In the SFA-treated group, recall scores also improved (p < 0.01), and the positive outcomes were maintained for about 3 months (p < 0.05). The Apathy Scale scores did not show significant change in either the SFA-treated or non-treated groups. Our present results suggest that the SFA program encouraged continuous participation to cognitive intervention and was useful for ameliorating dysfunctions in visuospatial memory and recognition in patients with neurocognitive disorder.We attempted to clarify positive benefits in cognitive abilities and motivation during our cognitive intervention [structured floral arrangement (SFA) program] for patients with neurocognitive disorder due to stroke, traumatic brain injury (TBI), and other related disorders. In this SFA program, participants are required to arrange cut flowers and leaves on absorbent foam according to an instruction sheet. In a previous study of patients with schizophrenia, our SFA program encouraged participants and contributed to stimulating their visuospatial process and memory. Here, 27 patients with neurocognitive disorders participated in this study. Sixteen patients were assigned to an SFA-treated group and participated in six sessions during two phases plus to daily activities. Eleven non-treated patients engaged only daily activities during the same period. We compared Apathy Scale scores and neuropsychological scores between the SFA-treated and non-treated patients. Their mean attendance rate was more than 90% during the two phases. SFA-treated patients copied a Rey–Osterrieth complex figure more accurately than non-treated patients ( Various cognitive interventions have been developed to improve cognitive functions of patients with neurocognitive disorder due to stroke, traumatic brain injury (TBI), Alzheimer’s disease, and other-related disorders . For patCognitive stimulation therapy (CST) is also a well-established cognitive intervention for elderly people with neurocognitive disorder. Participants engage in reality orientation and other enjoyable activities such as word games, being creative, and sound recognition . GeneralIn addition to program content, continuous participation to cognitive intervention appears important in achieving significant outcomes . In homeWe have developed a structured floral arrangement (SFA) program, combining benefits of CT and horticultural therapy, in which participants were required to arrange cut flowers and leaves (foliages) on absorbent foam according to an instruction sheet. The SFA treatment program contributed to stimulating abilities of visuospatial process and spatial memory like CT, and it also encouraged continuous participation . In patiAs training for visuospatial ability, the SFA procedure resembles that of a pegboard  or blockTo clarify cognitive outcomes, the present study investigated change using group comparisons of visuospatial abilities with SFA treatments. We also attempted to clarify SFA treatments’ horticultural effects. For one group, real flowers, and leaves were used as materials and, for the other, colored sticks; then cognitive outcomes and motivation for the program were compared between groups. We believe that the present study of SFA contributes to further development of cognitive interventions for people with neurocognitive disorder.Table 1). The number of SFA-treated patients was about 1.5 times larger than the number of non-treated Control group, because the 16 SFA-treated patients subdivided into two subgroups.We recruited patients with neurocognitive disorders from 2 day-care facilities whom directors agreed to cooperate on our study. In one facility, the number of daily users was from 35 to 50, and about 20 in the other one. These users were suffered from neurocognitive dysfunctions. They engaged daily in activities including light exercise, fancywork, and horticultural work, were able to communicate with easy oral language, and were physically capable of light work. We introduced our research to all users older than 18 years who had no congenital cognitive dysfunctions in the two facilities, and recruited 27 patients who agreed to participate our research. The 27 patients were randomized into two groups  nearly equal in age, years of education, illness, and MMSE score (Table 1).After assignment to the SFA-treated group, 16 patients participated in the SFA treatments in addition to the daily program. They showed mild to moderate neurocognitive deficits due to TBI, stroke, or other diseases . In the Control group, 11 patients showed mild to moderate neurocognitive dysfunctions because of TBI, stroke, or herpes encephalitis. Mean scores of the Mini-Mental State Examination (MMSE) were 24.4 in the SFA and 23.4 in the Control group; about 40% of all same were recognized as having dementia according to the MMSE cut-off score . No signThis study was approved by the Ethics Committees of the University of Ibaraki Prefectural University of Health Sciences Hospital. We obtained written informed consent from all adult participants and from the parents of non-adult participants before participating in the study. When participants could not sign due to their brain dysfunction, we obtained written informed consent from their family members after confirming participants’ agreements.Figure 1 shows the SFA group’s experimental schedule, that is, two treatment phases at intervals of 2 weeks. In each phase, three sessions of SFA treatment (30–40 min) were conducted within 8 days. Neuropsychological tests were examined before and after each phase . The 16 SFA patients were divided randomly into two similar subgroups that did not differ in age, years of education, time since onset of illness, and MMSE scores. The first subgroup, flower–stick SFA patients (n = 7) participated with real flowers and leaves  in phase 1, and, in phase 2, they used colored sticks instead . The second subgroup, the stick–flower SFA patients (n = 9) first used sticks and then real flowers and leaves, in reverse order from the first group. Both groups’ total treatment times were the same. We prepared two floral arrangement patterns with differing levels of difficulty . Patients were instructed to make the easier pattern  on the first day of each phase and the more complex pattern  on the latter 2 days.Figure 2C), which guided the order and location to place each material on the foam . For example, as a fifth procedure of Pattern 1, a pink rose was to be placed vertically on the foam’s No. 5 center circle impression .Structured floral arrangement program treatments were conducted in a group setting according to the procedure described in During the treatment’s second half, patients were instructed to complete the arrangement by referring to the instruction sheet without any staff assistance. To evaluate patients’ arrangement skills, we photographed their work and recorded with a stopwatch the time they took to finish the second arrangement on each SFA treatment session. After each SFA treatment with flowers and leaves, patients took their arrangements home with them.Figure 1: pre-1, post-1, pre-2, and post-2). The Rey–Osterrieth complex figure test was administered only to the SFA-treated patients about 3 months after phase 2 as a follow-up test. Control group patients spent their time as usual at the day-care facility with daily activities and participated only in neuropsychological tests on the same schedule as the SFA group, except for the 3-month follow-up. Because we predicted that Rey scores improved only in the SFA-treated group during two test phases, we did not plan the following test in the Control group. Using the Apathy Scale   and the hy Scale  before aThe SFA program’s mean attendance rates were 91.7% (with flowers and leaves) and 95.8% (with sticks). We observed that almost all SFA-treated patients continued to try to complete their arrangements and were able to finish them alone in the treatment’s second half. Once in the second session, one TBI patient failed to finish an arrangement with flowers and leaves by himself during phase 1, but he was able to finish his work without any assistance the next treatment day.Figure 3A shows changes in copy scores on the Rey–Osterrieth complex figure test both in SFA-treated and Control groups during the two phases. We found significant interaction between groups (SFA and Control) and test periods  (p < 0.05). The SFA group’s mean scores were significantly higher than those of the Control group both in pre-2 and post-2; SFA-treated patients could draw the complex figure more correctly than Control patients during phase 2. On recall scores in the Rey–Osterrieth complex figure test, we also detected significant interaction between groups and test periods . The SFA group’s mean scores gradually improved through treatments, and their post-2 mean score was significantly higher than those of the other three tests (p < 0.01). We also found a significant trend between the mean scores of the SFA and Control groups in the post-2 .Figure 4 shows these patients’ mean Rey–Osterrieth complex scores at each test point . Copy scores showed no significant differences among test points . By contrast, the mean recall scores at post-2 and after 3 months were significantly higher than those at pre-1 . SFA-treated patients maintained improvement of visuospatial memory for 3 months. We analyzed the Rey–Osterrieth complex data separately between flower–stick (n = 7) and stick–flower (n = 9) SFA subgroups . No significant differences in copy  and recall  scores were found between subgroups. In other words, SFA materials  did not affect outcomes of the Rey–Osterrieth complex figure test.Eleven of 16 SFA patients were followed about 3 months after the final SFA treatment. Figure 6 shows drawing samples by three SFA-treated patients in the Rey–Osterrieth complex figure recall test. Case 1 was a 24-year-old female (MMSE = 28), who had right-brain lesion due to TBI 4 years before her SFA program participation (flower–stick subgroup). Her performance improved gradually with SFA participation, and she maintained that improvement for 3 months . Case 2 was a male with mild dementia (MMSE = 22), 48 years old; he suffered from multiple sclerosis onset 2 years before SFA participation (stick–flower subgroup), with bilateral brain damage. Although he performed well on the pre-1 Rey–Osterrieth complex figure copy test (score = 34/36), he revealed severe memory deficit in the recall version . His recall scores improved through SFA treatments, and on the 3-month follow-up test, he appeared to maintain improvement . Case 3 was a 34-year-old male with moderate cognitive dysfunction (MMSE = 10) due to TBI 2 years before SFA participation (flower–stick subgroup). His copy scores improved from 26.0 (pre-1) to 31.5 (post-2) points during treatment. Although recall scores improved in the two SFA phases, his scores declined after 2-week and 3-month intervals . Even in older patients, improvements were found in Rey–Osterrieth complex figure recall test. Case 4  recorded 31.5 points in the post-2 test, while he scored 8 points in the pre-1 test. In Case 5 , recall scores were increased from 19 to 28 points during SFA interventions. The improvements in recall scores were observed over a wide age range.Table 2 shows other tests’ mean scores. Scores on the digit span, block tapping, and the Apathy Scale did not reveal any significant changes in either the SFA or Control patients. From the Apathy Scale, Japanese version, each patient was recognized as having apathy syndrome at over 16 points  without any support in the intervention’s second half. Although his copy score was 32.5 points, his recall score (0.5 point) was the worst among SFA group participants in the Rey–Osterrieth complex figure pre-1 test. He finished two arrangement patterns well with sticks, which were all placed correctly on the marked absorbent foam . He also arranged flowers and leaves well , but he failed correctly to place two cut flowers (orange roses) in Pattern 2 .Almost all patients could arrange materials correctly in Pattern 1 treatments, but they made mistakes occasionally in Pattern 2, which had a more complex design. Table 3 shows mean performance times for completing arrangements alone in the SFA treatment’s second half. Mean performance time with flowers and leaves was significantly longer than that with sticks (p < 0.01), and participants required more time to finish the arrangement on the second and third days (Pattern 2) than on the first day (Pattern 1) during each phase (p < 0.05). As we expected, the Pattern 2 design required a higher load for participants than Pattern 1. In addition, arranging flowers and leaves was more difficult than arranging sticks. We divided performance time by the number of materials (flowers and leaves or sticks) in each program pattern and calculated mean times required for placing each piece of material on the foam’s impression (Table 4). Mean times with flowers and leaves were significantly longer than with sticks (p < 0.01), and the time did not change within a phase.We found positive cognitive outcomes with the SFA program in the Rey–Osterrieth complex figure test. SFA-treated patients could draw a complex figure more accurately than Control patients in the later phase. Patients also improved their visuospatial memory through SFA treatments, and positive outcomes were maintained for about 3 months. Even for a patient with severe memory deficits, visuospatial memory scores improved gradually during treatment. Thus, our results revealed that the SFA program enhanced visuospatial analysis, recognition, and memory. These results were consistent with our previous studies’ results  and resuThe SFA program’s attendance rate was high, and almost all patients with mild to moderate neuropsychological disorders could finish their arrangements alone during the second half of treatment. The SFA program may be appropriate for a broad range of patients with neurocognitive dysfunctions, and it appears to provide CST-like elements that encourage participation through enjoyable activities . In addiFigure 6, Case 3). Patients with relatively severe neurocognitive disorder  might tend to show decline in cognitive benefits after non-intervention intervals. Participants with severe cognitive dysfunctions may need continuous treatment to maintain cognitive outcomes. Further study is needed to clarify the relationship between levels of neurocognitive dysfunction and retainment of cognitive outcomes.Present results showed that eleven followed participants maintained their improved recall scores for 3 months on the Rey–Osterrieth complex figure test. By contrast, we reported a patient who showed reduced recall scores after a 3-month interval ; this is unique among visuospatial training tools including puzzles, peg-boards, and block designs  in creating their arrangements because they were able to finish by alone more complicated work on later treatment days.Some differences appeared between the present study’s results and those of the previous study with schizophrenia patients. Present results showed improvement in scores of Rey–Osterrieth complex figure test, recall version, but not in the block-tapping task, whereas the previous study showed improvement in block-tapping task scores, but not in the Rey–Osterrieth complex figure test, recall version . PatientAgainst our prediction, we did not detect significant improvement in Apathy Scale scores. However, nine SFA-treated patients maintained their positive mental state based on the Apathy Scale, while three of five SFA non-treated patients showed gradual psychic deterioration and became to be recognized as having apathy syndrome at the experiment’s final test. Perhaps the SFA intervention contributed to patients’ maintaining good mental health. There are some limitations in the present study. First, the SFA-treated patients continued to engage daily day-care activities during SFA interventions, so an interaction between the SFA program and daily activities would not be clear. Second, the present sample size was small especially with regard of the age, whereas we found stable benefits in SFA program treated patients who were from twenties to sixties. The cognitive benefits may be affected by the age and other related factors, such as brain reserve capacity, which designates the ability to maintain cognitive performance well in the face of age-related brain modifications and pathology . The braIn the SFA program, patients took the flower and leaf arrangements home with them. We did not investigate the natural work’s influence in each home. The natural materials facilitate communication among participants’ family members and would ameliorate relationships between patients and caregivers, similar to CST interventions . In the HM-K conceived and designed the study. HM-K, IK, and YY administered the cognitive intervention. HM-K, IK, and SM executed cognitive tests. HM-K analyzed the data and wrote the manuscript with the help of other members .IK is currently employed by company Sase Total Care Center Co., Ltd. The present study was executed independent of the company. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Benzimidazoimidazoquinazoline (OCT). The compound was characterised using FTIR, 1H and 13C NMR, DIMS, as well as X-ray crystallography. The most significant peak in the 13C NMR spectrum is C-7 at 65.5 ppm which confirms the cyclisation process. Crystal structure analysis revealed that the molecule grows in the monoclinic crystal system P21/n space group and stabilised by an intermolecular hydrogen bond between the N1–H1A…N3 atoms. The crystal packing analysis showed that the molecule adopts zig-zag one dimensional chains. Fluorescence study of OCT revealed that it produces blue light when expose to UV-light and its’ quantum yield equal to 26%. Antioxidant activity, which included DPPH· and ABTS·+ assays was also performed and statistical analysis was achieved via a paired T-test using Minitab 16 software with P < 0.05. Also, the antimicrobial assay against two Gram-positive, two Gram-negative, and one fungus was screened for these derivatives.A comparative study between two methods, , was performed to synthesise a new quinazoline derivative from 2-(2-aminophenyl)-1Using microwave to synthesise OCT have drastically reduced reaction time, and increased yield. OCT show good antioxidant activity in one of the tests and moderate antimicrobial activity.The online version of this article (10.1186/s13065-018-0509-z) contains supplementary material, which is available to authorized users. Also, the literature survey resulted to only one study that have focused on antioxidant activities of benzimidazoquinazoline compounds [c]quinazoline resulting from two different synthetic methods.Since 6-heptyl-5,6-dihydrobenzoimidazo+ fragment. The remainder of the fragments with their molecular weights is illustrated in Fig. The molecular ion peak was determined for OCT and is equivalent to its molecular weight , which is precisely equal to the calculated molecular weight and the other fragmentation peaks, are also displayed. This molecular ion also underwent α-cleavage to eliminate 6-heptyl moiety to produce a fragment at m/z 220 with 100% abundance as a base peak. Further, under the same type of cleavage, a radical ion at m/z 110 formed by cutting off C15H15N moiety. However, under inductive cleavage (i-cleavage), a radical ion at m/z 92 was formed via cutting C15H19N2 off, , b = 17.14 (5), c = 11.27 (4) Å, α = 90°, β = 101.5 (2)°, ɤ = 90°, z = 4 and volume = 1773 (11) Å3. The crystal system and refinement parameters are given in Table 6-Hepty5,6-dihydrobenzoimidazo1,2--cquinazoThe molecule is discrete, having only one molecule in the asymmetric unit. The heptyl group is attached to the diazine ring at C7 atom. The molecular structure with the numbering scheme is illustrated in Fig. The benzimidazole ring N2/N3/(C8–C14) is planar with a maximum deviation of 0.012 (5) Å and 0.012 (7) Å for C8 and C11, respectively from the least square plane. The benzene ring (C1–C6) is planar with a maximum deviation of 0.007 (5) Å for C1 from the least square plane. The dihedral angle between the benzimidazole plane and the benzene ring is 7.26 (17)°.The diazine ring, N1/N2/C1/C6/C7/C14 adopts half-chair conformation with a maximum deviation of 0.209 (5) Å for atom C7 from the least square plane Fig. .Fig. 10Tc]quinazoline imidazo[1,2-Additional file 1. Additional tables.


Investments to reduce the spread of antimicrobial resistance (AMR) in the European Union have been made, including efforts to strengthen prudent antimicrobial use. Using segmented regression, we report decreasing and stabilising trends in data reported to the European Surveillance of Antimicrobial Consumption Network and stabilising trends in data reported to the European Antimicrobial Resistance Surveillance Network. Our results could be an early indication of the effect of prioritising AMR on the public health agenda. Escherichia coli and Klebsiella pneumoniae.Antimicrobial resistance (AMR) is one of the main public health challenges worldwide. In the European Union (EU) and European Economic Area (EEA), 33,000 deaths are estimated to be attributed to infections with AMR bacteria annually . EnsurinThe European Antibiotic Awareness Day, a European health initiative and annual event coordinated by the European Centre for Disease Prevention and Control (ECDC), was initiated in 2008 to support EU/EEA countries in their efforts to prevent and control AMR by raising awareness about prudent use of antibiotics , (ii) a E. coli (2002–18) and K. pneumoniae (2006–18) isolates were analysed with the same joinpoint methodology. E. coli and K. pneumoniae are among the pathogens contributing most to the burden of AMR in the EU/EAA [Supplementary Material, including an additional sensitivity analysis.To examine trends, we analysed population-weighted EU/EEA-wide data from the European Surveillance of Antimicrobial Consumption Network  and the European Antimicrobial Resistance Surveillance Network . Using joinpoint version 4.7.0.0 , we performed a regression analysis of data reported to ESAC-Net/ESAC on defined daily doses (DDDs) as listed in the Anatomical Therapeutic Chemical (ATC) Index for 2019  on the ce EU/EAA . Joinpoie EU/EAA . MateriaFor antimicrobial consumption, the joinpoint model identified two separate trend segments for third-generation cephalosporin consumption . For theThe most recent trend segments, starting between 2008 and 2016 depending on the antimicrobial group and sector, all showed stable or decreasing trends, while segments covering earlier years showed larger variation -3. For cE. coli  or an increase but with a smaller slope compared with previous segments   . For almE. coli) -3.E. coli and K.pneumoniae was most prominent up to 2012/13, after which there is a tendency towards stabilising levels. These trends may be less discernible in shorter time series analyses and underline the usefulness of data retrieved from longer reporting periods.The decreasing or stabilising EU/EEA trends in antimicrobial consumption of fluoroquinolones, third-generation cephalosporins and carbapenems described in this study during the last 6–10 years could be an early signal of the positive effects of antimicrobial stewardship initiatives in EU/EEA countries. Although the ecological study design and different data sources used cannot demonstrate a causal relationship between antimicrobial consumption and AMR, the similar, but less pronounced, stabilising trends or change in slope of AMR percentages are encouraging. Our analysis shows that the increase in AMR in Our study has several limitations. First, for EARS-Net/EARSS, we could not control for changes in the use of clinical breakpoints over time . HoweverE. coli and K. pneumoniae, but they only contribute to approximately 14% of the total consumption of antibacterials for systemic use (ATC group J01) . Combined resistance to multiple antimicrobial groups increased during our study period, especially for K. pneumoniae [. In addition, changes in infection prevention and control (IPC) practices in hospitals and other healthcare settings in EU/EEA countries during the study period could have affected AMR trends, in particular for K. pneumoniae for which most infections are healthcare-associated. Longitudinal data on IPC in hospitals in the EU/EEA are not available to analyse such potential impact. Finally, the trends presented here represent the population-weighted mean for the EU/EEA as a whole and individual countries may report different long-term trends and changes, therefore, each EU/EEA country should assess its own situation.Several other factors than consumption of specific antimicrobial groups may have affected the corresponding AMR trends. The antimicrobial groups included in this study were selected as they are of relevance to treat severe infections caused by eumoniae . ConsequResponding to the increasing public health threat of AMR is a priority in the EU ,6,7. The


Despite the high consumption of dietary supplements (DS), few reliable, relevant, and comprehensive online resources could satisfy information seekers. This research study aims to understand consumer information needs on DS using topic modeling, and to evaluate accuracy in correctly identifying topics from social media. We retrieved 16,095 unique questions posted on Yahoo! Answers relating to 438 unique DS ingredients mentioned in sub-section, “Alternative medicine” under the section, “Health”. We implemented an unsupervised topic modeling method, Correlation Explanation (CorEx) to unveil the various topics in which consumers are most interested. We manually reviewed the keywords of all the 200 topics generated by CorEx and assigned them to 38 health-related categories, corresponding to 12 higher-level groups. We found high accuracy (90–100%) in identifying questions that correctly align with the selected topics. The results could guide us to generate a more comprehensive and structured DS resource based on consumers’ information needs. Dietary supplements (DS) usage has gained popularity in recent years with almost 52% of U.S. adults reporting the use of one or more supplement . This hiIn recent years, the internet has emerged as an important source of health-related information providing an opportunity for people to search online for free health information. According to a Pew Research Center report, 80% of internet users have looked for health information online , 4. ThisThe rapid growth of digital data in today’s world, especially in the healthcare domain, offers great opportunities for secondary use in clinical research. Topic modeling  has beenThere are various social Questions and Answers (Q&A) sites and online forums within health communities, e.g., Yahoo! Answers, allowing one to seek information through posting questions and receiving answers from others users  . PreviouThe purpose of this research study is to understand the information needs of DS consumers by analyzing questions coming directly from consumers and in their own language. The goal is achieved by using Correlation Explanation (CorEx) - a topic modeling algorithm on the title and body of each question under the Q&A section of the Yahoo! Answers database in order to unveil the “topics” around DS information needs. We generated a list of coherent topics that more accurately represent the areas of DS-related information and associated DS ingredients that consumers are most interested in. We will also evaluate the accuracy of the CorEx method in correctly identifying the topics from social media. In the future, the knowledge gained from this study could be used as a guide for developing more meaningful DS resources for consumers that are better aligned with their information needs.We collected in total 2,820,179 Yahoo! Answer questions and the corresponding answers posted under 21 sub-categories belonging to the main category “Health”. We further extracted 112,090 questions (including their titles and contents) from one of the sub-categories “Alternative Medicine”. We then matched the preferred DS names in “iDISK”, the first Integrated Dietary Supplements Knowledge base where DS related information is represented in a comprehensive and standardized form , with thThese questions were further pre-processed by subject matter experts  and used for topic modeling. We removed all ingredient mentions within the questions to understand the information needs non-specific to certain DS. Each question was then lower-cased and tokenized. Special characters, hyperlinks, and common stop-words  were removed, and each word was normalized using the normalized string generator (Norm) from the SPECIALIST NLP tool . We onlyIn our preliminary investigation of different topic modeling strategies, we found that Correlation Explanation (CorEx)  discoverThe selected model was further analyzed and assigned topic names after mutual agreement between two assessors . The “topics” with similar themes were then merged into “categories”  that were further condensed into higher level “groups” . For the group, “uses and symptoms”, we utilized System Organ Classification (SOC) created by the Medical Dictionary for Regulatory Activities (MedDRA), a medical terminology used to classify adverse event information associated with the use of biopharmaceuticals and other medical products .To evaluate the accuracy of the topic modeling, we selected 15 topics and extracted their corresponding 10 questions with highest ranked probabilities. Manual review  was conducted to determine if the extracted questions correctly aligned with topics generated by the above topic modeling methods. The measure of correctness was reported as percentage accuracy. We also extracted the DS ingredient names corresponding to each topic in order to explore the distribution of ingredients names across various topics. We also reported the DS ingredients associated with most questions for selected topics.The final list consisted of 438 unique DS terms in total associated with 16,095 unique matching questions. After data pre-processing, our corpus contained a total of 213,790 tokens, which made up of 7,164 unique words.From the 200 topics generated by CorEx modeling method, the domain experts  identified topics with similar themes and classified them into 38 unique categories by . The 38 After evaluating the top 10 ranked questions for selected topics, we reported accuracy as number and percentage of questions that correctly aligns with the generated topic. We found the percent accuracy for most of the selected topics is between 90% - 100%, except sleep (80%) and frequency/time categories (70%). “Use and adverse effects” is the most dominant topic group and accounted for 50 topics out of 200. Under this topic, there were 15 categories classified based on MedDRA SOC .We also extracted the DS ingredient names associated with most questions corresponding to a particular topic in order to explore the distribution of most commonly discussed ingredients. Only DS ingredients associated with ≥10% of questions under a specific topic were reported .In this study, we employed CorEx topic modeling over user-generated questions coming from the Yahoo! Answers data in order to better understand the information needs of consumers. We also discovered interesting information on the distribution of DS ingredients across topics of special interest to consumers. This research effort further validates the feasibility of topic modeling to extract important information hidden in large corpus of social media data.Applying CorEx topic modeling methods, we were able to accurately identify 12 topic groups. The top three groups with the most number of respective assigned categories and topics, which can be regarded as the information most sought by consumers, are: “use and adverse effects”, “product-related”, and “healthy life style” . ExtractWe found high accuracy when we identified questions that correctly align with the topic categories/groups . We founThis research study had several limitations. We analyzed only questions belonging to alternative medicine sub-category under “health” section and might have missed dietary supplement occurrences under other sub-categories, e.g., mental health conditions, general health care. We only used preferred DS ingredient names and not their synonyms  to extract the corresponding questions. Also, there are inherent limitations to topic modeling e.g., topics were generated based on the statistical word distribution within the questions and thus topics with incoherent topic keywords were also generated.This research provides essential insights on extracting and understanding the information needs of consumers around dietary supplements using CorEx-based topic modeling that could identify the relevant topics embedded in a large corpus of Yahoo! Answers data with high accuracy. The knowledge gained here could be used to generate a more comprehensive repository of resource for consumers around dietary supplements usage. Thus, this study is an important contribution in further accentuating the potential benefits of using social media data in the clinical research.


Final year students also have more knowledge (13×) and awareness of contributory factors (3×) on AMR (p = 0.001) than other students. Unsatisfactory knowledge on AMR issues exists among veterinary students yet willingness to improve was observed. Identified knowledge, attitude, and gaps in AMR awareness should be targeted by veterinary schools in Nigeria.The challenge of antimicrobial resistance (AMR) is grave in developing countries. Antimicrobials are misused yet stakeholders’ contribution to antimicrobial stewardship is low. Veterinary students are future prescribers and their knowledge could influence progress in combating AMR; hence, there is a need to assess their knowledge, attitude, and awareness of AMR. A multi-institutional questionnaire was administered to undergraduates in Nigerian veterinary schools. It comprised demographics, own personal antibiotic usage, and knowledge, attitude, and awareness of AMR in humans and animals. Descriptive statistics and logistic regression were used for analyses. Of the 426 respondents, 39.2% reported personal antimicrobial use in the previous six months. Over 60% received knowledge scores lower than average and >87% requested more education on clinical use and prescriptions pre-graduation, monitored dispensing of antimicrobials, conducting AMR research, and confirmed link among human, animal, and environmental health. Less than 25% of respondents were aware of antimicrobial stewardship and global efforts/organizations for AMR. Final year students have 9-fold and 14-fold more satisfactory knowledge on antimicrobials in humans and animals compared with other students, respectively ( Globally, antimicrobial resistance (AMR) is a complex public, animal, and environmental health problem, primarily fueled by inappropriate use of antimicrobials. The trends and problems associated with AMR are observed naturally in pathogenic and commensal or non-pathogenic bacterial organisms ,2. CurreHuge antimicrobial consumption and misuse are prevalent in some low- and middle-income countries, such as Nigeria ,11. OverSpecifically, veterinarians are very important stakeholders and play a vital role in the mitigation of AMR ,19,20,21The scope of AMR in the academic training of undergraduate veterinary students in most Nigerian schools is not clearly defined. A previous survey of final year students in five conveniently selected veterinary schools in Nigeria indicated poor knowledge of AMR . In thisUndergraduate students in 10 of the 12 Nigerian universities that award the DVM degree were the target population. The selected universities include: University of Ilorin (UNILORIN); Federal University of Agriculture, Abeokuta (FUNAAB); University of Maiduguri, Maiduguri (UNIMAID); Usmanu Danfodiyo University, Sokoto (UDUS); University of Abuja, Abuja (UNIABUJA); Ahmadu Bello University, Zaria (ABU); University of Ibadan, Ibadan (UI); University of Agriculture, Makurdi (UAM); University of Jos, Jos (UNIJOS); and Michael Okpara University of Agriculture, Umudike (MOUAU) . NigeriaA cross-sectional multi-institutional questionnaire survey among veterinary students in Nigeria was carried out online from August to September 2018. The sample size formula for the cross-sectional study (random sample)  was usedThis instrument comprised questions on demographics, antibiotic usage, and knowledge, attitude, and awareness on antimicrobial resistance in humans and animals. We conducted a literature search ,29,30 anTo assess participants’ attitudes towards AMR, we utilized a 16-item questionnaire section. Respondents’ attitudes on the effect of AMR on human and animal health and production were surveyed. Attitudinal questions about respondents having sufficient knowledge on antibiotic use for future clinical practice, need for more education on AMR at the clinical levels, poor infection control practices by veterinarians and other health care professionals as causes of the introduction and spread of AMR, following existing guidelines on the future use of antimicrobials, prescription of antibiotics in closely controlled environment, dispensing of antibiotics without prescription or over the counter, and the link between human, animal, and environmental health, in terms of AMR were asked.Respondents were also asked about their general awareness of key terms, such as superbugs, AMR, antimicrobial stewardship, World Antibiotic Awareness Week, The Global Antimicrobial Resistance Surveillance System (GLASS), Global Antibiotic Research and Development Partnership (GARDP), Global Action Plan on Antimicrobial Resistance, and National Action Plan for Antimicrobial Resistance, Nigeria.https://forms.gle/X9yEDCsHwq87gunS9). This online survey was accessible from 17 August 2018, 3:10:59 PM GMT+1 to 8 September 2018, 12:24:30 PM GMT+1. Informed consent was obtained. Respondents were allowed to participate in the survey voluntarily (not under group setting) and had the opportunity to withdraw from participation at any period without bias based on recommendations of the 2013 World Medical Association Declaration of Helsinki Ethical principles for medical research involving human subjects [The questionnaire was pretested on five undergraduate DVM students from UNILORIN. Feedbacks received were used to standardize the questions before opening the online survey nationwide, for use by respondents (subjects . The Ethp < 0.05) were further subjected to stepwise backward likelihood multivariate logistic regression analysis to determine possible factors influencing knowledge levels among undergraduate veterinary students in Nigerian universities. Universities with <15 respondents were excluded from the multivariate logistic regression. A dependent variable was created from the thematic questions below, which were used to determine the knowledge levels of Nigerian undergraduate DVM students on antibiotic use and AMR; related questions (variables) were categorized together into various outcomes: (1) Antimicrobial resistance in humans; (2) antimicrobial resistance in animals; (3) contributory factors to antimicrobial resistance; and (4) overall knowledge on antimicrobial resistance. A numeric scoring system was used [Collected data were initially summarized in Microsoft Excel 2016 and analyzed using Open Source Epidemiologic Statistics for Public Health (OpenEpi), version 3.03a . Descripwas used  to assesA total of 426 participants (age: 23.08 ± 2.82 years) from 10 universities across Nigeria completed the questionnaire. The pattern of antibiotic usage among veterinary undergraduate students in Nigeria is presented in p < 0.001) than other age categories (p = 0. 043) and UNIABUJA  were two times and 11 times less likely to possess satisfactory knowledge of AMR than students in other universities in Nigeria. Satisfactory levels of knowledge of AMR tend to increase relative to the year of study of respondents (two < three < four < five < six) (p < 001) and animals . These students were also more likely to be aware of the contributory factors of AMR prevalence , and have general knowledge of AMR  than students in other levels.On the knowledge scores of students on AMR, general knowledge scores of AMR were low and ranged from 11 to 37 . Overalltegories . In the e < six) . ClinicaRespondents in this survey demonstrated positive attitudes toward AMR control . HoweverWith regard to the key terms and popular statements used in AMR fora, the awareness level among veterinary students appeared low . Only 18This multi-institutional national survey investigated the knowledge and attitude of undergraduate veterinary students on antibiotic use and antimicrobial resistance with key findings. It provides an insight into the importance of possible points of intervention. Whereas veterinary students demonstrated good personal use of antibiotics, the majority had an unsatisfactory level of knowledge of AMR. Veterinary students’ academic level of study and age groupings are important factors associated with a satisfactory knowledge of AMR. It should be understood that the higher-level classes roughly correlated with higher ages and specific training in pharmacology, microbiology, and medicine, and these may have influenced the observations above. Although the respondents demonstrated positive attitudes toward tackling AMR, their awareness of key terms and concepts of AMR is low. These findings are key to understanding the situation of antibiotic use in humans and animals as well as important to evaluate knowledge of AMR among veterinary students in Nigeria. Since these students are the future prescribers for the nation’s veterinary  and food industries, they remain important stakeholders who will be playing pertinent roles in combating AMR in food animals and the environment in the future.While over half of the respondents (64.6%) reported that the last time they used antibiotics was over six months back, based on these self-reports, the awareness of compliance with proper antibiotic usage in humans is heightened. Previously, undergraduate pharmacy students in Sri Lanka have similarly demonstrated a high degree of compliance with antibiotic use . In the The gender distribution of respondents to the questionnaire in this survey corresponds to the gender aggregation of students of veterinary medicine in Nigerian universities. Overall, the surveyed students demonstrated an unsatisfactory level of knowledge of AMR, particularly in areas of knowledge of the use of antibiotics, AMR in humans and animals, and on the contributory factors to incidences of AMR. These findings signify a deficiency in aspects of the clinical training curricula, especially microbiology, pharmacology, and medicine, or lack of integration of knowledge as well as poor exposure of Nigerian veterinary students to associated outcomes of AMR. Comparably, in another survey, low knowledge about AMR was documented among final year veterinary students It was not surprising that the senior-level students possess higher knowledge levels on AMR compared to others. In this study, satisfactory knowledge of AMR increased with an increasing academic year of the veterinary studentship. Clinical-level students (particularly those in year six) were generally better than the other students. However, more year 5 students (34.5%) had satisfactory knowledge scores of AMR in animals than year 6 students (22.4%). A possible reason could be that year 5 students have a better memory of pharmacology since in most veterinary schools in Nigeria, pharmacology lessons are concluded in year 5 with clinical or applied pharmacology. Previous evaluation, even among the senior-level non-medical students, showed that they have appreciable knowledge on AMR than those in lower levels . SakeenaRespondents to our survey exhibited positive attitudes toward AMR control. A similar observation has been observed among undergraduate paramedical students . The resOn key terms in AMR, only 42.3% of respondents were aware of the term “AMR”. This is confirmed by the previous observation of Anyanwu et al. . SimilarA major limitation of this study was the use of an online questionnaire tool for data collection, which may be associated with enrolment bias. However, the overall response rate is sufficient to eliminate or reduce this bias. Because of the peculiar situation of some institutions, some students could have had limited access to the online questionnaire due to poor availability and limited access to the internet, and this possibly affected the uneven distribution of respondents from various schools. Also, it is harder to draw probability samples based on e-mail addresses or website visitations as well as a lack of trained interviewers to clarify and probe responses. Nevertheless, it was believed that the pre-testing before actual data collection should improve the accuracy and internal quality control of the tool. The logistic regression model was run with year two students serving as the reference against other years of study. Only one year two student had satisfactory knowledge of AMR in animals, resulting in a small sample size of year two students with satisfactory knowledge. Thus, this affected the outcome resulting in a wide confidence interval for students in higher years of study (4–6) with more satisfactory knowledge scores of AMR in animals. Also, wording options, such as “inadequacy of drug regulation agencies”, sounds extremely bias, possibly influencing students to click on them. However, only about 58% of respondents picked these options. Thus, the bias was unpronounced as envisaged.This study expounded the unsatisfactory knowledge level of AMR issues existing among veterinary students in Nigeria, yet willingness to improve was observed. Identified knowledge, attitude, and awareness of AMR gaps should be targeted by veterinary schools in combatting AMR issues in Nigeria.


Plastic bronchitis is the expectoration of bronchial casts in the mold of the tracheobronchial tree. It is a rare occurrence of unknown etiology that has been primarily described in children with congenital heart disease. In this case report, we present the first reported case of plastic bronchitis in a patient with pulmonary Kaposi sarcoma and underlying HIV infection. Plastic bronchitis (PB) is the formation of casts in the mold of the tracheobronchial tree, leading to airway obstruction. It has been described in association with a variety of coexisting disorders, but most commonly in children with cyanotic congenital heart disease following the Fontan procedure. To our knowledge, we describe the first case of PB in the setting of human immunodeficiency virus (HIV) infection and pulmonary Kaposi sarcoma (KS).μL, viral load 307 copies/mL on antiretroviral therapy) and pulmonary KS on chemotherapy presented with progressive dyspnea and cough productive of rubbery red and white material ), and a percussion vest were unsuccessful. Nebulized ipratropium and albuterol and supplemental oxygen by nasal cannula afforded occasional symptomatic relief. Multiple bronchoscopic procedures were performed to remove the fibrinous material from the lung, but it quickly reaccumulated. The expectorated material did not dissolve with tissue plasminogen activator (TPA)In some cases, PB has been due to lymphatic leakage into the bronchi either from surgical trauma or pulmonary lymphatic abnormalities, with resolution of the condition after ligation of the thoracic duct . In thisOver approximately three months, the patient was repeatedly readmitted for respiratory distress and ultimately required endotracheal intubation and mechanical ventilation. Repeated bronchoscopy was performed in an effort to clear the casts, but it was unsuccessful. While on the ventilator, he empirically received multiple therapies for the reduction of lymphatic flow , midodrine, and octreotide) and the treatment of KS (with sirolimus) to curb cast production. Unfortunately, the patient developed refractory respiratory failure and was transitioned to comfort measures. An autopsy revealed extensive pulmonary KS with hepatization of the lung and near obliteration of the normal alveolar architecture with copious mucin and cellular debris within the airways Figures , and 8. venae arteriosae expectorantii, which translates literally to “expectorated arteries and veins” [ ventilgeraeusch (“sound of a fan”) or bruit de drapeau (“sound of a flag snapping”) may be present and indicate subtotal airway obstruction [Plastic bronchitis is a rare condition of unclear pathogenesis characterized by the expectoration of casts in the mold of the tracheobronchial tree. It was first described by Galen in the second century AD asd veins” , 2. Sincd veins” –5, allerd veins” , cystic d veins” , influend veins” , and pned veins” ); and did veins” –13). Patd veins” . In a mitruction . Seear etruction , 16. Thetruction , 18 and truction , 20, andtruction . On the truction , 22, or The development of PB has not been previously described in patients with KS, an angioproliferative disorder of the vascular and lymphatic endothelium. KS is well known to cause lymphatic obstruction and lymphedema of the extremities but lymphatic stasis in the lungs has not been well described , 24. OthPlastic bronchitis, although rare, is a life-threatening condition that has been seen in multiple underlying pulmonary conditions. This case report highlights an unusual cause of PB and the need for further investigation into its pathogenesis and therapeutic measures.


Molecular-logic based computation (MLBC) has grown by accumulating many examples of combinational logic gates and a few sequential variants. In spite of many inspirations being available in biology, there are virtually no examples of MLBC in chemistry where sequential and combinational operations are integrated. Here we report a simple alcohol-ketone redox interconversion which switches a macrocycle between a large or small cavity, with erect aromatic walls which create a deep hydrophobic space or with collapsed walls respectively. Small aromatic guests can be captured or released in an all or none manner upon chemical command. During capture, the fluorescence of the alcohol macrocycle is quenched via fluorescent photoinduced electron transfer switching, meaning that its occupancy state is self-indicated. This represents a chemically-driven RS Flip-Flop, one of whose outputs is fed into an INHIBIT gate. Processing of outputs from memory stores is seen in the injection of packaged neurotransmitters into synaptic clefts for onward neural signalling. Overall, capture-release phenomena from discrete supermolecules now have a Boolean basis. While many processes in biological cells can be understood in terms of molecular logic gates that process information sequentially and combinationally, the design and construction of such devices in the laboratory are unknown. Here the authors achieve this by the reversibly-controlled capture and release of guest molecules from host containers. Indeed, a molecular memory carrying a downstream processor is unknown. Nevertheless, inspiration can be found in stored neurotransmitters being released into the synaptic cleft23 for further neural processing of the original signal. Although molecular versions of computer memories or RS flip-flops32 are known, all-chemical versions where the device, inputs and outputs are chemical in nature are unavailable.Molecular-logic-based computation35. Molecules can also be imprisoned/liberated44 but it is harder to achieve in a reversible all-or-none manner without complications of side-reactions40 or incomplete conversion. On the other hand, related intramolecular studies are available as parts of molecular machines49.Reversible atom capture/release is well-established23 to previously unaffected areas of chemistry, e.g. cleanup of pollutants, generation of chemical signals50 and programmed release of functional species from discrete supermolecules. Logical release (but not capture) from materials (but not discrete molecules) is available51. Digitally capturing/releasing a molecular guest from a molecular host due to molecular inputs shows the ability to create sequential logic devices with input–output homogeneity for gate concatenation, which is a way information is processed inside biological cells. In the present instance, the memory component is serially integrated with an INHIBIT gate whose final output is fluorescence emission. It is also remarkable that all the output states of the device are optically self-indicating. The current work also demonstrates a useful supramolecular mechanism for reversible guest capture/release in water by erecting/collapsing phenylene ring walls in a p-cyclophane by chemical redox conversions. Reduction is the set(S) input which gives the output of guest capture. Oxidation is the reset(R) input which produces the output of guest release. The ideas underlying the RS flip-flop are explained in the Discussion section below.We now present a molecular memory carrying a downstream processor. In this case, it allows us to introduce an all-chemical RS flip-flop. This permits logically controlled reversible capture/release of molecules both in aqueous solution and in the solid state. This pushes the relevance of molecular-logic-based computation1/2 and 3/4  and 1,5-diiodopentane37. NaBH4 reduction52 and work-up of 1 gives 2. Importantly, 1/2 can be derivatized so that biocompatible water-soluble versions 3/4 become available. This begins with selective monoiodination at ortho-positions with respect to the oxygens of all four phenylene rings of 1 to give 6 in good yield53. Each iodo substituent is converted to a carboxylic acid moiety to give 3 in good yield by Pd0-catalyzed carbonylation54. NaBH4 reduction in water to the dialcohol tetracarboxylic acid 4 completes the synthesis 55 converts dialcohols 2/4 to diketones 1/3. 4 is also oxidized to 3 with KMnO452 in water at 40 °C . The large cavity captures aromatic guests, but the small cavity does not, as demonstrated by X-ray crystallography  owing to the wall collapse, and no benzene occupant is found. Smaller molecules, e.g. dimethylformamide, are known to be included in some related macrocycles60, though in a disordered fashion59. Examination of the packing diagrams shows that benzene is found only in the internal cavity of 2 and it is completely absent from 1.In contrast to those concerning henone58  showing 2 contained one benzene occupant whereas the wall-collapsed 1 lost its occupant, we next examine aqueous solution-phase data of solubilized counterparts 4 and 3, respectively. No suitable crystals of 3 and 4 could be grown from solvents in which they were soluble. 4 and 3 are employed at levels which are below their critical aggregation concentrations  are evaluated as potential occupants. If the geometric fit is suitable, the dications of these p-xylyldiammonium salts  should be electrostatically attracted to the tetraanionic macrocycle cavity, besides some favourable hydrophobic attractions between the central parts of both the hosts and guests. The p-xylyldiammonium unit is stable under our redox conditions. The 1H NMR spectra of 7 with/without 3 or 4 are shown in Fig. δ) are observed for 7 in the presence of dialcohol 4, but not in the presence of diketone 3. The corresponding Δδ values for dialcohol 4 in the presence of 7 are shown in the inset in Fig. δ values, which is evidence for an axial inclusion complex in the solution state, are similar to those seen for classical examples of cyclophane inclusion complexes62, but with some differences. No significant chemical shift differences are found for diketone 3 in the presence of 7. Somewhat analogous to the X-ray crystallography studies , inclusive complexation is clear between 4 and 7, and no binding is observed for 3 and 7 in spite of ion-pairing in both cases. Although p-cyclophane hosts are known to be shape-dependent binders37, reversible all-or-none switching between binding and releasing states in water has not been demonstrated until now. The complexation between 4 and 7–9 is quantified by measurement of concentration-dependent Δδ values to obtain binding constants (log β) which have values around 4  to the INHIBIT logic gate  due to energetically low-lying nπ* excited states69. The capturing states have low ε values of 3900 M−1 cm−1 at 290 nm owing to the smaller π-delocalization of the benzhydrol moieties, while the ΦFlu value is low (0.004) when the guest 7 is bound and high  when guest-free. All three output states, i.e. vacant capturing state, occupied capturing state and releasing state, are therefore distinguishable by optical spectroscopy.Fluorescence spectra also illuminate the interaction between nce of 4 63. Thus,n64 Fig. . Since aate Fig. , which iIn3 Fig. . This is −0.2 eV . No exci48, we felt that we could apply our experience of molecular switching12 to make a contribution to this area. Excellent examples of switchable cyclophanes and related macrocycles already exist44, but binary all-or-none switching of cyclophanes in water still remains a worthwhile goal. Since cyclophanes are well-known to bind shape-complementary organic guests in water37, a method of switching the cavity size/shape should enable the guest’s release. Such guest capture/release would have immediate consequences for dosing of drugs and other bio-signalling agents, as well as environmental beneficiation by toxin removal. Notably, each of these interventions could remain under the user’s external control or could be triggered by an existing (e.g. intracellular) condition. The capabilities of such a switchable system could be made more intelligent if it could be endowed with self-indicating properties. Self-indication is key for nanometric molecules to communicate their status to their far-larger human handlers, i.e. to say when a task has been accomplished successfully18.Having been inspired by the roles played by cyclophanes in supramolecular chemistry4. Computer scientists and electrical engineers have dealt with such situations in larger-sized systems. This relationship between the disciplines is clear to us since we had the pleasure to introduce the field of molecular-logic-based computation10.Such relatively complex molecular-logic behaviour can benefit from the insights of computer science70 suggests that a guest cannot be corralled if the walls have fallen down for some reason. The present work shows that these human situations are mirrored in the molecular world.The inclusive complexation of a guest by a cyclophane can be seen, from some viewpoints, to be a case of a guest being corralled by a surrounding wall, with the corral itself being specified by the macrocycle and with the wall height being determined by the width of the aromatic rings. Notably, the walls are perpendicular to the corral area, i.e. the aromatic ring planes are orthogonal to the mean macrocycle plane. Human experience37, a prominent feature of our particular cyclophanes is the presence of a pair of ketones or secondary alcohols with flanking phenylene moieties. The benzhydrol version behaves as a more-or-less normal cyclophane host by orienting its phenylene ring planes orthogonal to the macrocycle plane, thereby creating a rather deep and hydrophobic cavity which can include an aromatic guest61. On the other hand, the benzophenone version has additional cross-conjugation due to the ketones flanked by the phenylene units. This flattens one phenylene of the pair beside each ketone so that the ketone and the phenylene form a plane which is nearly in the macrocycle plane. Such phenylene ring rotation closes off a large part of the macrocycle cavity. Thus a suitable aromatic guest which is inclusively complexed by the benzhydrol version is rejected by the corresponding benzophenone.Although cyclophanes of this type are well-known52, each of which are efficient unidirectional chemical reactions. When the ketone is treated with a reducing agent, the corresponding alcohol is formed and it stays as an alcohol whether excess reductant is present or whether the reductant is removed entirely. Similarly, when the alcohol is treated with an oxidant, the corresponding ketone is formed which persists whether the oxidant is absent or present in excess.It is well-appreciated that ketones and secondary alcohols can be interconverted by appropriate reduction and oxidation1 = 1, Out2 = 0 in Fig. 1 = 0, Out2 = 1 in Fig. From a computer science perspective, the ketone/alcohol system can exist in either of two stable states, depending on the input condition. If a high concentration of reductant is applied as the input, the resulting output state is the alcohol because of the reduction of any starting ketone. This input is called the set(S) input because it sets the alcohol as the output state  is only permitted for the benzhydrol 4 with its erected phenylene walls. An all-chemical RS flip-flop feeding an INHIBIT gate is demonstrated since chemical redox inputs applied to the p-cyclophane devices 1/2 or 3/4 permit reversible all-or-none capture or release of the guests where guest occupancy is self-indicated by the fluorescence output being quenched. This augurs well for the small-scale serial integration of sequential and combinational logic devices in the context of guest capture/release applications.In conclusion, X-ray crystallography shows that, when π-conjugation opportunities are depleted in benzhydrol derivative All the procedures are given in the 1 and 2 were obtained by evaporation of concentrated solutions of the compounds in benzene. Suitable crystals were selected and measured on a Rigaku Saturn724 + (2 × 2 bin mode) diffractometer. The crystals were kept at 100 K during data collection. Using Olex271, the structures were solved with the ShelXS72 structure solution programme using direct methods and refined with the ShelXL72 refinement package using least squares minimization. X-ray crystallographic data for 1 and 2, as well those for control compounds are detailed in the Single crystals of 4 as a function of added guest are detailed in the Fluorescence spectroscopic data obtained on a Perkin-Elmer LS55 spectrometer running FLWinlab software for Supplementary InformationPeer Review File


Synthetic molecular machines designed to operate on materials surfaces can convert energy into motion and they may be useful to incorporate into solid state devices. Here, we develop and characterize a multi-component molecular propeller that enables unidirectional rotations on a material surface when energized. Our propeller is composed of a rotator with three molecular blades linked via a ruthenium atom to a ratchet-shaped molecular gear. Upon adsorption on a gold crystal surface, the two dimensional nature of the surface breaks the symmetry and left or right tilting of the molecular gear-teeth induces chirality. The molecular gear dictates the rotational direction of the propellers and step-wise rotations can be induced by applying an electric field or using inelastic tunneling electrons from a scanning tunneling microscope tip. By means of scanning tunneling microscope manipulation and imaging, the rotation steps of individual molecular propellers are directly visualized, which confirms the unidirectional rotations of both left and right handed molecular propellers into clockwise and anticlockwise directions respectively. Controlling the rotation direction of individual molecular machines requires precise design and manipulation. Here, the authors describe a surface-adsorbed molecular propeller that, upon excitation with a scanning tunneling microscope tip, can rotate clockwise or anticlockwise depending on its chirality, and directly visualize its stepwise rotation with STM images. Among them, molecular propellers have shapes similar to their macroscopic counterparts and they are formed by molecular scale blades that rotate along a shaft8. In nature, molecular propellers are vital in many biological applications ranging from the swimming of bacteria to intracellular transport10 but, unlike bio-molecular propellers, synthetic molecular propellers may be able to operate in much harsher environments12. However, the molecular machines can change their mechanical, and electronic properties upon adsorption on a surface, which can greatly impact their performance. In some cases, the surface can also impose chirality in the molecule as well13. Therefore, for designing molecular machines to be operated on surfaces, the effects of the surface should be considered, and if possible should be used as an advantage. Although there are a number of molecular systems having propeller shapes and functions that can be deposited on a solid surface14, further development of molecular machines demands exquisite and robust design of molecular propellers with precise control over their rotation direction.Synthetic molecular machines are fascinating and have a great promise to revolutionize scientific and technological fields. The immense interest on this research area is evident by the 2016 Nobel Prize in Chemistry awarded for the “design and synthesis of molecular machines”. To date, various molecular machines that can be operated on solid surfaces such as molecular vehicles and molecular motors have been developedHere, we develop a multi-component molecular propeller that enables unidirectional rotation suitable to operate on a material surface. An important aspect of molecular machine development is to test the properties and operations of individual molecular machines. We use a variety of scanning tunneling microscope (STM) tip manipulation schemes; electric field induced rotation, inelastic electron tunneling (IET) induced rotation, and rotation by mechanical force to investigate unidirectional rotation of the molecular propellers on one-molecule-at-a-time basis. In electric field induced rotations, the propeller rotates by using the electrical energy supplied from the STM tip. Here, the propeller can be rotated by the tip scanning with a high voltage above ±1 V during acquisition of images, as well as by positioning the tip statically above or beside the molecule. The threshold electric field required for the rotation, 0.25 V/Å, is determined from the slope of the linear relationship between the negative threshold voltage and the tip height. Using this value, the electrical energy stored in the molecular propeller is calculated as −0.66 eV. While a low current is used for the electric field induced rotation, IET induced rotations are performed with a high current above 2 nA at positive biases. The IET induced rotation is initiated by the transfer of electron energy from inelastic tunneling electrons via temporary electron attachment to the lowest unoccupied molecular orbital (LUMO) of the molecular propeller. The threshold energy for the IET induced rotation is determined as ~0.6 eV. Furthermore, mechanical rotations of both left and right handed propellers provide information on the detailed rotation mechanism, which reveals that swinging of the propeller blades occur before proceeding to a full rotation. Moreover, we are able to directly visualize the rotation steps of the individual propellers from the STM images acquired at each rotation steps on both left and right handed geometries. The statistical analyses gleaned from the STM images and mechanical rotation events further confirm the unidirectional rotation of the propeller. The experimental results are corroborated by density functional theory calculations, which reveal tilting of the stator phenyl rings that leads to the formation of a ratchet shape molecular gear.p-bromophenylene (Cp(PhBr)5) units covalently attached to it16. The Ru atom is coordinated to both the stator and to the three propeller blades  atom in a facial mode while retaining thioester (SEt) groups at opposite ends Fig. . The stades Fig. . For exp5 stator parallel to the surface plane. In our previous study5, the three propeller blades formed by the indazole groups have been used as a tripodal stator to anchor the molecular motor by preferentially binding the sulfur atoms from the three SEt groups on a Au(111) surface. In the current system however, the five bromophenyl groups of the Cp(PhBr)5 stator are preferentially positioned on a Au(111) surface and the propeller blades formed by the three indazole groups are located at the top with an angle of 120 degrees to each other  surface, large area STM images acquired at 80 K substrate temperature show individual molecular propellers preferentially located along the wider face centered cubic (FCC) regions of the herringbone surface reconstructions Fig. . The molher Fig. . Remarkaher Fig. . Thus, tInterestingly, the STM images of the molecular propellers reveal left or right handed chirality depending on the positioning of the three propeller blades Fig. . Statist18, either left or right. Here, the five phenyl-rings are tilted with different angles; 18°, 35°, 43.4°, 66.5°, and 76.7° respectively from the surface plane  surface, we have performed density functional theory (DFT) calculations. Geometrically relaxed calculations reveal that the Cp ring of the stator is located 3.7 Å above the top surface layer of Au(111) Fig. , and the11 surfacane Fig. , and theane Fig. . The DFTane Fig. . The strear Fig. . Owing tIndeed, stepwise rotation of the individual molecular propellers can be observed when the STM images are acquired with biases equal or greater than ±1 V. Fig. Since the three propeller blades are positioned on the 5-teeth ratchet gear formed by the stator there are 15 steps to complete a 360° rotation. This means that each step has a 24° rotation, and the propeller shown in Fig. The threshold bias required to rotate the propellers can be determined from the statistical analysis gleaned from the STM images acquired at different biases Fig. . In ordeTo check the rotational directions of both left and right-handed propellers, a statistical analysis for the rotations observed during STM imaging is performed. Here, only full step rotations of all three propeller blades are counted and from a total of 168 rotation events of both left and right-handed propellers, 132 rotations (78.6%) are observed as correct rotation directions, i.e., left-handed propellers rotate to the clockwise direction while the right-handed propellers rotate to the anticlockwise direction, respectively Fig. . Among tThe observed rotations during image acquisition invoke further questions concerning the required electric field strength and electrical energy stored in the molecular propeller. To determine the threshold electric field required for rotation, we perform controlled stepwise rotations of the propeller using STM tip-induced electric field manipulation as follows: Initially, the tip is positioned above the center of the propeller with a fixed height Fig. , and theThe electric field induced rotation can also be realized with the tip positioning next to the propeller. Figure 21. To trigger the IET process, the probability of electron capture by the molecule is important and a sufficient tunneling current is necessary. Unlike the electric field induced rotation where a low tunneling current (pA regime) and a high bias are used, the IET induced rotations are realized with a high current (greater then 2 nA) with a lower positive tunneling voltage of 0.6 V. The use of positive bias means that the electrons are tunneling from the tip to the unoccupied states of the sample in our case. Figures 5. This can be directly confirmed by a threshold energy measurement, which shows that ~0.6 V is necessary to trigger the IET manipulation while the dI-dV spectroscopy and the projected density of states unveil this energy as close to the energy of the LUMO orbital  processes22  surface next to a propeller blade and then the tip height is reduced for 3 Å to 4 Å from its initial position. Then the STM tip is laterally moved towards the propeller blade in a constant current mode, i.e., maintaining the tip height by the electronic feedback loop, until it mechanically pushes the blade22 Fig. . During From the mechanical rotation experiments, one can unravel the rotation mechanism as swinging the blade followed by passing the molecular gear teeth for each rotation step. Such mechanical manipulation could result in rotating just one propeller blade or the whole propeller. For instance, STM images of Fig. 23 borate in presence of thalium sulfate. The overall yield for 3 steps was 63%. The molecules were deposited onto atomically clean Au(111) sample using a custom built Knudsen source under an ultrahigh vacuum (UHV) environment. For the deposition, the molecular source was heated to ~450 K and the Au(111) substrate temperature was held at ~120 K. The sample was then transferred to a Createc GmbH STM scanner directly attached to the sample preparation chamber via a gate valve under UHV condition. The sample temperature was then lowered to 80 K and 5 K for respective experiments. For all the experiments, an electrochemically etched tungsten wire was used as the STM tip. The tip was prepared in-situ by gently dipping into the substrate prior to the manipulation experiments25 and thus it was assumed to cover with Au.The single crystal Au(111) sample was cleaned by a repeated sequence of ArThe STM tip induced electric field, inelastic tunneling electrons, and the mechanical rotations of the propellers were performed at 80 K substrate temperature. STM imaging was performed at 80 K and 5 K substrate temperatures, and a low tunneling current range between 10 pA and 70 pA was used. The mechanical rotations of the both chiral propellers with the STM tip were performed by using tunneling resistance range of 1.3 MΩ to 70 MΩ, the tunneling current range of 16 nA to 75 nA, and the bias range of 0.01 V to 0.1 V. The initial set point tunneling current and voltage before the manipulation were 50 pA, and ±1 V, respectively.26, with core electrons described by the projected augmented wave method27. Exchange-correlation was treated in the Generalized Gradient Approximation28. Because of the relative importance of non-bonding molecule surface interactions, van der Waals D3 functional was used29. The plane wave basis was expanded to a cutoff of 600 eV. The Au(111) surface was modeled by a three-layer slab containing 432 atoms and the first Brillouin zone was sampled at the Γ point only. The molecular propeller composed of 117 atoms was placed on top of a three-layer Au(111) slab representing the surface with a vacuum space of 20 Å. The geometry optimizations were converged within 2 meV per formula unit for the total energies.Spin polarized Density Functional Theory (DFT) calculations were carried out with the Vienna ab initio simulation package codeSupplementary InformationPeer Review FileDescription of Additional Supplementary FilesSupplementary Movie 1Supplementary Movie 2


Candida Antarctica lipase B (CALB) is a well-known enzyme, especially because of its promiscuous activity. Due to its properties, CALB was widely used as a benchmark for designing new catalysts for important organic reactions. The active site of CALB is very similar to that of soluble epoxide hydrolase (sEH) formed by a nucleophile-histidine-acid catalytic triad and an oxyanion hole typical for molecular structures derived from processes of α/β hydrolases. In this work we are exploring these similarities and proposing a Ser105Asp variant of CALB as a new catalyst for epoxide hydrolysis. In particular, the hydrolysis of the trans-diphenylpropene oxide (t-DPPO) is studied by means of quantum cluster models mimicking the active site of both enzymes. Our results, based on semi-empirical and DFT calculations, suggest that mutant Ser105Asp CALB is a good protein scaffold to be used for the bio-synthesis of chiral compounds. Epoxides are important molecules for producing chiral compounds. Because of their chemical versatility and ability to react readily with halides, carbon, nitrogen, oxygen, or sulfur nucleophiles, epoxides became crucial intermediate products in organic synthesis . EpoxideHerein we focus on the epoxide hydrolases  that serve as a catalyst to transform epoxide into the corresponding 1,2-diol by addition of a water molecule. In fact, mammalian EHs were widely studied mostly because of their biological functions. The main roles of EHs in mammalian organisms are detoxification, catabolism, and regulation of signaling molecules. However, EHs have mainly garnered interest because of their potential applications in chiral chemistry .in vitro purposes, the substrate is trans-diphenylpropene oxide (t-DPPO), and thus it will be used as a substrate in the present study. Recently, it was found that sEH plays an important role in the regulation of blood pressure and inflammation [Soluble epoxide hydrolase (sEH) hydrolyses a broad range of substrates such as gem-di-, trans-di-, cis-di, tri-, and tetra-substituted epoxides . Howeverammation ,11,12,13ammation ,15,16.sEH belongs to the α/β hydrolase fold family of proteins, which are characterized by a Nucleophile-His-Acid catalytic triad evolved to efficiently operate on substrates with different chemical composition or physicochemical properties and in various biological contexts . In the Candida antarctica lipase B (CALB), widely studied because of its high catalytic promiscuity [The structure of the sEH active site is very similar to other enzyme from the α/β hydrolases family named miscuity . The mecmiscuity  and theomiscuity , providimiscuity ,23,24. Mmiscuity . Interesmiscuity . In the The active site of CALB contains a Nucleophile-His-Acid catalytic triad formed by Ser105, His224, and Asp187. As in sEH, there is an oxyanion hole in the active site of CALB formed by Thr40 and Gln106 residues. The similarities between the sEH and CALB active sites are indicated on It is generally assumed that the mechanism of hydrolysis of epoxides catalyzed by sEH to the corresponding diols is known. It was demonstrated using experimental techniques that the epoxides can be opened by direct attack of a nucleophile on the epoxide ring or via an intermediate in which a covalent link between the enzyme and the substrate is formed. This variant of the mechanism, in which the nucleophile Asp333 was involved in the reaction, was recently confirmed by theoretical studies performed by Mulholland and co-workers . HoweverIn the case of hydrolysis of t-DPPO, attacks on both the C1 or C2 carbon atom of epoxide are possible, resulting in two different enantiomers, as presented in However, sEH shows regioselectivity by preferentially attacking the reactive C1 carbon atom rather than C2 with a 97:3 selectivity ratio . As was R,R)-t-DPPO in active sites of sEH and a variant of CALB is described using quantum cluster theoretical models.In the present work we are proposing a new catalyst for epoxide hydrolysis reaction based on the structure of the active site of CALB. In order to examine our proposal, the mechanism of the hydrolysis of  ,33,34,35wat–C1)-d(Oep°x–C1)) was used as one of the distinguished reaction coordinates. The other one was defined as the antisymmetric combination of distances defining the position of the transferred hydrogen atom between the oxygen atoms of the water molecule and the nitrogen atom in position ε of His523 (d(Owat–Hwat)-d(Nε–Hwat)).In the case of mechanism A, the nucleophilic attack can take place concurrently with the proton transfer from a water molecule to Nε-His523. This possibility was investigated by exploring the corresponding potential energy surface (PES), where the antisymmetric combination of distances defined between the oxygen of the water molecule and the carbon atom of epoxide, and between this carbon atom and the oxygen atom of epoxide (d(OAsp333–C1)-d(Oep°x–C1)) was used as one of the coordinates, while the antisymmetric combination of distances describing the transfer of hydrogen atom from oxygen of Asp333 to the nitrogen in position ε of His523 (d(OAsp333–HAsp333)-d(Nε–HAsp333)) were controlled as the second reaction coordinate. The resulting PESs are shown in −1 at AM1 level and 28.5 kcal·mol−1 after correction at M06-2X level. On the other side, mechanism B takes place through a step-wise process whose rate-determining step, the attack of the deprotonated oxygen atom of Asp333 to the carbon atom of the epoxide ring, presents an energy barrier of 39.5 kcal·mol−1 at AM1 level but is dramatically reduced to 20.3 kcal·mol−1 when recomputed at M06-2X/6-31+G level. Thus, our simulations predict mechanism B as the kinetically favorable mechanism and thus we will focus on the next steps of this mechanism. This conclusion is in agreement with experimental studies based on solvent KIEs that confirmed the role of Asp333 residue as the attacking nucleophile since the heavy 18O oxygen was incorporated into the Asp333 of sEH [−1) [−1 for β-methyl-styrene oxide [−1 for t-DPPO epoxide [In order to explore the second possibility of nucleophilic addition, the nucleophilic attack to C1 of t-DPPO(1) by Asp333, a similar PES was explored. In this case, the antisymmetric combination of distances between oxygen from Asp333 and the carbon of epoxide and between this carbon atom and the oxygen of epoxide (d(O3 of sEH . MoreovesEH [−1)  for the ne oxide ,33,34,35 epoxide . In our Based on the obtained energy barriers for the nucleophilic attack of Asp333 on either the C1 or C2 of epoxide based on our cluster models of the sEH active site, it is difficult to reach conclusions about its enantioselectivity. Comparing our energy barriers presented in After the first step of the reaction was studied, in which transformation of epoxide to oxyanion tetrahedral intermediate occurs, we focused on the second part of the mechanism, which is still an unsolved problem. The relevance of this step is evident since it was shown by experimental measurements that this step of the reaction, in which the oxyanion intermediate is transformed into a diol, seems to be the rate-limiting step.According to the obtained results after the nucleophilic addition, the oxyanion intermediate is formed and the protonation state of His523 is changed . The presence of a hydrogen atom in Nε-position does not allow His523 to activate a water molecule in order to attack the carbon of Asp333 and thus alternative scenarios, depicted in −1 at AM1 level and 57.3 kcal·mol−1 when corrected at M06-2X/6-31G+ level . Finally, the hydroxyl formation takes place in a single step in both mechanisms B1 or mechanism B2 & B3, with a potential energy barrier of 2.2 kcal·mol−1. All in all, the presented results allow us to conclude that hydroxyl formation from an oxyanion intermediate would preferentially occur through mechanism B2 or B3. The amount of tyrosinate formed during the nucleophilic attack was experimentally estimated to be 0.4 per enzyme molecule [The results obtained for mechanism B1 show that the nucleophilic attack of the water molecule on the carbonyl carbon atom of Asp333 takes place through a TS with an energy, related to the reactant complex, of 64.8 kcal·molevel see a. In them B2 see b, and thm B3 see c. Afterwm B3 see a with a molecule , indicatAs mentioned in the Introduction, the active site of CALB presents noticeable similarities with the active site of sEH. However, in the wild-type CALB the role of the nucleophile in its catalytic triad is played by a serine (Ser105) instead of an aspartate (Asp333) residue. In fact, as experimentally shown, the Asp333Ser variant of sEH loses its capability to catalyze the epoxide hydrolysis. Thus, it can be concluded that Ser105Asp CALB should perfectly mimic the active site of wild-type sEH. Apart from differences in the rest of the proteins’ scaffolds, the main difference between the active sites of sEH and Ser105Asp CALB is the residues involved in the formation of the oxyanion hole. Then, we can predict that the reaction in the latter should proceed through a path similar to mechanism B2 described for sEH. As we have already demonstrated, the role of the oxyanion hole in sEH is to stabilize the negatively charged oxygen of the intermediate, because the other possible role being a proton donor has already been excluded or does not present any energetic advantage (mechanism B3). Thus, the epoxide hydrolysis in the active site of wild-type CALB and Ser105Asp variant is studied assuming a reaction path similar to the most favorable one observed in sEH, mechanism B2.The resulting PESs of the nucleophilic attack to C1 carbon atom of t-DPPO(1) in wild-type and Ser105Asp CALB are shown in In order to compare the pathway of the reaction catalyzed by she or by the Ser105Asp variant of CALB, the rest of the steps from INT3 to PC were explored following the same strategy as the one employed in the study of mechanism B2 & B3 of the reaction catalyzed by sEH. The full energy profiles are summarized in −1·Å−1. It is important to note that no constraints were applied to any of the geometry optimizations. Although allowing more reliable energetics, this implies that possible artifacts, such as odd interaction complexes, can be obtained. Thus, proper orientation of the different structures in the starting point structures is a crucial step in the computational protocol. Also, keeping in mind that the reaction under study is a multi-step process, IRC calculations traced forward from a TS structure do not necessarily converge in the end of the backwards path traced from the following IRC. In this sense, efforts have been made to get a converged result; otherwise, the minimum energy structure, belonging to the reaction path, was selected. All AM1 calculations were performed with MOPAC2007 [The theoretical study of possible t-DPPO epoxide hydrolysis mechanisms has been performed within two different active sites models, as presented in OPAC2007 , while DOPAC2007 .et al. [The primary reaction of the sEH, the hydrolysis of epoxides, has been studied by means of the AM1 semi-empirical method and the M06-2X hybrid functional, using a cluster model to mimic its active site. Using this reaction as a template, the secondary activity of wild-type CALB as an epoxidase has been studied and the comparison with the reaction on sEH has suggested a mutation of one residue of the active site that could improve the catalytic activity of CALB. Indeed, our results carried out with t-DPPO as a substrate suggest that the multi-step mechanism that Ser105Asp employs to hydrolyze t-DPPO is similar to the one used by sEH. Our predicted rate limiting step of sEH is in agreement with the pre-steady-state kinetic analysis of epoxide hydration catalyzed by mEH performed by Tseng et al. , who conFrom the computational point of view, our results, obtained at the AM1 level, follow the same trend as the ones recomputed at a higher level of theory, M06-2X with the 6-31+G** basis set. These latter values are, in fact, quite close to the barriers that can be deduced from experimentally measured rate constant in related enzymatic processes. Nevertheless, it appears that the stereo-selectivity of the enzyme, as a result of the nucleophilic attack taking place on one or the other carbon atom of the epoxide, cannot be predicted from simulations based on reduced models. Thus, further studies should be carried out in the future with more complex and realistic models such as the ones based on the use of hybrid QM/MM models. Information derived from the present study will be of great help in setting up the model, in determining the size and level of theory to be employed in the QM region, and in the analysis of the role of the key residues of the active site.