{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,18]],"date-time":"2026-05-18T19:11:04Z","timestamp":1779131464034,"version":"3.51.4"},"update-to":[{"DOI":"10.12688\/f1000research.172637.1","type":"new_version","label":"New version","source":"publisher","updated":{"date-parts":[[2026,4,29]],"date-time":"2026-04-29T00:00:00Z","timestamp":1777420800000}}],"reference-count":41,"publisher":"F1000 Research Ltd","license":[{"start":{"date-parts":[[2026,4,29]],"date-time":"2026-04-29T00:00:00Z","timestamp":1777420800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["f1000research.com"],"crossmark-restriction":false},"short-container-title":["F1000Res"],"abstract":"<ns3:p>\n                    Background Diabetes mellitus (DM) is often associated with male sexual and reproductive dysfunction, including erectile impairment, reduced libido, hormonal disturbances, and poor semen quality. Black pepper (\n                    <ns3:italic>Piper nigrum<\/ns3:italic>\n                    L.) fruit extract has shown antidiabetic and reproductive benefits in experimental studies, but its role in diabetes-related sexual dysfunction remains unclear. Objective To evaluate the effects of black pepper fruit extract on hormonal, erectile, libido, sperm, histological, and molecular parameters in alloxan-induced diabetic male rats. Methods Thirty male Sprague Dawley rats were assigned to five groups: normal control, diabetic control, diabetic rats treated with black pepper extract at 122.5 or 245\u00a0mg\/kg body weight (BW) for 8 consecutive days, and diabetic rats treated with sildenafil citrate 1\u00a0mg\/kg BW. Outcomes included intratesticular testosterone (ITT), erectile parameters (quick flip, long flip, erection, total penile reflexes [TPR]), libido parameters (courtship latency [CL], mount latency [ML], mount frequency [MF]), sperm parameters (concentration, progressive motility, normal morphology), Leydig cell and spermatocyte counts, and eNOS mRNA expression. Data were analyzed using the Kruskal-Wallis test followed by Dunn\u2019s post hoc test. Results Compared with diabetic controls, the 122.5\u00a0mg\/kg BW black pepper group significantly improved TPR, CL, ML, and MF. The 245\u00a0mg\/kg BW group significantly improved ITT, sperm concentration, and normal sperm morphology. Both black pepper doses increased Leydig cell counts. Sildenafil significantly improved TPR, sperm concentration, progressive motility, and normal morphology. eNOS mRNA expression was higher in all treatment groups than in diabetic controls. Conclusion Black pepper fruit extract showed dose-dependent effects in alloxan-induced diabetic rats. The lower dose was more beneficial for erectile and libido-related parameters, whereas the higher dose had greater effects on ITT and selected sperm outcomes. These findings support its potential as a preclinical candidate for diabetic male sexual and reproductive dysfunction.\n                  <\/ns3:p>","DOI":"10.12688\/f1000research.172637.2","type":"journal-article","created":{"date-parts":[[2026,4,29]],"date-time":"2026-04-29T09:24:37Z","timestamp":1777454677000},"page":"38","update-policy":"https:\/\/doi.org\/10.12688\/f1000research.crossmark-policy","source":"Crossref","is-referenced-by-count":0,"title":["Effects of Black Pepper (Piper nigrum L.) Fruit Extract on Sexual Function, Hormonal Parameters, Sperm Parameters, Testicular Histology, and eNOS Expression in Alloxan-Induced Diabetic Rats"],"prefix":"10.12688","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0009-0003-0960-4521","authenticated-orcid":false,"given":"Exsa","family":"Hadibrata","sequence":"first","affiliation":[{"name":"Doctoral Program Bandar Lampung Lampung Universitas Lampung Fakultas Matematika dan Ilmu Pengetahuan Alam"},{"name":"Department of Surgery Bandar Lampung Lampung Universitas Lampung Fakultas Kedokteran"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sutyarso","family":"Sutyarso","sequence":"additional","affiliation":[{"name":"Department of Biology Bandar Lampung Lampung Universitas Lampung Fakultas Matematika dan Ilmu Pengetahuan Alam"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hendri","family":"Busman","sequence":"additional","affiliation":[{"name":"Department of Biology Bandar Lampung Lampung Universitas Lampung Fakultas Matematika dan Ilmu Pengetahuan Alam"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7646-0869","authenticated-orcid":false,"given":"Syazili","family":"Mustofa","sequence":"additional","affiliation":[{"name":"Department of Biochemistry and Biomolecular Bandar Lampung Lampung Universitas Lampung Fakultas Kedokteran"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Wawan","family":"Abdullah Setiawan","sequence":"additional","affiliation":[{"name":"Department of Biology Bandar Lampung Lampung Universitas Lampung Fakultas Matematika dan Ilmu Pengetahuan Alam"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ratna","family":"Dewi Puspita Sari","sequence":"additional","affiliation":[{"name":"Department of Obstetric and Gynecology Bandar Lampung Lampung Universitas Lampung Fakultas Kedokteran"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nuning","family":"Nurcahyani","sequence":"additional","affiliation":[{"name":"Department of Biology Bandar Lampung Lampung Universitas Lampung Fakultas Matematika dan Ilmu Pengetahuan Alam"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"2560","published-online":{"date-parts":[[2026,4,29]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"1364-1379","DOI":"10.2337\/dc12-0413","article-title":"Management of Hyperglycemia in Type 2 Diabetes: A Patient-Centered Approach: Position Statement of the American Diabetes Association (ADA) and the European Association for the Study of Diabetes (EASD).","volume":"35","author":"S Inzucchi","year":"2012","journal-title":"Diabetes Care."},{"key":"ref3","doi-asserted-by":"publisher","first-page":"1143067","DOI":"10.3389\/fendo.2023.1143067","article-title":"Microvascular and macrovascular complications of type 2 diabetes mellitus: Exome wide association analyses.","volume":"14","author":"A Mansour","year":"2023","journal-title":"Front. 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Comment: <\/b>The manuscript states that data are non-parametric, yet both Kruskal-Wallis test and ANOVA are reported. This represents a fundamental methodological contradiction. (ANOVA + Tukey post hoc (if parametric), OR Kruskal\u2013Wallis + Dunn\u2019s test (if non-parametric)) \n<b>Response:<\/b> We sincerely thank the reviewer for identifying this inconsistency. We agree that the original statistical description contained a methodological contradiction. Although the manuscript incorrectly stated that one-way ANOVA with LSD post hoc testing was used, the analyses actually performed in this study were Kruskal-Wallis tests followed by Dunn\u2019s multiple-comparison post hoc tests, consistent with the non-parametric nature of the data. This error has now been corrected in the revised manuscript, and the statistical methods have been rewritten to accurately reflect the analyses conducted. We have also carefully reviewed the Results section and tables to ensure consistency with the corrected statistical approach.  \n<b>&nbsp;2. Comment: <\/b>Several results are described as \u201cincreased\u201d or \u201cimproved\u201d despite p &gt; 0.05. E.g. sperm parameters (p = 0.877) are interpreted as improvements. \n<b>Response:<\/b> We sincerely thank the reviewer for this careful and important comment. We agree that it is not appropriate to interpret non-significant differences as definite improvements. In response, we have revised the Results and Discussion sections to ensure that only statistically significant findings are described as significant improvements, whereas outcomes with p &gt; 0.05 are now reported more cautiously as numerical differences without statistical significance. For example, the statements regarding sperm parameters have been corrected accordingly. We have also rechecked the manuscript throughout to ensure consistency between the statistical results and their interpretation.  \n<b>&nbsp;3. Comment: <\/b>The term \u201cgland penis\u201d is incorrect. Should be &quot;glans penis&quot;. \n<b>Response:<\/b> We sincerely thank the reviewer for this valuable comment. We agree that the correct anatomical term is \u201cglans penis\u201d rather than \u201cgland penis.\u201d The manuscript has been revised accordingly in the Materials and Methods section to ensure anatomical accuracy.  \n<b>&nbsp;4. Comment: <\/b>The term &quot;male kisses&quot; is non-scientific. Rephrase appropriately. \n<b>&nbsp;Response:<\/b> We sincerely thank the reviewer for this valuable comment. We agree that the expression \u201cmale kisses\u201d is not appropriate for scientific reporting of animal sexual behavior. In the revised manuscript, we have replaced this phrase with more formal and objective terminology to describe the observed behavior in the Libido Assessment subsection. We changed the sentence into \u201cCourtship latency was defined as the time from opening of the partition until the male initiated investigatory or courtship behavior toward the female, including anogenital sniffing or contact with the female\u2019s body.\u201d  \n<b>&nbsp;5. Comment: <\/b>Include units for sperm count and fluid intake. \n<b>Response: <\/b>We sincerely thank the reviewer for this valuable observation. We agree that the omission of units for sperm count and fluid intake reduced the clarity of data presentation. In response, we have revised the manuscript into units of millions\/mL to include the appropriate units for these variables throughout the text and tables.  \n<b>6. Comment:<\/b> Avoid overgeneralization. \n<b>Response<\/b>: We sincerely thank the reviewer for this valuable observation. We agree that the original manuscript contained several statements that were overly broad in relation to the data presented. In response, we have revised the manuscript throughout to ensure that the conclusions are stated more cautiously and are fully supported by the results. Specifically, we avoided definitive claims, reduced overstatement of non-significant findings, and clarified that the observed effects should be interpreted as dose-dependent, endpoint-specific, and preliminary findings in a short-term alloxan-induced diabetic rat model.  \n<b>7. Comment:<\/b> There is contradictory literature use, i.e. &quot;piperine increases testosterone&quot;, and &quot;piperine inhibits spermatogenesis&quot;. These two statements contradict each other. \n<b>Response<\/b>: We sincerely thank the reviewer for this valuable comment. We agree that the earlier wording may have oversimplified the published evidence. In the revised manuscript, we clarified that the available literature suggests complex and context-dependent effects of piperine, rather than uniformly beneficial or uniformly harmful reproductive effects. We now explicitly state that piperine may enhance Leydig-cell function and steroidogenic activity while, under different experimental conditions, adversely affecting spermatogenesis. We therefore revised the Discussion to reflect that these findings represent different reproductive endpoints and may vary according to dose, exposure duration, and study conditions.  \n<b>8. Comment<\/b>: &quot;ddp-4&quot; should be written as &quot;DDP-4&quot; \n<b>Response:<\/b> We sincerely thank the reviewer for this careful observation. We agree that the abbreviation should be written consistently in uppercase. In the revised manuscript, \u201cdpp-4\u201d has been corrected to \u201cDPP-4\u201d throughout the text.  \n<b>9. Comment<\/b>: Establish the relevance of the oxidative stress pathway and nitric oxide signalling in your discussion. \n<b>Response: <\/b>We sincerely thank the reviewer for this important comment. In response, we further revised the Discussion to more explicitly establish the relevance of the oxidative stress pathway and nitric oxide signaling to the findings of the present study. Specifically, we clarified that hyperglycemia-induced oxidative stress can reduce nitric oxide bioavailability and impair eNOS-mediated endothelial signaling in the corpus cavernosum, which contributes to diabetic erectile dysfunction. We also strengthened the discussion linking the directional increase in eNOS mRNA expression after black pepper treatment with the possibility of partial restoration of endothelial NO signaling. In addition, we added relevant recent literature to support the mechanistic relationship among oxidative stress, endothelial dysfunction, and impaired erectile function in diabetes. \n<b>&nbsp; 10. Comment: <\/b>Conclusion is overstated. Conclude using the specific parameters evaluated. Correct typo such as &quot;randmoized&quot; to &quot;randomized&quot; \n<b>Response:<\/b> We sincerely thank the reviewer for this valuable observation. We agree that the previous Conclusion was somewhat overstated. In the revised manuscript, we have rewritten the Conclusion to refer specifically to the measured outcomes, including total penile reflexes, libido-related parameters, intratesticular testosterone, sperm concentration, sperm morphology, Leydig cell count, and eNOS expression, rather than making broad claims about diabetic male reproductive dysfunction overall. We have also revised the language to ensure that the findings are interpreted as dose-dependent, endpoint-specific, and preliminary. However, we don\u2019t used the term \u201crandomized\u201d in our manuscript anymore.  \n<b>11. Comment:<\/b> The phrase &quot;known to be the cause of.....&quot; used in the second paragraph of the discussion section should be rephrased as &quot;....is associated with..... &quot; \n<b>Response:<\/b> We sincerely thank the reviewer for this valuable comment. We agree that the original phrasing implied a stronger causal interpretation than was warranted. In the revised Discussion, we have replaced \u201cknown to be the cause of\u201d with newer sentences in our revised manuscript.  \n<b>12. Comment: <\/b>The discussion attributes effects to increased testosterone and antioxidant activity. However, no biochemical assays (e.g., testosterone, SOD, catalase, MDA) were performed. Remove or clearly qualify speculative statements and limit discussion to observed data unless supported by references. \n<b>Response: <\/b>We sincerely thank the reviewer for this valuable observation. We agree that the original Discussion contained mechanistic statements that could be interpreted as stronger than warranted by the available data. In the revised manuscript, we have clarified that intratesticular testosterone was directly measured and may therefore be discussed as an observed endpoint, whereas oxidative stress-related mechanisms were not directly evaluated, as no assays such as SOD, catalase, or MDA were performed. We therefore revised the Discussion to remove over-speculative language, qualify mechanistic interpretations more carefully, and restrict causal inferences to the measured outcomes. Where oxidative stress or antioxidant effects are mentioned, these are now presented explicitly as literature-supported hypotheses rather than direct findings of the present study. We have also included this into our limitation.  \n<b>13. Comment:<\/b> Extremely large standard deviations (e.g., sperm count: &plusmn;52.9), which may indicate poor experimental control or insufficient sample size. \n<b>Response:<\/b> We sincerely thank the reviewer for this valuable comment. We acknowledge that some outcomes, particularly sperm count, showed relatively large standard deviations. We carefully re-examined the raw data and confirmed that these values were accurate and not due to transcription error. We elected to retain the original values to preserve transparency and to avoid underreporting biological variability. We agree that the relatively small group size may have contributed to the wide dispersion observed. Accordingly, we used non-parametric statistical analysis (Kruskal-Wallis followed by Dunn\u2019s post hoc test) and revised the text to ensure that the findings are interpreted cautiously. However, we also include this topic into our limitation to keep our study transparent.  \n<b>14. Comment:<\/b> The title was duplicated in reference 3. The author's name(s) are not included in reference 20, and two journal names were mentioned within the same reference. Some references have the journal names abbreviated, while some do not (be consistent). Is the work clearly and accurately presented and does it cite the current literature? \n<b>Response:<\/b> We sincerely thank the reviewer for this important comment. We have carefully re-examined the entire reference list and corrected the identified errors, including the duplicated title in Reference 3, the incomplete author and journal information in Reference 20, and inconsistencies in journal-name formatting. The references have now been standardized throughout the manuscript to ensure consistency and accuracy. Furthermore, we revised the Discussion extensively to strengthen the scientific presentation of the work and to ensure that the findings are interpreted in light of relevant and current literature.","URL":"https:\/\/f1000research.com\/articles\/15-38\/v2#referee-comment-15952","order":3,"name":"referee-comment-15952","label":"Referee Comment","group":{"name":"article-reports","label":"Article Reports"}},{"value":"The author(s) declared that no grants were involved in supporting this work","order":4,"name":"grant-information","label":"Grant Information"},{"value":"This is an open access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.","order":0,"name":"copyright-info","label":"Copyright"}]}}