Although correct selection of pedicle screw dimensions is indispensable to achieving optimum results, manufacturer-specified or intended dimensions may differ from actual dimensions. Here we analyzed the reliability of specifications made by various manufacturers by comparing them to the actual lengths and diameters of pedicle screws in a standardized experimental setup. We analyzed the actual length and diameter of pedicle screws of five different manufacturers. Four different screw lengths and for each length two different diameters were measured. Measurements were performed with the pedicle screws attached to a rod, with the length determined from the bottom of the tulip to the tip of the screw and the diameters determined at the proximal and distal threads. Differences in length of > 1 mm were found between the manufacturers’ specifications and our actual measurements in 24 different pedicle screws. The highest deviation of the measured length from the manufacturers’ specification was 3.2 mm. The difference in length between the shortest and longest screw with identical specifications was 3.4 mm. The highest deviation of the measured proximal thread diameters and the manufacturer’s specifications was 0.5 mm. The diameter of the distal thread depends on the shape of the pedicle screw and hence varies between manufacturers in conical screws. We found clear differences in the length of pedicle screws with identical manufacturer specifications. Since differences between the actual dimensions and the dimensions indicated by the manufacturer may vary, this needs to be taken into account during the planning of spine instrumentation. Accurate selection of the screw diameter is essential to achieve optimal insertion into the pedicle and avoid fracturing of the cortical bone around the pedicle.Transpedicular instrumentation is a frequent procedure for dorsal stabilization of the thoraco-lumbar spine. Pullout strength of inserted pedicle screws depends on the design characteristics of the screws, the bone structure and quality, the accuracy of screw placement, and insertion depth.In relation to insertion depth, in most cases avoidance of anterior cortical breach is crucial due to risk of neurovascular injury. In some cases, however, intentional bicortical fixation might be justified in the interest of increased stability.diameter × length by the manufacturers. However, due to missing standards to adhere to, the actual dimensions might vary across manufacturers due to different measurement paradigms. The aim of this study was to analyze the lengths and diameters of thoraco-lumbar pedicle screws of different manufacturers in a standardized manner and compare our results with the manufacturers’ specifications.Pedicle screws consist of two elements, the tulip and the shaft. Morphology of the shaft is characterized by its shape, which can be cylindrical or conical, and by its dimensions. The dimensions of the shaft are specified by length and diameter, the latter can be given as the outer diameter of the thread and the inner diameter of the shaft without the thread. To achieve optimal screw fixation, exact knowledge of screw dimensions is essential. Dimensions of pedicle screws are usually indicated as No ethical approval was required for this in vitro biomechanical study.®, Aesculap—S4® Spinal System, DePuySynthes—Expedium®, K2M—Everest®) in an experimental setup. Four different lengths  and for each length two different diameters (5 and 6 mm or 5.5 and 6.5 mm or 6 and 7 mm) as indicated by the manufacturers were measured. For each type, two identical screws were analyzed.We analyzed the actual length and diameter of pedicle screws for thoraco-lumbar instrumentation from five different manufacturers .A detailed overview of the results is depicted in The labeled diameter of pedicle screws conformed to the diameter of the proximal thread in all screws, irrespective of whether they were of conical or cylindrical shape. The highest deviation of the measured proximal thread diameter and the manufacturer’s specification was 0.5 mm. The diameter of the distal thread depends on the shape of the pedicle screw. The proximal and distal diameters of the cylindrical pedicle screws were identical across all manufacturers. In conical-shaped screws, the dimensions of the distal thread diameter vary from manufacturer to manufacturer.2 However, the actual dimensions of pedicle screws with the same specifications might vary between different manufacturers due to different measurement modalities. In this study, we, therefore, analyzed the lengths and diameters of pedicle screws of various manufacturers in a standardized manner. We found notable discrepancies in length between pedicle screws with the same dimension specification of up to 3.4 mm.Characteristics of pedicle screws have a crucial impact on the biomechanics and strength of pedicle screw-based spinal instrumentation. The main features of pedicle screws are characterized by their length and their diameter, parameters which are clearly labeled on the screws by the manufacturers. The pullout strength in turn is proportionate to the diameter of the screw utilized and the intrinsic bone density. However, to maintain fixation strengths, the fracturing of the cortical bone around the pedicle must be avoided. This must especially be taken into account in osteoporotic patients with a thinner cortical bone, with screws with a too-large diameter posing a high risk of fracturing it and thus impairing stability.79 Accurate fitting of screws in the pedicle is thus crucial for achieving optimal results.Approximately 60% of the pullout strength of vertebral screw fixation depends on the fixation in the pedicle.12 However, when penetrating the anterior cortex exact knowledge of the pedicle screw dimensions is crucial to avoid violation of anterior structures which could lead to vascular injuries.14While exact planning of screw insertion depth is crucial for obtaining good strengths, this is of critical importance when a bicortical fixation is planned, as engagement of the anterior cortex has been shown to significantly increase the pullout strengths and improve bone fixation especially in osteoporotic patients.Therefore it is imperative for a surgeon to be exactly aware of the diameter of the utilized screws in the given pedicle segment. As opposed to clear deviations of screw lengths among different manufacturers, we did not find notable deviations of the diameters of the proximal threads in our measurements. The diameter of the proximal thread generally conformed to the labeled diameter on the screws, representing the largest individual screw shaft diameter in conical-shaped screws.16 Nevertheless, surgeons must take possible advantages or shortcomings into account. Our current data shows that due to a lack of uniform standards regulating the labeling of screw dimensions across various manufacturers, surgeons may not be able to rely on specified and labeled dimensions alone and would need to take relevant deviations into consideration.Our present study focused on differences in lengths and diameters of pedicle screws with the same labeling between different manufacturers. There are also other screw design elements significantly influencing biomechanics and stability of pedicel screw-based spinal instrumentation, such as the thread design or the shaft shape, which were not analyzed in this study due to manifold variations making a clear systematization impossible.Since we found dimensions of pedicle screws to exhibit relevant differences between different manufacturers despite bearing the same specifications, this might become of critical value when surgeons change instrumentation systems. Being accustomed to the specific dimensions of pedicle screws of one manufacturer, switching to another manufacturer might lead to unexpected results when actual screw dimensions may differ from the formerly used screws.At present, there are no uniform standards for defining dimensions in the manufacture of pedicle screws. Since these dimensions of different manufacturers do vary, we recommend the introduction of standards that would be obligatory for all manufacturers to adhere to.


We present a record of glacier ice dynamics and ice melt rate at the boundary between grounded ice and ocean—or grounding line—of Petermann Glacier, a major outlet glacier in Northwest Greenland. The traditional view of grounding lines implemented in ice sheet models in charge of projecting sea level rise is that they not migrate during the tidal cycle and experiences no ice melt. Instead, the satellite record reveals kilometer-size grounding line migrations—or grounding zones—with preferential intrusions along preexisting subglacial channels. The highest melt rates of ice are recorded within the grounding zone. Vigorous ice-ocean interaction in kilometer-wide grounding zone will make projections of sea level rise from glaciers potentially double. Warming of the ocean waters surrounding Greenland plays a major role in driving glacier retreat and the contribution of glaciers to sea level rise. The melt rate at the junction of the ocean with grounded ice—or grounding line—is, however, not well known. Here, we employ a time series of satellite radar interferometry data from the German TanDEM-X mission, the Italian COSMO-SkyMed constellation, and the Finnish ICEYE constellation to document the grounding line migration and basal melt rates of Petermann Glacier, a major marine-based glacier of Northwest Greenland. We find that the grounding line migrates at tidal frequencies over a kilometer-wide (2 to 6 km) grounding zone, which is one order of magnitude larger than expected for grounding lines on a rigid bed. The highest ice shelf melt rates are recorded within the grounding zone with values from 60 ± 13 to 80 ± 15 m/y along laterally confined channels. As the grounding line retreated by 3.8 km in 2016 to 2022, it carved a cavity about 204 m in height where melt rates increased from 40 ± 11 m/y in 2016 to 2019 to 60 ± 15 m/y in 2020 to 2021. In 2022, the cavity remained open during the entire tidal cycle. Such high melt rates concentrated in kilometer-wide grounding zones contrast with the traditional plume model of grounding line melt which predicts zero melt. High rates of simulated basal melting in grounded glacier ice in numerical models will increase the glacier sensitivity to ocean warming and potentially double projections of sea level rise. The Greenland Ice Sheet has lost billions of tons of ice to the oceans in the last few decades, increasing global sea level by 14 mm since 1972 2. In the Prior modeling studies have questioned the assumption of a grounding line with zero melt. Parizek et al.  used a cA number of recent studies using radar interferometry data have recently pointed out the existence of broad, kilometer-size “grounding zones” in Antarctica, e.g., on Pine Island Glacier , Thwaite12 kg) for the period 1972 to 2017  . With it to 2017 . The ret to 2017 .In prior studies, we used less differential interferometric synthetic aperture radar (DInSAR) data to map the grounding line with high precision : two DInMaterials and Methods. Previous attempts at mapping ice shelf melt rates using optical-derived DEMs conservatively stopped several kilometers seaward from the grounding line to be sure to remain in a region of hydrostatic equilibrium  after verifying that hydrostatic equilibrium still applies in that zone. We document how the grounding zone evolves from the 1990s to the present and observe how the melt rates change during the retreat. We conclude on melt rates in the grounding zone before and during the retreat and discuss the impact of the results on projecting glacier evolution in a warming climate.To map the ice sheet surface elevation, we use a monthly time series of digital elevation models (DEM) from the German Space Agency (DLR) TanDEM-X SAR mission for the period 2011 to 2021  Fig. 1)Fig. 1). ilibrium . Here, w1.The 1992 to 2011 grounding zone (GZ) of Petermann is broadly rectilinear with two extension lobes on each side . The GZ 2, respectively; that is, it nearly doubled. Most of the change took place at the glacier center and along the western and eastern extension lobes.After calibration of the DEMs, we find that the height above flotation (HAF) of the ice surface is zero at the center of the inferred GZ and less than 2 to 3 m within the GZ, which is consistent with the hypothesis that ice is floating in the GZ. From 1992 to 2013, 2011 to 2015, 2016 to 2017, 2020 to 2021, the GZ area grew in size from 74.5, 86.7, 81.9, and 125 kmThe GZ retreated along an area of retrograde slope, i.e., where bed elevation drops in the inland direction . The retMaterials and Methods). The main channel (Profile C in 3/s on average) of the total subglacial water discharge in winter from geothermal and frictional heat versus 27% for the secondary channel (Profile D) and 3% for each of the other two channels (Profiles A and E). In summer (June to August), the main channel drains 43% of the total water discharge (or 86 m3/s on average for the three summer months) versus 45% for the secondary channel, and 6% each for the other two channels. The region of the most pronounced retreat is therefore aligned with the main channel, but we do not observe a similarly large retreat along the secondary channel, which discharges almost as much subglacial water. Over the time period 2001 to 2022, subglacial water discharge increased from 149 to 301 m3/s in summer. This doubling in subglacial water discharge must have enhanced the ice melt rates but cannot be the only factor contributing to the retreat  but with improved precision. The highest melt rates are recorded in the GZ, not farther out on the ice shelf. As predicted by the plume model, we observe a secondary peak about 10 to 15 km downstream. Melt rates, however, average 60 ± 15 m/y at the center of the GZ and exceed 80 to 100 ± 15 m/y along the east and west lobes , the total freshwater flux increased from 0.38 mSv in 2011 to 2015 to 0.50 mSv and 0.46 mSv in 2016 to 2017 and 2020 to 2021. For comparison, the freshwater flux for the entire ice shelf has been previously estimated at 0.33 mSv in year 2011 , the water height varies almost linearly from the full positive tide (2 m at the entrance for a ± 1-m tide) at the seaward edge of the GZ to zero at the upstream edge. This height is much thinner than further downstream, outside of the GZ and outside of the flexure zone, where it rapidly increases to several hundred meters . The wat3 in 2011 to 2015 and 0.125 km3 in 2020 to 2021 within 6 h. If we assume a water temperature T = 0.3 °C at 500-m depth in the cavity , of 2.6 °C. Using a heat capacity of cold seawater, Cp = 3,974 J kg−1 °C−1, and converting the water volume to a mass transfer, m, we obtain an ocean heat transfer, Q = mCp(T − Tf) of 0.36 1011 W to 6.0 1011 W to the ice. If all the ocean heat is available to melt ice with a latent heat of fusion, Lf = 334,000 J/kg, this yields a freshwater flux increasing from 0.11 mSv in 2011 to 2015 to 0.18 mSv in 2020 to 2021 or a mass flux of 3.5 to 5.7 Gt/y. These estimates, which assume that the warmest ocean waters can reach the GZ, are consistent with melt rates deduced from remote sensing data, hence providing additional confidence in the results.For a tidal amplitude of ±1 m, the cavity will fluctuate from 0 to 2 m at the entrance and taper linearly down to zero at the end, which is equivalent to an average water thickness of 1 m. The water volume in the cavity will therefore change by 0.075 kme cavity , a freezI). Point #1, at the seaward edge of the GZ, thinned at 2.2 ± 0.1 m/y vs. 1.2 ± 0.1 m/y on grounding ice at point #4. Since point #1 is fully afloat, the surface thinning of 22 m in 10 y translates into a total thickness reduction of 204 m. In contrast, point #4 remains grounded during the entire period, hence lost 12 m of thickness. If we attribute this ice lowering to dynamic thinning, i.e., a longitudinal stretching of grounded ice caused by ice flow acceleration, the same rate will apply to the proximate floating ice. With that correction, we deduce that ice shelf melt alone must have removed (204 − 12) m = 192 m of ice in 10 y at point #1 for a total cavity height of 204 m, which is a rapid rate of ice removal by Greenland and Antarctic standards. At points #2 and #3, in the middle of the cavity, melting is accompanied by thinning until 2019, followed by a small amount of thickening in 2019 to 2021. We suspect that the bending stresses, which push ice below hydrostatic equilibrium immediately seaward of the grounding line, relaxed as the grounding line retreated, which resulted in a gain in surface elevation instead of an expected drop in elevation caused by ice melt  from hydrostatic equilibrium. Using a Lagrangian approach with precision DEM data, we find that the GZ experiences the highest melt rates anywhere on the ice shelf, with values averaging from 60 ± 15 m/y at the glacier center to 80 to 100 ± 15 m/y at the glacier sides. During the retreat and formation of a new cavity, the average melt rates dropped to 40 ± 11 m/y before increasing again to 60 ± 15 m/y as the cavity became fully formed. In light of these results, we recommend revisions in the representation of ice melt rates at grounding lines in numerical ice sheet models. We suggest that ice sheet models adopt subshelf melt schemes that include melting on elements at and upstream of the grounding line . SeparatA.3 for ice and 1.028 g/cm3 for seawater. A height above flotation is calculated assuming that thickness minus free-board height times water density equals thickness times the density of ice. Thickness of grounded ice is surface elevation from TDX minus bed elevation from BedMachine.We use 302 digital elevation models (DEM) from the TanDEM-X (TDX) mission generated by the German DLR for the period 2011 to 2021 on a monthly basis. The data are automatically calibrated at high elevation using ICESat-1 data  and assuB.We use measurements of annual ice velocity from European Space Agency (ESA)’s Earth Remote Sensing (ERS) 1 and 2, Envisat ASAR, Sentinel-1 and 2, NASA’s Landsat 7 and 8, Canadian Space Agency (CSA)’s RADARSAT-1, Japanese Space Agency (JAXA)’s ALOS PALSAR, and the United States Geological Survey (USGS)’s Landsat-4/5 . VelocitC.We use prior mappings of the grounding line from synthetic aperture radar interferometric data  acquiredD.3, 2) an ocean water layer with a density of 1.028 g/cm3, and 3) a rock layer with a density of 2.67 g/cm3. A forward model of the gravity is calculated using the bed elevation from BMv5 and IBCAOv3. We calculate the direct current (DC) shift between modeled and observed gravity in areas where bed elevation is known from conductivity, temperature, and depth (CTD), BMv5, or multibeam echo sounding data. We interpolate the DC shift onto a regular grid using a minimum curvature algorithm, correct the observed gravity with the interpolated DC shift, fill the data gaps with the model results, and invert the resulting gravity field where bed elevation is not known. The interpolation of the DC shift accounts for natural variations in underlying geology across the model domain caused by variations in crustal thickness, sedimentary basins, or intrusions. The inversion stops when the misfit between observed and modeled gravity is less than 0.1 mGal, which corresponds to a nominal error in sea floor depth of ±60 m. The inversion reveals the presence of a ridge 10 km from the 1996 grounding line, which was previously reported using a two-dimensional inversion of coarser resolution, sparser sampling, NASA’s Operation IceBridge gravity data  . The surity data .E.2 (milli-Watt per square meter), basal friction assuming full sliding of the glacier at the bed, using the ice surface velocity, and assuming that 50% of the available heat is used to melt ice. The calculation is repeated with the addition of summer runoff from the Regional Atmospheric Climate Model Version p2.3 (RACMOp2.3) (We use a D-infinite channel model  to modelCMOp2.3)  for yearF.Two approaches are possible to express mass conservation on floating ice shelves and deduce the rates of bottom melt : 1) an EAppendix 01 (PDF)Click here for additional data file.


In vivo, Sprague–Dawley (SD) male rats were administered oxycodone with or without icotinib or apatinib. Blood samples were collected and subjected to UPLC-MS/MS analysis. An enzyme incubation assay was performed to investigate the mechanism of drug–drug interaction using both rat and human liver microsomes (RLM and HLM). The results showed that icotinib markedly increased the AUC(0–t) and AUC(0–∞) of oxycodone but decreased the CLz/F. The Cmax of oxycodone increased significantly upon co-administration of apatinib. In vitro, the Km value of oxycodone metabolism was 101.7 ± 5.40 μM and 529.6 ± 19.60 μM in RLMs and HLMs, respectively. Icotinib and apatinib inhibited the disposition of oxycodone, with a mixed mechanism in RLM  and a competitive and mixed mechanism in HLM . In conclusion, both icotinib and apatinib inhibit the metabolism of oxycodone in vitro and in vivo. Therefore, the dose of oxycodone should be reconsidered when co-administered with icotinib or apatinib.This study aimed to investigate the interactions between icotinib/apatinib and oxycodone in rats and to unveil the underlying mechanism. An ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) method was developed and validated to determine oxycodone and its demethylated metabolite simultaneously.  Comorbidities and multiple medicines are common during cancer therapy. Therefore, the incidence of drug–drug interactions is high in clinical practice . HoweverCancer pain is one of the most common comorbidities of cancer. Its incidence is 39.3% after curative treatment, 55% during anticancer treatment, and 66% in advanced or metastatic status . Oxycodoin vitro evidence that icotinib inhibits CYP3A  and human liver microsomes (HLMs) to investigate the pharmacokinetic inhibition of oxycodone by icotinib and apatinib. The outcomes of this investigation could provide valuable theoretical support for precision medicine, enabling the avoidance of adverse reactions and ineffective treatments.In this study, we employed a library of tyrosine kinase inhibitors (TKIs) to conduct a comprehensive screening for drug–drug interactions. We successfully identified icotinib and apatinib as highly potent inhibitors, based on their significant inhibitory level on oxycodone metabolism. Furthermore, we evaluated the drug–drug interactions through We obtained standard substances noroxycodone and midazolam  from Sigma-Aldrich . Additionally, we purchased icotinib from Inviwo Chemical Technology  and acquired apatinib from Sunflower Technology Development Co., Ltd. . Liquid chromatography (LC)–grade acetonitrile (ACN) was obtained from Merck . Ultrapure water was produced using a Milli-Q purification system. Finally, we obtained HLMs and RLMs from Corning Life Sciences Co., Ltd. .18 column, and incubation was carried out at 40 °C. The mobile phase consisted of 0.1% formic acid (A) and acetonitrile (B), with a gradient elution at a flow rate of 0.40 mL/min for 3.0 min. The gradient conditions were as follows: 10%–90% B (0–1.0 min), 90%–10% B (1.0–2.0 min), and 10% B (2.1–3.0 min). Multiple reaction monitoring was performed in the positive ion mode.An ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) method was developed to detect oxycodone and noroxycodone as previously described . This mem values of oxycodone in RLMs and HLMs.A 200-μL culture system was used, which included 1 M phosphate buffer, 0.2 mg/mL RLM or HLM, 1 mM NADPH, and 8–2,000 μM oxycodone. Before measurement, the mixture without NADPH was pre-incubated at 37 °C for 5 min. Then, 1 mM NADPH was added, and the reaction was terminated by cooling to −80 °C after 30 min. Next, 400 μL acetonitrile and 20 μL midazolam  were added to the mixture. After shaking for 2 min, the supernatant was obtained by centrifugation at 13,000 rpm for 10 min and subjected to UPLC-MS/MS analysis. With this method, we obtained the Km values, the concentration of oxycodone was also set at 100 μM. With this method, we obtained the inhibition rates of these 24 drugs on oxycodone metabolism and identified icotinib and apatinib as those with high inhibition rates. To determine the half-maximal inhibitory concentration (IC50) of icotinib and apatinib, the concentrations of icotinib and apatinib were set at 0.01, 0.1, 1, 10, 25, 50, and 100 μM, while the concentration of oxycodone was set at 100 μM (in RLMs) and 500 μM (in HLMs), based on the corresponding Km values. To determine the inhibition mode, the concentration of oxycodone was set at 25, 50, 100, and 200 μM in RLMs and at 50, 250, 500, and 1,000 μM in HLMs. The concentration of icotinib was set at 0, 0.75, 1.5, and 3 μM in RLMs and at 0, 5, 10, and 20 μM in HLMs, based on the IC50 value. The concentration of apatinib was set at 0, 0.25, 0.5, and 1 μM in RLMs and at 0, 0.125, 0.25, and 0.5 μM in HLMs. After incubation, the samples were prepared and measured by UPLC-MS/MS in accordance with the abovementioned method. Data were collected as previously described  rats were obtained from Viton Lever Laboratory Animal Center . Rats with a body weight of 200 ± 20 g were used for the experiment. The rats were acclimated in animal rooms for 1 week, with the room temperature maintained at 20–25 °C and the humidity ranged from 50% to 65%. To simulate day and night cycles, the light and dark conditions were alternated every 12 h. During the acclimation period, the rats were provided with unlimited access to food and water. The study was conducted in accordance with the guidelines of the Ethics Committee of Wenzhou Medical University (wydw2020-0322). The experimental animal center has a license number SCXK  2019-0001. Prior to the start of the experiment, the rats were fasted for 12 h. For euthanasia, 2% isoflurane was used, while 5% isoflurane was used for pharmacokinetic experiments.n = 6), namely the control group, the icotinib + oxycodone group, and the apatinib + oxycodone group. We dissolved icotinib and apatinib in 0.5% CMC-Na solution to a final concentration of 37.5 and 40 mg/mL, respectively. Based on the bioequivalent dose, we converted the concentrations of the two inhibitors. The icotinib + oxycodone group received 37.5 mg/kg icotinib per mouse by oral administration, while the apatinib + oxycodone group received 40 mg/kg apatinib per mouse by gastric gavage. After 30 min, 3 mg/kg oxycodone was subcutaneously injected into each mouse in the three groups. To dilute oxycodone to a final concentration of 2 mg/mL before injection, we used saline solution. At 0.083, 0.167, 0.25, 0.5, 1, 2, 4, 6, 8, and 10 h after injection, we collected blood samples from the tail vein as previously described  software 3.0, utilizing a noncompartmental model. The drug–time curves were plotted using Origin 8.0. All data are presented as the mean ± standard deviation. Statistical analysis was performed using an independent-samples The monitoring transitions of oxycodone, noroxycodone, and midazolam are m/z 316.2 → 241.1, m/z 302.2 → 187, and m/z 326.1 → 291.1, respectively. The chromatogram condition was optimized. As shown in m value of oxycodone was 101.7 ± 5.40 μM, as shown in m value of oxycodone was 529.6 ± 19.60 μM, as shown in 50 values of icotinib/apatinib in RLMs and HLMs  and AUC(0–∞) values of oxycodone in the oxycodone + icotinib group were significantly increased by 1.4 and 1.2 times, respectively, compared with those in the control group (P < 0.05). In addition, the CLz/F value decreased by about 0.7 times (P < 0.05). Upon co-administration of apatinib and oxycodone, the Cmax value of oxycodone significantly increased (by 1.5 times) compared with the control group (P < 0.05).To further evaluate the interaction between icotinib/apatinib and oxycodone i value was 0.65 μM for apatinib and 1.04 μM for icotinib. In HLMs, the inhibition mode of apatinib was the same as that in RLMs, with Ki = 1.04 μM, whereas icotinib competitively inhibited the metabolism of oxycodone, with Ki = 9.42 μM.As shown in e.g., voriconazole and diphenhydramine) can interact with oxycodone  and AUC(0-∞) values of oxycodone and decreased the CLz/F value by approximately 0.7-fold. Additionally, apatinib significantly increased the Cmax value of oxycodone. These findings suggest that icotinib and apatinib enhance the exposure to oxycodone in rats, and icotinib has a significant impact on oxycodone clearance in vivo. This indicates that icotinib and apatinib likely exert inhibitory effects, possibly through their strong inhibition of liver enzyme CYP3A4, thereby leading to decreased enzyme activity and inhibition of oxycodone metabolism. Previous literature reports have shown that the toxic blood concentration of oxycodone is 0.06 mg/L  . This discrepancy may explain why apatinib exhibited strong inhibitory effects in vitro but a weaker inhibitory ability in vivo.To further investigate the inhibitory effects of these two inhibitors on oxycodone, we examined their inhibitory mechanisms in RLMs and HLMs. As for the experimental design, adding ice-cold acetonitrile directly to the sample can effectively terminate the reaction. However, in experiments with large samples, this procedure may not be feasible in a timely manner. We conducted comparisons and found that the results obtained with the freezing method had a smoother linearity compared to adding ice-cold acetonitrile directly. The findings revealed that in RLMs, both icotinib and apatinib inhibited the metabolism of oxycodone through a mixed mechanism. In HLMs, icotinib acted as a competitive inhibitor, while apatinib acted as a mixed inhibitor. Interestingly, there was inconsistency between their inhibitory abilities 0.12 μM) , which dIn summary, this study examined the interaction between two TKIs (icotinib and apatinib) and oxycodone. The results showed that both TKIs inhibited the metabolism of oxycodone in RLMs and HLMs, leading to the changes in its pharmacokinetic characteristics. Thus, cancer patients may experience adverse reactions of varying degrees when oxycodone is used alongside icotinib or apatinib. In clinical practice, it is important to carefully consider and monitor the concurrent use of these drugs, as the pharmacokinetics of oxycodone could be significantly altered even at therapeutic doses. Therefore, the dosage of oxycodone should be appropriately reduced. However, it is important to note that this study focused primarily on the impact of genetic polymorphisms and drug interactions on oxycodone metabolism and did not take gender into account as an additional variable. Hence, the findings of this experiment can only be applied to male individuals. Overall, our study provides accurate guidance for the combined use of oxycodone and TKIs such as icotinib or apatinib.10.7717/peerj.16601/supp-1Supplemental Information 1Click here for additional data file.


Cement-augmentation pedicle screws have been widely used in spinal internal fixation surgery combined with osteoporosis in recent years, which can significantly improve the fixation strength, but compared with conventional methods, whether it has more advantages is still inconclusive of evidencebased medicine. To systematically evaluate the efficacy and safety of cement-augmented pedicle screw in the treatment of thoracolumbar degenerative diseases with osteoporosis.We searched PubMed, Embase, and Cochrane Library for studies published from the establishment of the database up until June 2023. We included studies that concerning the cement-augmented pedicle screw and the traditional pedicle screw placement for thoracolumbar degenerative diseases with osteoporosis. We excluded repeated publication, researches without full text, incomplete information or inability to conduct data extraction and animal experiments, case report, reviews and systematic reviews. STATA 15.1 software was used to analyze the data.P > 0.05).A total of 12 studies were included in this meta-analysis. The sample size of patients were totally 881, of which, 492 patients in cement-augmented screw group and 389 patients in conventional screw group. Meta-analysis results showed that Japanese Orthopaedic Association (JOA) score , intervertebral space height  and post-operation fusion rate  were higher in the cement-augmented screw group than those in the conventional screw group. Operation time was longer in the cement-augmented screw group than that in the conventional screw group . Screw loosening rate was lower in the cement-augmented screw group than those in the conventional screw group . However, hospitalization time, intraoperative blood loss and Visual analog scale (VAS) score were not significantly different between the two groups (Compared with conventional pedicle screw placement, cement-augmented pedicle screw is more effective in the treatment of osteoporotic thoracolumbar degenerative disease by improving fusion rate and interbody height, reducing the incidence of screw loosening, and elevating long-term efficacy.The online version contains supplementary material available at 10.1186/s13018-023-04077-w. Thoracolumbar degenerative diseases are common diseases in middle-aged and elderly people, which often cause patients with low back pain, lower extremity pain and limited movement. Finally, the pedicle screw system is required for internal fixation to improve spinal stability and alleviate symptoms, which brings serious mental and economic burden to patients , 2 With With the development of science and technology, spinal surgery techniques have been continuously improved, among which methods to improve the stability of pedicle screws have also been increasing. At present, three methods are mainly used: (1) Using bone cement to strengthen pedicle screws ; (2) IncCement-augmented pedicle screw technology is one of the common techniques to improve the stability of internal fixation of pedicle screw. Compared with traditional pedicle screw fixation, this technique can effectively improve the success and rate of internal fixation. However, the comparison of the clinical efficacy of the treatment of osteoporotic thoracic and lumbar degenerative diseases is still lack of evidence-based medical conclusion . In thisInclusion criteria: the study type is retrospective or prospective study; studies that reports the cement-augmented pedicle screw and the traditional pedicle screw placement for thoracolumbar degenerative diseases with osteoporosis; the language is limited to English.Exclusion criteria: repeated publication; studies without full text, incomplete information or inability to conduct data extraction; animal experiments; case report; reviews and systematic reviews.In this meta-analysis, we searched Pubmed, Embase, Cochrane Library from establishment of the database to June, 2023. The search terms are as follows: (“osteoporosis”) AND (“pedicle screw” OR “fixation”) AND (“cement” OR “polymethyl methacrylate” OR “PMMA” OR “augmentation”).Two researchers independently carried out literature search, screening and information extraction. When a question or dispute arises, a decision is made after discussion or negotiation with a third person. The data extraction included the author, publication year of articles, study design, sample size, age, sex and outcomes including hospitalization time (day), operation time (minute), intraoperative blood loss (ml), Japanese Orthopaedic Association (JOA) score, Visual analog scale (VAS) score, intervertebral space height, fusion rate and screw loosening rate.The Newcastle–Ottawa Scale (NOS) for evaluating the quality of published literature is carried out separately by two academics , and it I2 and Q test were used to evaluate heterogeneity. If the heterogeneity test is P ≥ 0.1 and I2 ≤ 50%, it indicates that there is homogeneity between studies, and the fixed effects model is used for combined analysis; if P < 0.1 or I2 > 50%, it indicates that there is heterogeneity and sensitivity analysis was used to find the source of heterogeneity. If the heterogeneity is still large, use the random effects model or give up the combination of results and use descriptive analysis. Funnel plot was used to assess the publication bias.All data analyzed by STATA 15.1  . WeighteIn this meta-analysis, a total of 763 studies were retrieved from the database including Pubmed, Embase and Cochrane Library. After eliminating duplicate studies, 394 were obtained. After browsing titles and abstracts, 123 studies were obtained. Finally, 6 articles were included in the meta-analysis . The pooled results showed that the difference in hospitalization time between the cementaugmented screw group and the conventional screw group was not statistically significant  . The pooled results show that the operation time of the cement augmented screw group was significantly longer than that of the conventional screw group  . The pooled results showed that the difference in intraoperative blood loss between the cement augmented screw group and the conventional screw group was not statistically significant  . The pooled results show that the JOA score of the cement augmented screw group was significantly higher than that of the conventional screw group  . The pooled results showed that the difference in VAS score between the cement augmented screw group and the conventional screw group was not statistically significant   Fig. .Fig. 6FoI2 = 0.0%, P = 0.480). The pooled results show that the intervertebral space height of the cement augmented screw group was significantly higher than that of the conventional screw group   Fig. .Fig. 7FoI2 = 0.0%, P = 0.480). The pooled results show that the post-operation fusion rate of the cement augmented screw group was significantly higher than that of the conventional screw group  . The pooled results show that the screw loosening rate of the cement augmented screw group was significantly lower than that of the conventional screw group  , while the JOA after cement treatment with fenestrated pedicle screws was significantly higher than that of conventional screw treatment   Fig. .Fig. 10DP = 0.476), or fenestrated pedicle screws  and conventional screw treatment in JOA were all not statistically significant  or fenestrated pedicle screws  were all significantly higher than that of conventional screw treatment  and below 0.6 g/cm2 for early screw loosening [The development of medical technology has led to the continuous aging of the population, and the lack of calcium supplements, sedentary and smoking, and the increasing number of the elderly population, leading to the increasing number of people suffering from osteoporosis. In osteoporosis patients with lumbar disc herniation, spinal stenosis, lumbar spondylolisthesis instability and scoliosis, and other degenerative diseases, spinal fusion and fixation with pedicle screw system is one of the main treatment methods . Howeveroosening .At present, many scholars use a variety of ways and technology efforts to improve the stability of spinal fixation, such as some scholars in the original fixed, on the basis of supplementary anterior fixation, laminar hook and install transverse connector, and more scholars to enhance pedicle screw, pull force, reduce pine, nail and other related complications risk, constantly improve technology . Bone ceA total of 12 articles were included in this study, and the results of meta-analysis showed that the operation time of the bone cement-strengthened pedicle screw group was more than that of the traditional pedicle screw group, and considering the cement-reinforced pedicle screw group increased the cement injection link, the number of fluoroscopy required also increased significantly, which in turn increased the operation time to a certain extent. There was no difference between the two groups in terms of intraoperative blood loss, hospital stay and postoperative drainage, considering that spinal canal decompression was basically concentrated in spinal canal decompression with spinal surgery bleeding and greater damage to patients, while surgical bleeding and injury were closely related to postoperative recovery, so the recent postoperative recovery speed of the two was similar, resulting in the same length of hospital stay .JOA and VAS are common criteria for evaluating the efficacy of spinal surgery . In thisIn addition, in terms of maintaining the height of intervertebral space, improving the fusion rate and reducing the screw loosening rate, the cement-reinforced pedicle screw group was still better than the conventional pedicle screw group. This shows that the use of bone cement strengthening technology can effectively improve the strength of the vertebral body, reduce the risk of loosening and nail retraction, achieve strong internal fixation, reduce the collapse of the final stub, better maintain the height of the intervertebral space, and provide a good and stable mechanical environment for intervertebral fusion, improve the fusion efficiency , and achFurthermore, we performed subgroup analyses based on cement strengthening methods. The summary results suggest that in the analysis of JOA and fusion rate, cement treatment with fenestrated pedicle screws had higher postoperative JOA and fusion rate than cement treatment with conventional pedicle screws, indicating that fenestrated pedicle screws may be a better treatment and could be further promoted in the future.This meta-analysis has several limitations. First, all of the studies included in this research were cohort studies. The literature quality is lower than that of randomized controlled trials, which may lead to selection bias. In addition, the included sample size was too small, and some studies had large statistical heterogeneity, resulting in reduced the reliability of the analysis results. The conclusions obtained in this paper need to be further verified by more rigorous high-quality, large-sample clinical studies.Compared with conventional pedicle screw placement, cement-augmented pedicle screw is more effective in the treatment of osteoporotic thoracolumbar degenerative disease by improving fusion rate and interbody height, reducing the incidence of screw loosening, and elevating long-term efficacy.Additional file 1. Plots of sensitivity analysis.


Salt bridges, π–π stacking, π–cations, and hydrogen bonds contributed to the high binding characteristics of the inhibitors and ACE. Introducing eight Trp into rabbit skeletal muscle protein (no Trp in wide sequence) endowed the protein with a more than 90% ACE inhibition rate, further suggesting that meat with a high content of Trp could have potential utility in hypertension regulation. This study provides a clear direction for the development and screening of ACE inhibitory peptides.Chronic diseases, such as hypertension, cause great harm to human health. Conventional drugs have promising therapeutic effects, but also cause significant side effects. Food-sourced angiotensin-converting enzyme (ACE) inhibitory peptides are an excellent therapeutic alternative to pharmaceuticals, as they have fewer side effects. However, there is no systematic and effective screening method for ACE inhibitory peptides, and the lack of understanding of the sequence characteristics and molecular mechanism of these inhibitory peptides poses a major obstacle to the development of ACE inhibitory peptides. Through systematically calculating the binding effects of 160,000 tetrapeptides with ACE by molecular docking, we found that peptides with Tyr, Phe, His, Arg, and especially Trp were the characteristic amino acids of ACE inhibitory peptides. The tetrapeptides of WWNW, WRQF, WFRV, YYWK, WWDW, and WWTY rank in the top 10 peptides exhibiting significantly high ACE inhibiting behaviors, with IC Hypertension  is a chrThe antihypertensive drugs currently used in clinical practice have different degrees of side effects, such as captopril  and ramiPan et al.  used molBy evaluating the inhibitory effect of 8000 tripeptides, our group, Chen et al. , showed 50 values determination, and validation of ACE inhibitory activity of the hydrolysed rabbit-derived protein indicated that peptides containing Trp, Tyr, Phe, His, Arg, and especially Trp at the C- and N-terminal ends, had high ACE inhibitory activity. Therefore, we have successfully established an efficient and accurate screening strategy for ACE inhibitory peptides. The sequence characteristics of the ACE inhibitory tetrapeptide were systematically characterized for the first time, and proved to be correct by calculations and experiments. This study provides a clear direction for the development and screening of ACE inhibitory peptides.In this study, the ACE inhibitory effects of 160,000 tetrapeptides were systematically evaluated by molecular docking for the first time, and Tyr, Phe, His, Arg, and especially Trp were found to be the characteristic amino acids of ACE inhibitory peptides by statistical analysis. Molecular dynamics simulations, binding free energy calculations, interaction relationship analysis, in vitro ICFirst, the Protein Preparation Wizard module in Schrödinger ,28 was uTetrapeptides were selected for association analysis by RapidMiner Studio version 7.4. In order to fit association rule studies, physical properties of the amino acids were assigned as a dataset. The data (CSV file format) were imported to workflow processing by using RapidMiner’s import operator. The main process in this workflow contains FP-growth and creates association rules, which algorithms run on Rapidminer. The prediction association rules were evaluated by using support and confidence as the main filtering parameters in the system. The strength of an association rule can be measured in terms of its support and confidence. Association rules are created by analysing data for frequent if/then patterns, and using the criteria support and confidence to identify the most important relationships. Support is an indication of how frequently the items appear in the database. Confidence indicates the number of times the if/then statements were found to be true. The formal definitions of these metrics are:2) constraint. The SHAKE algorithm [The structure of the complex generated by Schrödinger docking was subjected to parallel MD simulations to investigate the binding mechanism of ACE and the tetrapeptide. The cationic dummy atom (CaDA) ,30 was ulgorithm  was usedCalculation of the free energy of binding, consisting of enthalpy and entropy changes, was performed using the MM/GBSA and IE methods, respectively.gasG and ∆solvG represent gas-phase energy and solvation free energy, respectively. Among them, ∆gasG consists of electrostatic interaction and van der Waals (vdW) interaction, as follows:The MM/GBSA method, one of the most widely-used methods, can be expressed by the following equation:solvG contains polar and nonpolar solvation free energy of two parts, as follows:∆gbG is calculated by the GB module, and the nonpolar solvation free energy ∆npG is calculated by:The polar solvation free energy ∆SASA represents the solvent-accessible surface area, and is calculated by the MSMS program. The γ and β are set to 0.005 kcal·(mol∙Å2)−1 and 0.00 kcal·mol−1, respectively.The The IE formula is derived from the free energy of binding in the gas phase.The contribution of electrostatic forces to the free energy of polar solvation is calculated using the GB model, where igb = 2.All the amino acids on ACE within 5 Å of the tetrapeptide were mutated to alanine. Potential ACE inhibitory peptides, selected based on the docking results, were solid-phase synthesized from Bankpeptide Biological Technology Co., Ltd. The purity (>95%) of these peptides was verified by RP-HPLC on an Inertsil ODS-SP column . Sequences were verified by mass spectrometry .50 refers to the semi-inhibitory concentration; here, the principle of in vitro IC50 determination of the potential ACE inhibitory peptide is that ACE can hydrolyse HHL into marena and dipeptide. HHL and tetrapeptide are competitive, and when ACE binds to the tetrapeptide, it will reduce the production of the product marena; the IC50 activity of the tetrapeptide is when it is reduced by half of the original concentration. The water bath temperature was controlled at 37 °C, and the initial mass concentration of each tetrapeptide was 2 mg/L. The tetrapeptide was diluted with borate buffer solution into 7 concentration gradients, and coincubated with HHL buffer for 5 min. Then, the ACE solution was added to initiate the reaction, and the reaction was terminated by the addition of HCl after 30 min of system reaction. Afterwards, the reaction was analysed on a C18 column, with HPLC detection at 228 nm and an injection volume of 10 μL. The mobile phases were water and acetonitrile at 75% and 25%, respectively, with a flow rate of 1 mL/min, and isocratic elution for 7 min (acetonitrile and water containing 0.1% trifluoroacetic acid). ACE inhibition was calculated as ACE inhibition = ((b − c) − (a − c))/(b − c) × 100% , and the inhibition rate was converted into IC50 values by the software graphpad prism 7.0.ICE. coli DH5α and E. coli BL21(DE3) cells , which were used as the host for cloning, and for the expression of chymotrypsin and the mutants with higher Trp content. Primers were ordered by Personalbio Co., Ltd. . The plasmid extraction kits, Gel/PCR Purification Kit and Seamless Cloning Kit, were provided by Solarbio Science and Technology Co., Ltd. . Substrate peptides were synthesized by ZhuanPeptide Inc. . All other commercial chemicals were provided by the China National Pharmaceutical Group Corporation.The pET-22b (−) plasmid  was used for subcloning E. coli BL21(DE3). The steps are as follows: (1) Single colonies of good growth on the transformed plates are picked and inoculated into 5 mL LB medium (containing 100 μg/mL ampicillin), and incubated for 12 h at 37 °C, 200 rpm. (2) Inoculate the culture in (1) with 1% of the inoculum into 50 mL LB medium (containing 100 μg/mL ampicillin), and incubate for 8 h at 37 °C, 220 rpm, in a baffled conical flask. (3) Inoculate the culture in (2) with 1% of the inoculum into 500 mL LB medium (containing 100 μg/mL ampicillin), and incubate in a baffled conical flask at 37 °C, 220 rpm, until OD600 = 0.8–1. (4) Add induction agent IPTG to (3) for a final concentration of 0.1 mM, induce at 20 °C, 200 rpm, for 12 h. (5) Take (4) and centrifuge at 8000× g for 5 min to collect the bacterium, and discard the supernatant. (6) Resuspend the bacterium in 0.9% saline, centrifuge again, discard the supernatant, repeat twice, resuspend with 50 mL PBS buffer (pH 7.4), and sonicate for 30 min . Centrifuge at 18,000× g and 4 °C for 10 min, and filter the supernatant by 0.22 μm filter membrane; the filtrate is used for subsequent purification. (7) Flush 10 column volumes with ultrapure water, flush 10 column volumes with equilibrium solution, top sample the crushed bacterial solution at a flow rate of 1 mL/min, and collect the perforated solution. (8) Rinse the nickel column sequentially with 25 mM imidazole buffer, 150 mM imidazole buffer, and 300 mM imidazole buffer, collect the eluate, and desalinate and concentrate the purified protein and store at −80 °C.The rabbit-derived skeletal myosin light chain C and its mutants with higher Trp content were expressed by g for 20 min, and filtered through a 0.22 μm filter. The optimum temperature for chymotrypsin was 30 °C. After hydrolysis for 2 h, the enzyme was inactivated by increasing the temperature to 95 °C, while the hydrolysis product remained stable at this temperature. The substrate HHL was broken down into hippuric acid (Hip) and the dipeptide His-Leu in the presence of ACE. The product hippuric acid showed maximum absorption at 228 nm, and was detected by HPLC on a C18 column. Incubation for 30 min at 37 °C was followed by the addition of 50 µL HCl to terminate the reaction. The hydrolysis products were first subjected to reductive alkylation, after which the treated samples were analysed by liquid–mass spectrometry (LC–MS/MS) to obtain the raw mass spectrometric results, which were analysed by the software Byonic to match the data and obtain the identification results.The purified rabbit skeletal muscle actomyosin and tryptophan mutants were dissolved in 0.1 M PBS buffer (pH 7.5). Proteins, at 5 mg/mL, with 25 μg/mL chymotrypsin at 37 °C for 2 h, were centrifuged at 20,000× In this study, 160,000 rational conformations of tetrapeptides were created for the first time by Python and ChemScript scripts. Using Zn (II) in the ACE structure (PDB ID: 1o8a) as the docking centre, 160,000 complex structures of tetrapeptides with ACE were obtained, with scoring functions ranging from −12.125 to −1.536 . A lower50 values  contained six Trp, and mutant 2 (Mut 2) contained eight Trp, and their sequences were compared in proteins . WT, Mutproteins C. Considproteins  further Food-derived ACE inhibitory peptides are excellent substitutes for highly effective drugs with many side effects. Efficient and accurate screening strategies, and knowledge of the sequence characteristics of inhibitory peptides and related mechanisms, are essential for the screening and development of ACE inhibitory peptides. In this paper, the low energy complex conformations of ACE with 160,000 tetrapeptides were systematically generated by molecular docking, and then, the ACE inhibitory effect of 160,000 tetrapeptides were preliminarily evaluated by a docking scoring function. The possible characteristics of the ACE inhibiting peptides were statistically analysed, and the presence of Trp, Tyr, Phe, His, or Arg residues at each site of the tetrapeptide had a strong ACE inhibitory effect, especially when Trp appeared at the C- and N-terminal. The top 10 and the last 2 ACE–tetrapeptide complexes were further analysed by molecular dynamics simulations, as well as MM/GBSA and IE methods. By calculating the binding free energy and analysing the possible reasons for the inhibitory effect of the top-ranked tetrapeptides containing these features, Van der Waals forces were found to play a major role in scoring the top-ranked complexes. The hot spot residue decomposition by the AS-GBIE method revealed that residues, including some binding pockets and some outside the binding pockets, contributed mainly to the binding of the tetrapeptides, including Q281, H353, W357, E411, H513, Y520, R522, and Y523. π–π stacking and π–cations also contributed to the stabilization of the structure. However, the tetrapeptides with a low docking score could not form most of the mentioned interactions that stabilize the complex conformation, and thus could not easily bind to ACE to form a stable complex for ACE inhibition.50 assay. It was found that the top 10 tetrapeptides all had a significant inhibition effect on ACE, with different degrees of action. This result was also consistent with the calculations, suggesting that the results of the 160,000 tetrapeptide systematic docking analysis were valid. Potential ACE tetrapeptide inhibitory peptides may prefer amino acids Trp, Tyr, Phe, His, or Arg at each site. In vitro hydrolysis experiments of rabbit-derived skeletal myosin light chain C demonstrated that the antihypertensive capacity of the protein was activated with increasing Trp content in the sequence. Thus, Trp is extremely important in the sequence profile of ACE inhibitory peptides. Enzymatic hydrolysis of Trp-rich food-derived proteins is an important tool for developing ACE inhibitory peptides.The inhibition effects of the top 10 tetrapeptides and the last 2 tetrapeptides in the docking score were verified by in vitro ICIn summary, this article constructed a library of binding characteristics of all 160,000 tetrapeptides to ACE for the first time. Based on energy analysis, the sequence features of ACE inhibitory peptides were comprehensively and systematically elucidated. The molecular mechanism of ACE inhibitory peptide was clearly explained by calculation and experiment. This paper showed the direction for the screening and development of ACE inhibitory peptides.


The geology beneath Thwaites Glacier, the Antarctic glacial catchment most vulnerable to climate change, is unknown. Thwaites Glacier lies within the West Antarctic Rift System, but details of the subglacial geology relevant to glacial flow, including sediment availability, underlying lithology, and heat flux, are lacking. We present the first sketch map of the subglacial geology of Thwaites Glacier, interpreted from maps of airborne gravity, magnetic and radar data, supported by 2D models and 3D inversion of subsurface properties, and the regional geological context. A zone of Cretaceous mafic magmatism extending ~200 km inland from the coast is interpreted, while sedimentary basins are restricted to a region 150 to 200 km inboard of the coast, underlying just 20% of the catchment. Several granitic subglacial highlands are identified, forming long-lived topographic highs. Our geological interpretation places constraints on the basal properties of Thwaites Glacier, laying the foundation for both improved predictions of ice sheet change and studies of West Antarctic tectonics. Geological sketch map provides insight into the basal conditions controlling the flow of the massive Thwaites Glacier. The West Antarctic Ice Sheet (WAIS) has been losing mass at an accelerating rate and is feared to be on a similar trajectory of rapid retreat as it was in previous warm periods  has seenThwaites Glacier differs from other WAIS ice streams in that ice generally flows across, rather than along, the tectonic fabric  province of West Antarctica . Before During the later stage of continental rifting, as the system moved toward continental breakup, it is suggested that a series of mafic magmatic bodies were emplaced along the Amundsen Sea and Zealandia conjugate margins, as revealed by linear magnetic anomalies  (21). InAfter the main Cretaceous rifting events, West Antarctica underwent further continental extension related to oceanic spreading within the adjacent Adare Basin 43 to 26 Ma  (22). ThP- and S-wave velocities, relative to the model mean, beneath the MBL Dome are interpreted to reflect upwelling of warm asthenospheric mantle, while further inboard faster velocities suggest cooler mantle   methods suggest that sources are typically between ~0 and 5 km below the ice-bed interface, with isolated estimates at 8 km or below . Tilt deDepth-to-magnetic source results from profile data show a broad scatter of solutions .The 2D forward models of upper crustal structure fit the observed Airy isostatic gravity and magnetic anomalies with source bodies ~6 km thick, with their tops located close to the ice bed, broadly consistent with depth-to-source solutions . Bodies −3 and 60 × 10−3 in the International System of Units (SI) suggests that within the inversion domain, 10 to 14% of the crust shallower than 10 km can be attributed to the high magnetic susceptibility sources. Given the nonunique nature of inversion results, this volume must be considered with caution. However, our maps show that magnetic sources underlie approximately 20% of the inversion region by area, and forward models suggest that bodies are ~6 km thick, giving an alternative but consistent estimate of ~12% of the upper 10 km of crust being formed from high-susceptibility material.The results of the 3D inversion show relatively good agreement between the input and predicted magnetic and gravity fields (fig. S7). However, the amplitude of the negative magnetic anomaly between anomalies M1 and M2 is not matched as well as other parts of the model. This suggests that the inversion may require negative susceptibility in this region, indicative of a component of unmodeled magnetic remanence in this area. Estimates of magnetic source geometry and properties from 3D inversion generally predict high-susceptibility bodies lying within the area of the bodies used in the forward models, and broadly coincident with the locations of depth-to-source clusters  and 5A. We interpret four geological units based on our aerogeophysical maps and models . Each unThe first unit we interpret is a series of elongate but relatively restricted sedimentary basins between 80 and 200 km long and ~30 km wide . The preWeak and deformable till in the areas of low basal roughness is revealed by swath radar showing elongate till ridges known as Mega Scale Glacial Lineations (MSGLs) (fig. S8C) km thick , but thekm thick  confirmskm thick , in contA thick basin containing relatively poorly lithified sedimentary rocks underlying the areas of smooth bed provides a mechanism for how these regions gain and maintain their relatively smooth appearance . Poorly The deforming till layer is likely an actively evolving feature associated with the current ice sheet  (42). Th−3 denser than the background material) with relatively high magnetic susceptibility (48 × 10−3 to 70 × 10−3 SI). A mafic body would be a reasonable source for the modeled positive Airy isostatic gravity anomalies, given the required densities . ThThe next unit is interpreted as a series of felsic granitoid intrusions. The geophysical signatures defining this unit are negative Airy isostatic anomalies associated with elevated topography (Gn1 to Gn4) . ThThe final unit is interpreted as an intermediate composition igneous rock, i.e., not strongly mafic or felsic in composition. This unit corresponds to magnetic anomalies >200 nT with no clear association with Airy isostatic gravity anomalies (The age of the intermediate unit is unknown. The dominant grid east to west to northeast to southwest orientation contrasts with the trend of the proposed mafic intrusions (fig. S4), suggesting emplacement in a different tectonic setting. Leaky transfer faulting within a failed continental rift producing the mafic intrusions, as suggested on the conjugate margin . Both prThe geological sketch map we have produced is the first detailed picture of the subglacial geology of the Thwaites Glacier catchment. Our interpretation has wide-ranging implications for understanding how the region’s tectonic evolution affects the dynamics of the overlying ice sheet.−1. The location of these basins in the upper reaches of Thwaites Glacier is important as they can act as a source of deformable till capable of reducing basal friction and enhancing ice flow downstream . Our objective in this study is to create a series of geophysical maps and models unbiased by regional trends, or survey layout, enabling the underlying geology to be interpreted. Two aerogeophysical surveys called AGASEA and BBAS underpin our study, providing regional airborne magnetic, gravity, and radar data . A 30-km low-pass filter before extraction of the tilt angle zero contour helped isolate the dominant trends.The edges of magnetic sources were located using both tilt angle and TDX enhancements calculated from the gradients of the reduced-to-the-pole magnetic field . After leveling and continuation, the line free-air data had an internal crossover error of 2.94 mgal.−3 for rock, ice, and water, respectively. Where the density of the topography differs from the standard, gravity anomalies correlated with the topography will be retained in the Bouguer anomaly. These retained anomalies will be proportional to any discrepancy in density, allowing the effects to be correctly considered in subsequent modeling. We note that the topography beneath the ice shelves is based on inversion of gravity data, and interpretation of Bouguer and derived gravity anomalies over the ice shelves is not valid. However, our study area is onshore, and use of gravity-derived topography in adjacent areas, required to create a seamless Bouguer correction grid, will not distort the shape or amplitude of the onshore anomalies.The signature of the subglacial topography dominates the free-air gravity anomaly (fig. S5A). The gravity effect of the subglacial topography, ice sheet, and offshore bathymetry calculated from our topographic compilation is the Bouguer correction, for which we assumed standard densities of 2670, 915, and 1028 kgmThe Bouguer gravity anomaly map (fig. S5B) shows little short wavelength resemblance to the subglacial topography, indicating that local topographic effects have been well modeled. However, isostatic compensation of the ice and topographic loads by low-density material at depth creates the observed long wavelength Bouguer gravity anomalies anticorrelated with the topography. The gravity effect of this compensation is modeled assuming Airy isostatic balance and subtracted from the Bouguer gravity anomaly to give the final residual Airy isostatic anomaly. The Airy isostatic model assumes no lateral strength in the crust and that the modeled gravity anomaly results from deflection of the Moho beneath elevated topography about a fixed compensation depth of 29 km. Such a model is not intended to be a true and robust reflection of crustal thickness, or the compensation method; rather, it is intended to quickly capture the gravity effect of the density variations required to support the observed topography in a simple and repeatable manner. The derived residual Airy isostatic anomaly is theoretically due only to upper crustal density variations . For det−3 as the Bouguer correction used this value and is assumed to account for density contrasts at and above the ice-bed interface. Where the assumed Bouguer correction density is incorrect, topography will be associated with local residual gravity anomalies, which can be appropriately modeled. The Airy isostatic correction is assumed to account for gravity anomalies associated with variations in crustal thickness or other regional isostatic compensation mechanisms. Variations in Moho depth or mantle density are therefore not considered in our 2D models as their signatures have theoretically been removed. The lack of active source seismic constraints means that additional mid to lower crustal sources for the observed magnetic and gravity anomalies, or complex upper crustal geometries, cannot be justified. In addition, estimates of Curie depth in this region of between 10 and 20 km suggest that few mid to lower crustal magnetic sources are present (To test the hypothesis that bodies with both elevated density and elevated magnetic susceptibility can cause the spatially correlated anomalies beneath the trunk of Thwaites Glacier, we constructed 2D forward models of the residual Airy isostatic gravity and magnetic anomalies . This coAn alternative to manual forward modeling is 3D inversion, which recovers an optimum, but nonunique, 3D distribution of magnetic susceptibility or rock density giving modeled anomalies matching those observed. We used the VOXI 3D inversion module of the Geosoft software suite . For det