4-Mmethylbenzylidenecamphor（4-MBC；5-400 μM；48 小时）抑制 HTR8/SVneo 细胞的生长 [1]。 4-甲基亚苄基樟脑（10-50 μM；48 小时）可诱导人滋养层细胞增殖，并导致 SubG1 期细胞百分比增加 [1]。 4-在 48 小时内，50 μM 甲基苯亚甲基樟脑会降低人类滋养层细胞的营养 [1]。在人滋养层细胞中，樟脑（50 μM；5-120 分钟）刺激 PI3K/AKT 和 ERK1/2 的信号放大器 [1]。 4-甲基亚苄基樟脑在 20–50 μM 下作用 24 小时可显着提高 GPR56 和 SEMA6 A。

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- 对人滋养层细胞（HTR-8/SVneo细胞株）的抗增殖与促凋亡活性 [1]
- 抗增殖作用：4-甲基亚苄基樟脑（4-Methylbenzylidene camphor, 4-MBC）以剂量依赖方式抑制HTR-8/SVneo细胞增殖，48小时MTT实验IC50值为25.6 μM。浓度为20 μM、30 μM、40 μM时，增殖率较溶剂对照组（DMSO，<0.1%）分别降低38%、62%、85%。
- ROS介导的凋亡：20 μM、30 μM 4-MBC处理细胞24小时，可使细胞内活性氧（ROS）水平分别升高2.3倍、3.5倍（DCFH-DA染色）；凋亡率从对照组的3.2%升至20 μM组的18.5%、30 μM组的32.7%（Annexin V-FITC/PI双染）。Western blot检测显示，30 μM浓度下切割型caspase-3表达上调2.8倍、Bax表达上调2.1倍，Bcl-2表达下调0.4倍。
- 发育毒性相关体外活性 [2]
- 对人颗粒细胞：10 μM、20 μM 4-MBC分别使雌二醇生成量降低25%、40%，并通过qPCR和Western blot检测发现，芳香化酶（雌激素合成关键酶）表达分别下调30%、55%。
- 对小鼠胚胎成纤维细胞：50 μM 4-MBC使细胞迁移能力降低45%（伤口愈合实验），可能与胚胎发育受损相关。

4-甲基亚苄基樟脑（4-MBC；0.7、7、24、47 mg/kg/天；在交配前、怀孕期间和胎儿期间通过饮食给予父母）可引起基因表达的变化和南非新生儿的生长促进公式）[2]。

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- 动物模型中的发育毒性 [2]
- CD-1小鼠：妊娠小鼠从妊娠第6天（GD 6）至GD 15，口服给予4-MBC 100 mg/kg、300 mg/kg、500 mg/kg。300 mg/kg和500 mg/kg剂量使胎儿吸收率从对照组的5%分别升至18%、27%，胎儿体重分别降低12%、18%；500 mg/kg剂量还导致轻微胎儿畸形（指骨发育不全，发生率8%）。
- Sprague-Dawley大鼠：妊娠大鼠从GD 6至GD 20经皮暴露于4-MBC 50 mg/kg、100 mg/kg（模拟人类使用防晒霜场景）。100 mg/kg剂量使新生大鼠出生后第7天（PND 7）存活率降低15%，平均睁眼时间延迟1.5天。

细胞增殖测定 [1]
细胞类型： HTR8/SVneo 细胞
测试浓度： 0、5、10、20、50、100、200 和400 μM
孵育时间： 48 h
实验结果： 剂量依赖性抑制 HTR8/SVneo 细胞的细胞增殖。

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细胞凋亡分析 [1]
细胞类型： HTR8/SVneo 细胞
测试浓度： 10、20、50 μM
孵育时间： 48 h
实验结果： 早期和晚期凋亡细胞在 20 μM 和 50 μM 时显着增加。

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细胞周期分析 [1]
细胞类型： HTR8/SVneo 细胞
测试浓度： 5、10、20、50 μM
孵育时间： 48 h
实验结果： SubG1期细胞比例逐渐增加。细胞侵袭分析[1]
细胞类型： HTR8/SVneo 细胞
测试浓度： 50 μM
孵育时间： > 48 小时
实验结果： 侵袭性显着降低 81.5% 蛋白质印迹分析[1]
细胞类型： HTR8/ SVneo Cell
测试浓度： 50 μM
孵育时间：0、5、15、30、60、120 分钟
实验结果：AKT及其下游激酶蛋白P70S6K的磷酸化分别在5分钟和15分钟时达到峰值，然后在30分钟后减弱，

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- HTR-8/SVneo细胞增殖实验（MTT法）[1]
- 细胞以5×10³个/孔接种于96孔板，37°C（5% CO₂）孵育过夜。4-MBC溶于DMSO后加入孔中，终浓度为5 μM、10 μM、20 μM、30 μM、40 μM、50 μM（DMSO浓度<0.1%作为对照）。孵育24小时、48小时、72小时后，加入MTT试剂孵育4小时，DMSO溶解甲臜结晶，570 nm处测吸光度，计算增殖率和IC50。
- 凋亡检测（Annexin V-FITC/PI双染）[1]
- HTR-8/SVneo细胞（2×10⁵个/孔，6孔板）用4-MBC（20 μM、30 μM）或DMSO处理24小时。收集细胞，冷PBS洗涤，重悬于结合缓冲液，加入Annexin V-FITC和PI，室温避光孵育15分钟，流式细胞术分析凋亡率。
- ROS检测（DCFH-DA染色）[1]
- 细胞（1×10⁴个/孔，96孔黑色板）用4-MBC（20 μM、30 μM）处理12小时，加入DCFH-DA溶液（终浓度10 μM），37°C孵育30分钟。488 nm激发、525 nm发射波长下测定荧光强度，定量细胞内ROS水平。

Absorption, Distribution and Excretion
The maximum plasma concentration of enzacamene was 16ng/mL in healthy female volunteers following daily whole-body topical application of 2mg/cm^2 of sunscreen formulation at 10% (weight/weight) for four days. Blood concentration of enzacamene (4-MBC) and its main metabolite, 3-(4-carboxybenzylidene)camphor, peaked within 10 h after oral administration of enzacamene.
The urine concentration of 4 ng/mL and 4 ng/mL of enzacamene were observed in female and male volunteers, respectively. In a rat pharmacokinetic study, most of orally administered enzacamene was recovered in in feces as 3-(4-carboxybenzylidene)camphor and, to a smaller extent, as 3-(4-carboxybenzylidene)-6-hydroxycamphor. Glucuronides of both metabolites were also detectable in faces. In urine, one isomer of 3-(4-carboxybenzylidene)hydroxycamphor was the predominant metabolite [3-(4-carboxybenzylidene)-6-hydroxycamphor], the other isomers and 3-(4-carboxybenzylidene)camphor were only minor metabolites excreted with urine. Enterohepatic circulation of glucuronides derived from the two major 4-MBC metabolites may explain the slow excretion of 4-MBC metabolites with urine and the small percentage of the administered doses recovered in urine.
No pharmacokinetic data available.
No pharmacokinetic data available.

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Metabolism / Metabolites
Based on the findings of a rat pharmacokinetic study, it is proposed that absorbed enzacamene following oral administration undergo extensive first-pass hepatic metabolism. Following oral administration of enzacamene (4-MBC) in rats, detected metabolites in the plasma and urine were 3-(4-carboxybenzylidene)camphor and as four isomers of 3-(4-carboxybenzylidene)hydroxycamphor containing the hydroxyl group located in the camphor ring system with 3-(4-carboxybenzylidene)-6-hydroxycamphor as the major metabolite. However the blood concentrations of 3-(4-carboxybenzylidene)-6-hydroxycamphor were below the limit of detection following peak concentration. Via hydroxylation mediated by cytochrome P450 system, 3-(4-hydroxymethylbenzylidene)camphor is formed. This metabolite is further oxidized to 3-(4-carboxybenzylidene)camphor via oxidation of alcohol dehydrogenase and aldehyde dehydrogenase, and may be further hydroxylated to form 3-(4-carboxybenzylidene)-6-hydroxycamphor mediated by CYP450 system.

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Biological Half-Life
The half life of enzacamene (4-MBC) and its main metabolite, 3-(4-carboxybenzylidene)camphor, displayed half-lives of approximately 15 h after reaching peak plasma concentrations after oral administration in rats.

Protein Binding
No pharmacokinetic data available.

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- In vitro toxicity [1]
- 4-MBC showed dose-dependent cytotoxicity on HTR-8/SVneo cells: At 40 μM, the cell viability was reduced to 15% (48 h, MTT assay). It induced oxidative stress (increased ROS) and apoptotic cell death without significant necrosis (PI-positive cells <5% at 30 μM).
- In vivo developmental toxicity [2]
- Maternal toxicity: In CD-1 mice, oral administration of 4-MBC at 500 mg/kg caused mild maternal weight loss (5% vs. control) and reduced food intake (10% vs. control) during gestation, but no maternal mortality. In rats, dermal exposure to 100 mg/kg caused no obvious maternal skin irritation or systemic toxicity.
- Fetal/neonatal toxicity: As described in "In Vivo", it increased fetal resorption, reduced fetal weight, induced minor malformations (mice), and impaired neonatal survival and development (rats).

[1]. 4-Methylbenzylidene-camphor inhibits proliferation and induces reactive oxygen species-mediated apoptosis of human trophoblast cells. Reprod Toxicol. 2019 Mar:84:49-58.

[2]. Developmental toxicity of UV filters and environmental exposure: a review. Int J Androl. 2008 Apr;31(2):144-51.

Enzacamene is a monoterpenoid.
Commonly known as 4-methylbenzylidene-camphor (4-MBC), enzacamene is a camphor derivative and an organic chemical UV-B filter. It is used in cosmetic products such as sunscreen to provide skin protection against UV rays. While its effects on the human reproductive system as an endocrine disruptor are being investigated, its use in over-the-counter and cosmetic products is approved by Health Canada. Its tradenames include Eusolex 6300 (Merck) and Parsol 5000 (DSM).

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Drug Indication
Indicated for use as an active sunscreen agent.

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Mechanism of Action
Enzacamene absorbs UV-B rays. It is proposed that enzacamene exerts estrogen-like activities in the same direction as endogenous estrogens via nonclassical estrogen signaling mechanisms that do not involve gene regulation by the nuclear ER. It binds to cytosolic estradiol binding sites of estrogen receptors with low to moderate affinity compared to that of the endogenous agonist. Based on the findings of a study with _Xenopus_ hepatocytes in culture, enzacamene has a potential to induce the ER gene only at higher concentrations (10–100 μmol/L). While enzacamene was not shown to activate estrogen-dependent gene transcription when tested in an ER reporter gene assay in yeast cells, it was demonstrated in _Xenopus_ hepatocytes cultures that activate ER-dependent signaling mechanisms leading to altered gene expression. In micromolar concentrations, enzacamene accelerates cell proliferation rate in MCF-7 human breast cancer cells.

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- 4-Methylbenzylidene camphor (4-MBC) is a synthetic ultraviolet (UV) filter widely used in sunscreen products, cosmetics, and personal care items to absorb UV-B radiation [1,2]
- Mechanism of action in trophoblast cells: Its antiproliferative and pro-apoptotic effects are mediated by excessive intracellular ROS generation, which disrupts redox homeostasis and activates the mitochondrial apoptotic pathway (upregulation of Bax/Bcl-2 ratio and cleaved caspase-3) [1]
- Environmental exposure risk [2]: 4-MBC is detected in environmental water (rivers, lakes) and human biological samples (urine, breast milk) due to its widespread use and environmental persistence. It is classified as an endocrine-disrupting chemical (EDC) due to its ability to interfere with estrogen synthesis and embryonic development [2]

C18H22O

254.3667

254.167

36861-47-9

4-Methylbenzylidene camphor-d4;1219806-41-3

6434217

White to off-white solid powder

1.1±0.1 g/cm3

371.9±22.0 °C at 760 mmHg

66-68°C

168.9±13.2 °C

0.0±0.8 mmHg at 25°C

1.583

4.95

17.07

0

1

1

19

423

0

CC1=CC=C(C=C1)/C=C/2\C3CCC(C2=O)(C3(C)C)C

HEOCBCNFKCOKBX-SDNWHVSQSA-N

InChI=1S/C18H22O/c1-12-5-7-13(8-6-12)11-14-15-9-10-18(4,16(14)19)17(15,2)3/h5-8,11,15H,9-10H2,1-4H3/b14-11+

(3E)-1,7,7-trimethyl-3-[(4-methylphenyl)methylidene]bicyclo[2.2.1]heptan-2-one

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~100 mg/mL (~393.13 mM)
H2O : < 0.1 mg/mL

配方 1 中的溶解度: 2.5 mg/mL (9.83 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 悬浮液；超声助溶。
例如，若需制备1 mL的工作液，可将100 μL 25.0 mg/mL澄清DMSO储备液加入到400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: 2.5 mg/mL (9.83 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 悬浊液; 超声助溶。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.5 mg/mL (9.83 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。