中华大蟾蜍(Bufo bufo gargarizans Cantor)，为我国名贵中药材蟾酥、蟾衣和蟾皮的重要基原动物，隶属于脊索动物门蟾蜍科蟾蜍属，在我国各省均有分布。蟾酥现已被《中华人民共和国药典》（2020年版，一部）收录，但因产量有限，其价格近年持续走高。目前，野生中华大蟾蜍由于前期人为滥捕和栖息地环境退化，资源量急剧下降；同时中华大蟾蜍由于被“三有动物”保护名录收录而不得随意捕获；故开展中华大蟾蜍人工养殖是缓解蟾酥供需矛盾的关键途径，而在中华大蟾蜍人工养殖技术研究中多聚焦于探究幼蟾的养殖技术，关于蝌蚪养殖技术探究及人为环境调控与蝌蚪生长质量的相关性鲜见报道。

本研究在课题组前期研究工作的基础上，以人工养殖条件下的中华大蟾蜍蝌蚪作为试验对象，首先依据戈斯纳（Gosner）分期表和养殖实际情况，确定了适用于观测辽宁本溪中华大蟾蜍蝌蚪生长发育的关键时期，明确其生物学特性，其次通过养殖水温和pH的单因素多水平调控试验，筛选出辽宁本溪地区中华大蟾蜍蝌蚪人工养殖适宜水温和酸碱度范围，以期通过生长关键环境参数的人为调控而提高蝌蚪生长质量，促进中华大蟾蜍人工养殖技术的规范。主要研究结果如下：

（1）通过对辽宁本溪地区人工养殖条件下中华大蟾蜍卵和蝌蚪的生物学特性观测，发现人工养殖和野生条件下受精卵和蝌蚪的生长发育节律和形态存在差异。结果表明：受精卵，卵带孵化时间在人工养殖条件下相对短，孵化率和蝌蚪整齐度人工养殖也优于野生条件。蝌蚪，人工养殖条件下的蝌蚪生长速度较快，体型较大，变态时间提前。养殖与野生的蝌蚪在摄食、活动和变态行为等三个方面的生活习性上存在显著区别。

（2）以戈斯纳（Gosner）分期为基础，确定了辽宁本溪地区蝌蚪养殖的5个关键时期，分别为：胚胎发育期、后肢生长期、五趾分化期、变态发育期和变态期。在各个时期可根据其生长特点，有针对性的优化管理条件以提高养殖管理的便捷性和准确性。

（3）通过单因素多水平试验，探究了不同水温及酸碱度与中华大蟾蜍蝌蚪生长发育的相关性。水温：在15-30℃范围内，水温与蝌蚪的多个生长指标呈显著负相关，随着水温升高，蝌蚪生长周期缩短，体增重减少，存活率下降；在15-19℃范围内水温能显著促进蝌蚪体重的生长并提高存活率，为蝌蚪适宜的养殖水温。酸碱度：在pH 6.5-8.5动态变化的酸碱环境更有利于蝌蚪生长，能有效促进蝌蚪生长并降低养殖风险。而养殖水体恒定的酸碱度不利于蝌蚪发育，低于pH 5.0或高于pH 10.0的极端的酸碱环境甚至导致大规模死亡。

本研究结论如下：在辽宁本溪人工养殖条件下，中华大蟾蜍受精卵孵化时间缩短，孵化率及蝌蚪整齐度优于野生环境；蝌蚪生长更快、体型更大且变态提前，其摄食、活动与变态行为显著区别于野生种群。基于戈斯纳分期确立胚胎发育期、后肢生长期、五趾分化期、变态发育期和变态期5个关键阶段，为精细化养殖管理提供依据。环境调控试验表明：15-19℃为蝌蚪适宜水温，动态pH 6.5-8.5环境最利于生长，而极端酸碱度（pH<5.0或>10.0）可致大规模死亡，为药用蟾蜍高效养殖提供关键技术参数。

Bufo bufo gargarizans Cantor, a chordate species belonging to the family Bufonidae, is an important source of traditional Chinese medicinal materials such as Bufonis venenum, toad clothing, and toad skin. With a wide distribution across China, it has been included in the Chinese Pharmacopoeia (2020 Edition, Volume I). However, the limited yield of Bufonis venenum has led to a continuous increase in its market price in recent years. Due to overharvesting and habitat degradation, wild populations of B. gargarizans have sharply declined, prompting its inclusion in the “List of Terrestrial Wildlife with Ecological, Scientific, and Social Value” by the National Forestry and Grassland Administration to restrict unauthorized capture. Therefore, the development of artificial breeding techniques for B. gargarizans has become crucial for alleviating the supply-demand imbalance of Bufonis venenum. Current research on captive breeding predominantly focuses on juvenile toad cultivation techniques, while systematic investigations into tadpole rearing methodologies and the correlation between artificial environmental regulation and tadpole growth quality remain notably underexplored in existing literature.

Building on prior research, this study investigated tadpoles of B. gargarizans under controlled breeding conditions. Key developmental stages were identified using the Gosner staging system and practical breeding experience, particularly focusing on biological characteristics specific to populations in Benxi, Liaoning Province. Single-factor multi-level experiments were conducted to optimize water temperature and pH ranges for tadpole rearing. The goal was to enhance growth quality through environmental parameter regulation and advance standardized breeding protocols. Major findings include:

(1) Comparative analysis revealed distinct biological differences between captive-bred and wild tadpoles in Benxi, Liaoning Province. Under captive conditions, the embryonic incubation periods were shorter, hatching rates were higher, and tadpole uniformity was greater. Captive tadpoles exhibited accelerated growth, larger body size, earlier metamorphosis, and significant behavioral differences in feeding, activity, and metamorphic patterns.

(2) In accordance with Gosner staging, five critical developmental phases were identified: embryonic development, hindlimb growth, five-toe differentiation, metamorphic development, and metamorphosis. To improve breeding efficiency, targeted management strategies were proposed for each phase.

(3) The experiment explored the effects of different water temperatures and pH levels on the growth and development of B. gargarizans tadpoles. Results showed that water temperature had a significant negative correlation with multiple growth indicators of tadpoles in the range of 15 - 30℃. As water temperature increased, the tadpoles' growth cycle shortened, weight gain decreased, and survival rate declined. The optimal water temperature range for tadpole farming was found to be 15 - 19℃, where water temperature significantly promoted weight gain and improved survival rates.

Regarding pH levels, a dynamic pH range of 6.5 - 8.5 was most conducive to tadpole growth, effectively promoting their development and reducing farming risks. Maintaining constant pH levels in aquaculture water proves detrimental to tadpole development, with extreme pH environments (below pH 5 or exceeding pH 10) inducing mass mortality events.

Under artificial rearing in Benxi, Liaoning, B. gargarizans exhibits accelerated embryonic development with enhanced hatching rates and larval synchrony, alongside tadpoles demonstrating superior growth metrics (larger size, earlier metamorphosis) and distinct behavioral phenotypes compared to wild counterparts. Application of Gosner staging delineated five critical phases-embryogenesis, hindlimb budding, digital differentiation, metamorphic progression, and climax-enabling precision husbandry. Environmental optimization identified 15-19°C and dynamic pH 6.5-8.5 as growth-optimal regimes, while extreme pH (< 5.0 or > 10.0) induced catastrophic mortality, establishing key technical parameters for medicinal toad aquaculture.

[1] 陈继军, 张旋, 杨绍军, 等. 贵州雷公山自然保护区两栖动物调查报告[J]. 四川动物, 2007, (04): 826-830.

[2] 陈强. 棘胸蛙蝌蚪的生理生态研究及全年候生长养殖技术探讨[D]. 福州大学, 2017.

[3] 陈强威, 唐春萍, 王秋新, 等. 蟾酥注射液对脂多糖发热模型大鼠解热及体外抑菌作用的研究[J]. 中药药理与临床, 2018, 34(02): 58-62.

[4] 陈瀛澜, 郝艳艳, 郭夫江, 等. 蟾酥化学成分及药理活性研究进展[J]. 中草药, 2017, 48(12): 2579-2588.

[5] 程亚美, 赵金良, 宋凌元, 等. 盐度、碱度对尼罗罗非鱼生长和肌肉品质的影响[J]. 水产科学, 2020, 39(03): 341-349.

[6] 崔岩. 中国林蛙蝌蚪人工养殖技术及管理研究[D]. 黑龙江: 东北林业大学, 2005.

[7] 戴劭. 棘胸蛙蝌蚪配合饲料中蛋白源优化的研究[D]. 集美大学, 2024.

[8] 邓莎, 李梦园, 武小玲, 等. 蟾酥化学成分及抗肿瘤作用研究进展[J]. 中成药, 2022, 44(03): 884-890.

[9] 佟庆. 东北林蛙肠道菌群多样性及其影响因素研究[D]. 东北农业大学, 2019.

[10] 杜伟, 郑重莺, 马文君, 等. 浙江省重点水产养殖区水体pH值和浮游植物季相变化[J]. 浙江农业科学, 2017, 58(06): 1048-1050.

[11] 费梁, 胡淑琴, 叶昌媛, 等. 中国动物志·两栖纲[M]. 北京: 科学出版社, 2009.

[12] 费梁, 叶昌嫒, 江建平. 中国两栖动物彩色图鉴[Ｍ]. 成都: 四川科学技术出版社, 2010.

[13] 冯秀娟, 陈大健, 李忠生, 等. 蟾蜍的化学成分及其临床应用[J]. 中兽医学杂志, 2007, (03): 36-38.

[14] 高波, 中华大蟾蜍标准化养殖研究[Z]. 安徽省, 安徽华润金蟾药业股份有限公司, 2019.

[15] 高延亮. 盐碱池塘高pH水体物理调控方法研究[D]. 上海海洋大学, 2018.

[16] 高子阳, 姜鹏, 詹常森. 中华大蟾蜍幼蟾的摄食节律研究[J]. 特种经济动植物, 2022, 25(02): 65-68.

[17] 龚双姣, 马陶武. 中华大蟾蜍蝌蚪的摄食节律和日摄食率[J]. 吉首大学学报(自然科学版), 2005, (03): 69-71.

[18] 龚钰雯, 黄春红, 覃日锐, 等. 低pH值养殖水对罗非鱼肠道菌群结构的影响[J]. 黑龙江畜牧兽医, 2024, (08): 117-125.

[19] 苟新元, 洪英科. 中药蟾酥在人工流产手术中麻醉作用的研究[J]. 中国计划生育学杂志, 2003 (01): 55-56.

[20] 国家林业和草原局公布新调整的“三有”野生动物名录[J]. 浙江林业, 2023, (07): 2+1.

[21] 国家药典委员会. 中华人民共和国药典. 一部[M]. 北京: 中国医药科技出版, 2020.

[22] 韩庆, 龚丽军, 凌卯梅, 等. pH对蒙古鲌肝胰脏和肠道中消化酶活性的影响[J]. 水产学杂志, 2020, 33(06): 39-43.

[23] 韩婷, 李艳芳, 刘金旗, 等. 中华大蟾皮化学成分研究[J]. 现代中药研究与实践, 2019, 33(05): 16-18.

[24] 郝爽. 蟾酥活性成分对神经元兴奋性的影响[D]. 大连理工大学, 2012.

[25] 胡红珍, 刘萍, 曾海涛. 蟾酥水提取物对人宫颈癌HeLa细胞生物学活性的抑制作用[J]. 新医学, 2013, 44(05): 351-355.

[26] 胡梦如. 雌二醇、温度、猪胎衣对中国林蛙性别及蝌蚪生长发育的影响[D]. 沈阳农业大学, 2016.

[27] 黄东宇, 杨璐铭, 钟映琪, 等. 蟾酥活性成分及其药理作用的研究进展[J]. 沈阳药科大学学报, 2023, 40(01): 124-136.

[28] 姬诚, 谢媛媛, 王义明, 等. 国产蟾酥药材质量综合评价研究[J]. 中国药学杂志, 2011, 46(19): 1466-1470.

[29] 姜鹏, 高子阳, 詹常森. 基于蟾酥品质评价的我国中华大蟾蜍资源分布特征[J]. 中国中药杂志, 2022, 47(11): 2924-2931.

[30] 解景田, DEY Lucy, 吴吉安, 等. 脂布福吉宁的心脏毒性作用:电生理学证据(英文)[J]. Acta Pharmacologica Sinica, 2001, (04): 3-11.

[31] 寇冠军, 秦姿凡, 邓雅芳, 等. 蟾酥的研究进展[J]. 中草药, 2014, 45(21): 3185-3189.

[32] 赖慧, 赖洪林, 陈立, 等. 蟾酥注射液对人肝癌smmc-7721细胞增殖与凋亡的影响[J]. 医学研究杂志, 2021, 50(04): 69-73.

[33] 李红梅, 聂金芬. 抚仙湖周边两栖动物栖息环境现状调查[J]. 畜牧与饲料科学, 2009, 30(10): 94-95.

[34] 李军德, 赵群, 康彦, 等. 密度与饲料对中华蟾蜍蝌蚪生长发育的影响[J]. 中国中药杂志, 2016, 41(06): 1001-1007.

[35] 李顺才. 中华大蟾蜍养殖与开发利用[M]. 化学工业出版社, 2014.

[36] 厉旭云, 陆源, 单绮娴, 等. 华蟾素对大鼠胸主动脉的缩血管作用及其机制[J]. 浙江大学学报(医学版), 2006, (02): 178-181.

[37] 立志, 李晓晨. 温度对中国林蛙卵孵化和孵出热耐受性的影响[J]. 动物学杂志, 2007, (01): 121-127.

[38] 梁晓萍, 张政, 胡坪, 等. 蟾酥急性毒性的代谢组学研究[J]. 高等学校化学学报, 2011, 32(01): 38-43.

[39] 梁智佳. 中华大蟾蜍(Bufo gargarizans)和中国林蛙(Rana chensinensis)尾吸收分子机制探究[D]. 陕西师范大学, 2021.

[40] 刘娟, 童明慧, 郭英球, 等. 蟾酥抗肿瘤临床应用及其疗效机制的研究进展[J]. 天然产物研究与开发, 2022, 34(08): 1430-1438.

[41] 刘乐, 蔡敏, 陈非洲, 等. 模拟酸雨对不同营养水平湖泊pH值的影响[J]. 生态与农村环境学报, 2018, 34(10): 917-923.

[42] 刘莉, 李成, 李乃兵, 等. 不同水温条件下中华蟾蜍蝌蚪的表型可塑性研究[J]. 四川动物, 2006, (02): 214-217.

[43] 吕敬才, 张倩, 王宇俊, 等. 三都县拉揽乡两栖爬行动物资源研究[J]. 贵阳学院学报(自然科学版), 2013, 8(01): 10-19.

[44] 陆文娟. 蟾酥的心脏毒性评价和减毒药物筛选[D]. 南京中医药大学, 2012.

[45] 毛铭廷, 苏秀俭, 张紫菊. 中华大蟾蜍(Bufo bufo gargaraizns)冬眠期的人工催产和人工授精及其早期发育的观察[J]. 甘肃师范大学学报(自然科学版), 1964, (01): 26-31.

[46] 苗春云, 刘毅, 刘衍忠, 等. 西玛津对中华大蟾蜍生存和性腺发育的影响[J]. 环境与健康杂志, 2011, 28(12): 1075-1077.

[47] 明仁月, 陈可嘉, 李兰英, 等. 农药环境污染对两栖动物蛙类毒性效应研究进展[J]. 现代农药, 2023, 22(06): 22-32.

[48] 聂捷夫, 罗蓉, 辜永河. 贵阳地区中华大蟾蜍越冬习性观察[J]. 动物学杂志, 1987, (01): 26-29.

[49] 乔莉, 段文娟, 姚遥, 等. 蟾酥中强心甾类化学成分的分离与鉴定[J]. 沈阳药科大学学报, 2007, (10): 611-614.

[50] 秦姿凡, 王保和. 中药蟾酥的研究进展概况[J]. 中国药物评价, 2014, 31(05): 306-309.

[51] 邵玲, 任江华, 曹茂银. 低温及蟾酥对麻醉家兔心肌缺血再灌注损伤的保护作用研究[J]. 中西医结合心脑血管病杂志, 2005, (02): 134-136.

[52] 邵然. 除草剂丁草胺对中华大蟾蜍脑及肝脏的毒性研究[D]. 沈阳师范大学, 2011.

[53] 宋涛. 蟾酥中有效成分蟾毒灵的含量测定及其对心肌细胞钙信号的影响[D]. 河北医科大学, 2018.

[54] 苏发文. 大丰盐碱养殖虾塘浮游植物群落结构组成及其对水环境pH值的影响[D]. 上海海洋大学,2016.

[55] 苏娟. 湖南八大公山中华大蟾蜍冬眠习性及栖息地选择[D]. 华中师范大学, 2017.

[56] 孙崇峰, 范圣此, 罗毅, 等. 蟾酥化学成分及其人工合成的研究进展[J]. 中草药, 2018, 49(13): 3183-3192.

[57] 陶志英, 马保新, 余智杰, 等. 环境因子对棘胸蛙蝌蚪生长发育的影响[J]. 湖南农业科学, 2015, (02): 55-56.

[58] 田园, 杨沛刚, 檀碧波, 等. 蟾酥注射液联合阿帕替尼治疗晚期胃癌患者的疗效[J]. 实用医学杂志, 2020, 36(18): 2583-2586.

[59] 王安纲. 中华大蟾蜍的生物学特性及养殖技术[J]. 内陆水产, 2004, (04): 18-19.

[60] 王传海, 李宽意, 文明章, 等. 苦草对水中环境因子影响的日变化特征[J]. 农业环境科学学报, 2007, (02): 798-800.

[61] 王恒杰, 戴梦杨, 王倩, 等. 环境胁迫因子对鱼类健康影响的研究进展[J]. 中国农学通报, 2025, 41(02): 157-164.

[62] 王立志. 恒温和变温驯化对大蟾蜍蝌蚪热耐受性的影响[J]. 生态学报, 2014, 34(04): 1030-1034.

[63] 王倩. 中华蟾蜍蝌蚪发育对温度的响应[D]. 华中师范大学, 2019.

[64] 王信海, 姜爱兰, 蔺玉华, 等. 不同盐度和碱度对卡拉白鱼精子活力和受精率的影响[J]. 江西农业学报, 2020, 32(05): 93-98.

[65] 王守红, 李豪, 刘露莎, 等. 温度对饰纹姬蛙蝌蚪生长的影响[J]. 动物学杂志, 2018, 53(02): 191-197.

[66] 王文霞. 中华蟾蜍四个种群蝌蚪发育的地理差异及其进化机制[D]. 华中师范大学, 2017.

[67] 王晓磊, 余道军, 侯晓丽. 中药蟾酥抗多重耐药鲍曼不动杆菌体外实验研究[J]. 中华中医药学刊, 2016, 34(11): 2801-2804.

[68] 危晓莉, 汪晓莺, 汤伟. 华蟾素对正常人外周血来源树突状细胞的影响[J]. 南通大学学报(医学版), 2007, (02): 98-100.

[69] 吴旻, 付议颉, 常雷, 等. 温度对东北林蛙蝌蚪生长发育和性别比例的影响[J]. 特种经济动植物, 2023, 26(2):47-49.

[70] 辛少鲲, 司南, 王宏洁, 等. 蟾皮中亲水性成分的化学研究[J]. 中国中药杂志, 2016, 41(20): 3767-3772.

[71] 辛秀兰, 张宝璟, 苏东海, 等. 中药蟾酥的药理作用研究进展[J]. 现代生物医学进展, 2012, 12(03): 588-590.

[72] 徐迪辉. 基于质谱技术对蟾酥中吲哚类生物碱的鉴定及镇痛抗炎活性评价[D]. 南京中医药大学, 2021.

[73] 许海鑫, 朱洪赓, 张家敏, 等. 碳酸盐碱度对大鳞副泥鳅胚胎发育和仔鱼活力的影响[J]. 南方水产科学, 2024, 20(02): 56-62.

[74] 徐骁骁, 赵文阁, 刘鹏. 环境温度对东北林蛙不同地理种群繁殖期体温和胚胎发育的影响[J]. 生态学报, 2018, 38(08): 2965-2973.

[75] 许友卿, 张仁珍, 丁兆坤. pH对鱼类繁育及生长发育的影响[J]. 水产科学, 2014, 33(02): 133-136.

[76] 徐瑱璐. 蟾酥活性成分酯蟾毒配基抑制lps诱导巨噬细胞炎症的作用及机制研究[D]. 北京协和医学院, 2023.

[77] 杨爱文, 范雪梅, 李雪, 等. 基因芯片研究蟾酥急性毒性及配伍减毒机制[J]. 高等学校化学学报, 2011, 32(05): 1058-1064.

[78] 杨程. pH对棘胸蛙蝌蚪若干生理指标的影响效应[D]. 浙江海洋大学, 2017.

[79] 殷佩浩. 中药蟾酥的研究进展[J]. 上海医药, 2015, 36(10): 3-6.

[80] 于华芸. 蟾酥的古今应用概述[J]. 辽宁中医药大学学报, 2008, (08): 53-54.

[81] 余姣姣. 四川地区中华蟾蜍种群遗传多样性及高原适应研究[D]. 四川农业大学, 2018.

[82] 于业辉, 张守纯, 刘超. 中华大蟾蜍的胚胎发育观察[J]. 沈阳农业大学学报, 2009, 40(06): 723-727.

[83] 詹常森. 中华大蟾蜍养殖模式与技术[M]. 上海交通大学出版社, 2023.

[84] 张盛周, 朱联九, 钱玉, 等. pH和温度对黑斑蛙消化道脂肪酶活力的影响[J]. 安徽师范大学学报(自然科学版), 2008, (02): 159-162.

[85] 张薇, 刘玉兰, 徐从云, 等. 蟾酥的镇痛活性成分[J]. 沈阳药科大学学报, 1998, (04): 37-40.

[86] 张小琴, 丁洪霞. 光照度对棘腹蛙蝌蚪行为的影响[J]. 湖北农业科学, 2021, 60(11): 99-101.

[87] 张晓冬. 黑龙江药用蛙类资源介绍[J]. 中国中医药科技, 2004, (06): 397.

[88] 张笑春, 蔡文栅, 俞乐, 等. 温度对黑斑蛙蝌蚪的生长发育和适应性影响[J]. 现代畜牧科技, 2024, (03): 52-54.

[89] 张秀高. 蟾蜍的人工饲养[J]. 中草药, 2000, (11): 66-67.

[90] 张志强, 彭秀娟, 裴鑫怡, 等. 增温影响中华蟾蜍蝌蚪表型可塑性的生理代价[J]. 生态学报, 2024, 44(08): 3329-3336.

[91] 赵进哲, 姜治国, 包卓旋, 等. 动物在热应激和冷应激条件下生理与代谢变化的比较分析[J]. 动物营养学报, 2025, 37(01): 169-181.

[92] 赵联芳, 朱伟, 莫妙兴. 沉水植物对水体pH值的影响及其脱氮作用[J]. 水资源保护, 2008, (06): 64-67.

[93] 赵威. 蟾酥清热解毒作用研究进展[J]. 临床军医杂志, 2016, 44(01): 105-107.

[94] 周波, 周霞. 远华蟾毒精通过调控mmp-2､mmp-9合成的相关信号传导通路抑制乳腺癌细胞增殖的作用与机制探讨[J]. 中国社区医师, 2020, 36(05): 4-6.

[95] 周可欣, 文鑫, 邓成, 等. 豹纹鳃棘鲈体色变异的色素细胞差异分析[J]. 水生生物学报, 2021, 45(05): 1164-1170.

[96] 周晓艳. 复方蟾酥散外敷治疗中度癌痛的临床研究[D]. 湖南中医药大学, 2007.

[97] 朱成悦, 张姗姗, 杨飞, 等. 基于斑马鱼模型的蟾酥甲醇提取物抑制血管生成和抗炎活性研究[J]. 药物评价研究, 2021, 44(07): 1392-1397.

[98] 朱海燕, 苏秀芳, 梁桂霞. 中华大蟾蜍受精过程的细胞学研究: 中国动物学会第十四届会员代表大会及中国动物学会65周年年会[C], 1999.

[99] 朱艳芳, 高波, 张爱民, 等. 温度和光照时间对中华大蟾蜍胚胎发育的影响[J]. 安徽农业科学, 2012, 40(30): 14748-14749.

[100] 郑振华, 董双林, 田相利. pH不同处理时间的周期性变动对凡纳滨对虾生长的影响[J]. 中国海洋大学学报(自然科学版), 2008, (01): 45-51.

[101] 邹李昶. 棘胸蛙蝌蚪对温度的耐受与响应特征[D]. 浙江海洋学院, 2015.

[102]左之良, 彭治桃, 田璐, 等. 不同介质､温度对中华鳖胚胎发育与幼鳖形态特征及运动机能的影响[J]. 云南农业大学学报(自然科学), 2024, 39(04): 38-44.

[103]Abbink W, Garcia B A, Roques A J, et al. The effect of temperature and pH on the growth and physiological response of juvenile yellowtail kingfish Seriola land in recirculating aquaculture sys-tems[J]. Aquaculture, 2012, 330: 130-135.

[104]Altwegg R, Reyer H U. Patterns of natural selection on size at metamorphosis in water frogs[J]. Evolution, 2003, 57(4): 872-882.

[105]Boaro L B, Lima D P E, Maria A D, et al. Cinobufagin: Unveiling the hidden bufadienolide's prom-ise in combating alimentary canal cancer development and progression-a comprehensive review[J]. Naunyn-Schmiedeberg's Archives of Pharmacology, 2025, (prepublish): 1-15.

[106]Dalpasso A, Seglie D, Eusebio Bergò P, et al. Effects of temperature and precipitation changes on shifts in breeding phenology of an endangered toad[J]. Scientific Reports, 2023, 13(1): 14573.

[107]Dastansara N, Vaissi S, Mosavi J, et al. Impacts of temperature on growth, development and surviv-al of larval Bufo (Pseudepidalea) viridis (Amphibia: Anura): implications of climate change[J]. Zo-ology and Ecology, 2017, 27(3-4): 228-234.

[108]Devi, M. L, Thabah, et al. Scanning Electron Microscopic Studies on the Effect of pH on the Gill Morphology of Polypedates teraiensis Hatchling (Amphibia: Anura)[J]. Journal of Advanced Mi-croscopy Research, 2014, 9(2): 115-120(6).

[109]Farquharson C, Wepener V, Smit J N. Acute and chronic effects of acidic pH on four subtropical frog species[J]. Water S.A., 2016, 42(1): 52-62.

[110]Freitas J S, Almeida E A. Antioxidant defense system of tadpoles (Eupemphix nattereri) exposed to changes in temperature and pH[J]. Zoological science, 2016, 33(2): 186-194.

[111]Gamidova D M, Rabadanova A I. Effect of pH on the Cytomorphological Parameters of Erythro-cytes of Rana macrocnemis Tadpoles[J]. Biology Bulletin, 2023, 50(5): 959-968.

[112]Gao J, Li X, Lu K, et al. Optimal dietary protein level for the growth and metamorphosis of bullfrog (Lithobates catesbeianus) tadpoles[J]. Aquaculture, 2024, 580: 740265.

[113]Helfman G S, Collette B B, Facey D E, et al. The diversity of fishes: biology, evolution, and ecolo-gy[M]. John Wiley & Sons, 2009.

[114]Hu Y L, Wu X B, Jiang Z G, et al. Population genetics and phylogeography of Bufo gargarizans in China[J]. Biochemical Genetics, 2007, 45: 697-711.

[115]Hubáček J, Gvoždík L. Terrestrial amphibians respond to rapidly changing temperatures with indi-vidual plasticity of exploratory behaviour[J]. Journal of Thermal Biology, 2024, 119: 103757.

[116]Kingsolver J G, Huey R B. Introduction: the evolution of morphology, performance, and fitness[J]. Integrative and Comparative Biology, 2003, 43(3): 361-366.

[117]Laurila A, Pakkasmaa S & Merila J. Influence of seasonal ti me constraints on growth and devel-opment of common frog tadpoles: a photoperiod experi ment[J]. Oikos, 2001, 95: 451-460.

[118]Lease H M, Hansen J A, Bergman H L, et al. Structural changes in gills of Lost River suckers ex-posed to elevated pH and ammonia concentrations[J]. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology, 2003, 134(4): 491-500.

[119]Li H, Meng Q, Chen T, et al. Transcriptomic response to low pH stress in gills of the pacific white shrimp, Litopenaeus vannamei[J]. Aquaculture Research, 2020, 51(1): 175-186.

[120]Lu H, Hu Y, Kang C, et al. Cadmium-induced toxicity to amphibian tadpoles might be exacerbated by alkaline not acidic pH level[J]. Ecotoxicology and Environmental Safety, 2021, 218: 112288.

[121]Nicholas S, Jelena M, Anssi L, et al. Short-term responses of Rana arvalis tadpoles to pH and pred-ator stress: adaptive divergence in behavioural and physiological plasticity[J]. Journal of compara-tive physiology. B, Biochemical, systemic, and environmental physiology, 2022, 192(5): 669-682.

[122]Pan Z, Luo Y, Xia Y, et al. Cinobufagin induces cell cycle arrest at the S phase and promotes apop-tosis in nasopharyngeal carcinoma cells[J]. Biomedicine & Pharmacotherapy, 2020, 122: 109763.

[123]Patil H S, Joshi S N. Effect of Water Temperature pH and Hardness on the Toxicity of Chromium Trioxide to Female Frog Rana cyanophlyctis[J]. Proceedings of Indian National Science Academy, 2015, 58(6B): 347.

[124]Pounds A, Masters K. Extinction in our times: Global amphibian decline[J]. Nature, 2009, 462(7269): 38-39.

[125]Reading C J. The effects of variation in climatic temperature (1980-2001) on breeding activity and tadpole stage duration in the common toad, Bufo bufo[J]. Science of The Total Environment, 2003, 310(1-3): 231-236.

[126]Saderne V, Wahl M. Effect of ocean acidification on growth, calcification and recruitment of calci-fying and non-calcifying epibionts of brown algae[J]. Biogeosciences Discussions, 2012, 9(3): 3739-3766.

[127]Sanuy D, Oromi N, Galofre A. Effects of temperature on embryonic and larval development and growth in the natterjack toad (Bufo calamita) in a semi-arid zone[J]. Animal Biodiversity and Con-servation, 2008, 31(1): 41-46.

[128]Shi X, Xiao K, Peng G, et al. Embryo development indices for the endangered Chinese sturgeon, Acipenser sinensis: the role of temperature on incubation time[J]. Environmental Biology of Fishes, 2024, 107(8): 899-907.

[129]Sun B, Si N, Wei X, et al. Multi-omics reveals bufadienolide Q-markers of Bufonis Venenum based on antitumor activity and cardiovascular toxicity in zebrafish[J]. Phytomedicine, 2024, 133, 155914.

[130]Tao B, Li X, Li X, et al. Derivatives of postbiotics (cell wall constituents) from Bacillus subtilis (LCBS1) relieve soybean meal-induced enteritis in bullfrog (Aquarana catesbeianus)[J]. Interna-tional Journal of Biological Macromolecules, 2024, 279: 135359.

[131]Tucker C S, D’Abramo L R. Managing high pH in freshwater ponds[M]. Stoneville: Southern Re-gional Aquaculture Center, 2008.

[132]Van Dongen J T, Gupta K J, Ramírez-Aguilar S J, et al. Regulation of respiration in plants: a role for alternative metabolic pathways[J]. Journal of Plant Physiology, 2011, 168(12): 1434-1443.

[133]Verma D K, Satyaveer N K M, Kumar P, et al. Important water quality parameters in aquaculture: An overview[J]. Agriculture and Environment, 2022, 3(3): 24-29.

[134]Wang J, Cai H, Liu Q L, et al. Cinobufacini inhibits colon cancer invasion and metastasis via sup-pressing Wnt/β-Catenin signaling pathway and EMT[J]. American Journal of Chinese Medicine, 2020, 48(3): 703-718.

[135]Wei W L, Hou J J, Wang X, et al. Venenum bufonis: An overview of its traditional use, natural product chemistry, pharmacology, pharmacokinetics and toxicology[J]. Journal of Ethnopharmacol-ogy, 2019, 237: 215-235.

[136]Wheeler C A, Bettaso J B, Ashton D T, et al. Effects of water temperature on breeding phenology, growth, and metamorphosis of foothill yellow‐legged frogs (Rana boylii): a case study of the reg-ulated mainstem and unregulated tributaries of California's Trinity River[J]. River Research and Ap-plications, 2015, 31(10): 1276-1286.

[137]Wilkie M P. Mechanisms of ammonia excretion across fish gills[J]. Comparative Biochemistry and Physiology Part A: Physiology, 1997, 118(1): 39-50.

[138]Yang F, Shao Q, Li J, et al. Adaptability of abnormal tadpole (Rana chensinensis) to water pH, sa-linity and alkalinity in Changbai Mountain of China[J]. The Journal of Applied Ecology, 2004, 15(8): 1411-1415.

[139]Zhan X, Wu H, Wu H, et al. Metabolites from Bufo gargarizans (Cantor, 1842): A review of tradi-tional uses, pharmacological activity, toxicity and quality control[J]. Journal of Ethnopharmacology, 2020, 246: 112178

[140]Zhu B, Zhong L, Shao C, et al. Effects of dietary lipid and protein levels on metamorphosis, growth, metabolism and gut microbiota of tadpole (Lithobates catesbeianus)[J]. Aquaculture, 2024, 587: 740900.