动物实验给药剂量及溶剂选择
实验动物相关生理参数
Table 1. Relevant physiological parameters for different animals1
| Animal species | Weight (kg) | Blood volume (mL) | Blood flow (mL/min) | Total surface area (m^2) | Life-span (year) | Normal oral (mL/kg) | Normal i.v. (mL/kg) |
|---|---|---|---|---|---|---|---|
| Mouse | 0.02 | 1.7 | Liver: 1.8; kidney: 1.3 | 0.008 | 2.7 | 10 | 5 |
| Rat | 0.25 | 13.5 | Liver: 13.8; kidney: 9.2 | 0.023 | 4.7 | 5–10 | 2.5 |
| Rabbit | 2.5 | 165 | Liver: 177; kidney: 80 | 0.17 | 8.0 | 5–10 | 1–2 |
| Rhesus monkey | 5 | 367 | Liver: 218; kidney: 138 | 0.32 | 22 | 5–8 | 0.5–1 |
| Dog | 10 | 900 | Liver: 309; kidney: 216 | 0.51 | 20 | 5–8 | 0.5–1 |
不同途径推荐给药体积
Table 2. Recommended Dose Volumes for Common Laboratory Animals2
| Species | Dose Volume | Oral (mL/kg) | IV bolus (mL/kg)* | IP (mL/kg)# | SC (mL/kg)# | IM (mL/kg)# |
|---|---|---|---|---|---|---|
| Mouse | Ideal | 10 | 5 | 5–10 | 5 | 0.05 total mL/site (2 sites/day) |
| Maximum | 20 | 25 (slow injection) | 20 | 20 (divided in 2–3 sites) | 0.1 total mL/site (2 sites/day) | |
| Rat | Ideal | 10 | 5 | 5–10 | 5 | 0.1 total mL/site (2 sites/day) |
| Maximum | 20 | 20 (slow injection) | 20 | 10 (divided in 2–3 sites) | 0.2 total mL/site (2 sites/day) | |
| Hamster | Ideal | 10 | 5 | 5–10 | 5 | 0.1 total mL/site (2 sites/day) |
| Maximum | 20 | 20 (slow injection) | 20 | 10 (divided in 2–3 sites) | 0.2 total mL/site (2 sites/day) | |
| Guinea Pig | Ideal | 10 | 1 | 10 | 5 | 0.1 total mL/site (2 sites/day) |
| Maximum | 20 | 5 (slow injection) | 20 | 10 (divided in 2–3 sites) | 0.2 total mL/site (2 sites/day) | |
| Rabbit | Ideal | 10 | 1–5 | 3–5 | 2.5 | 0.25 |
| Maximum | 20 (empty stomach) | 10 (slow injection) | 10 (rare) | 10 (divided in 2–3 sites) | 0.5 (Max 1 mL limit) | |
| Cat | Ideal | 10 | 5 | 5 | 2 | 0.25 |
| Maximum | 15 | 10 (slow injection) | 20 | 5 | 0.5 (Max 1 mL limit) | |
| Ferret | Ideal | 10 | 5 | 5 | 2 | 0.25 |
| Maximum | 15 | 10 (slow injection) | 20 | 5 | 0.5 (Max 1 mL limit) | |
| Marmoset | Ideal | 10 | 2.5–5 | 5 | 2 | 0.25 |
| Maximum | 15 | 10 (slow injection) | 20 | 5 | 0.1 total mL/site (2 sites/day) | |
| Dog | Ideal | 5–8 | 1–5 | 1 | 1 | 0.25 |
| Maximum | 15 | 10 (slow injection) | 20 | 2 (divided in 2–3 sites) | 0.5 (Max 3 mL limit) | |
| Macaque | Ideal | 5–8 | 1–5 | 3 | 1 | 0.25 |
| Maximum | 15 | 10 (slow injection) | 10 | 2 (divided in 2–3 sites) | 0.5 (Max 2 mL limit) | |
| Minipig | Ideal | 10 | 1–5 | 1 | 1 | 0.25 |
| Maximum | 15 | 10 (slow injection) | 20 | 3 (divided in 2–3 sites) | 0.5 (Max 5 mL limit per site) |
- * Bolus injections are typically dosed in less than 1 minute. Slow intravenous injections are typically dosed over 3-10 minutes. Solution properties such as tonicity, pH, etc. must be taken into account when approaching the volume limits or determining the volume to be infused IV. The recommended working range for pH is 4.5 to 8.0. The order of degree of tolerance of pH for different dosing routes is oral > intravenous > intramuscular > subcutaneous > intraperitoneal. Animal health must also be taken into consideration, such as kidney function and cardiovascular function. These systems must be normal to handle increased fluid volumes.
- * When larger volumes are to be administered, consider the rate of metabolism of osmolytes with respect to route of administration since apparently innocuous solutions can be rapidly metabolized leaving an equivalent of water to dilute body fluids, which can result in hyponatremia depending on the rate of administration and/or total volume administered.
- # When administering a solution IP, SC or IM, the viscosity, concentration, tonicity, and pH of the solution need to be taken into account.
给药溶剂选择
对于水溶性良好的化合物，可以直接使用生理盐水（Saline） 或 PBS 配制。
但大多数小分子化合物水溶性通常不好，可以先使用 DMSO 溶解后，再使用其他组分稀释。MCE 和 Selleck 官网给出溶剂体系有：
| 步骤 | 溶剂 | 终浓度 |
|---|---|---|
| 1 | DMSO | 10% |
| 2 | Saline | 90% |
注：一般细胞实验中 DMSO 含量终浓度尽量在 0.1% 以下，而在动物实验中，对于正常鼠，建议 DMSO 的终浓度不大于 10%；对于裸鼠或体弱鼠，建议 DMSO 的终浓度尽量不要超过 2%3。
| 步骤 | 溶剂 | 终浓度 |
|---|---|---|
| 1 | DMSO | 10% |
| 2 | PEG300 | 40% |
| 3 | Tween 80 | 5% |
| 4 | Saline | 45% |
注：PEG300 在静脉和肌肉注射剂型中的终浓度可达 50%，无任何毒性作用。口服时，PEG300 的最高浓度可达 90%4。
| 步骤 | 溶剂 | 终浓度 |
|---|---|---|
| 1 | DMSO | 10% |
| 2 | Corn oil | 90% |
注：Corn oil 推荐用于腹腔注射或灌胃，优选灌胃，不能用于静脉注射5。
| 步骤 | 溶剂 | 终浓度 |
|---|---|---|
| 1 | DMSO | 10% |
| 2 | 20% SBE-β-CD in Saline | 90% |
注：SBE-β-CD 是增加疏水性药物溶解度的理想媒介，已在 FDA 批准的多种注射剂和众多临床候选药物中使用。SBE-β-CD 溶液可以用于多种给药方式，例如腹腔、灌胃、静脉等6。
如果上述体系仍不溶解，可以参照 Table 3 尝试不同溶剂组分，并在剂量范围内使用。
Table 3. Recommend excipient(s) use range in early formulations for oral and i.v.1
| Empty Cell | Solvent or excipient with full name* | Dosing route/common usage range# |
|---|---|---|
| Aqueous | Water | oral, i.v. |
| 0.9% NaCl | i.v. | |
| D5W – 5% dextrose in water | i.v. | |
| Buffered solutions pH: 2–8 | oral, i.v. | |
| Cosolvent | NMP – N-methylpyrrolidon | 10–20% (oral, i.v.) |
| DMSO – dimethyl sulfoxide | 10–20% (oral or i.v.) | |
| Ethanol | 10% (oral, i.v.) | |
| DMA – N,N-dimethylacetamide | 10–30% (i.v.) | |
| PG – propylene glycol | 30–60% (oral, i.v.) | |
| PEG400 – polyethylene glycol 400 | 40–100% (oral, i.v.) | |
| Transcutol – diethylene glycol monoethyl ether | 30% (oral) | |
| Cyclodextrin | HPβCD – hydroxyl-β-cyclodextrin | 20–40% (oral, i.v.) |
| SBEβCD – sulfobutylether-β-cyclodextrin | 20–40% (oral, i.v.) | |
| Surfactant | Posorbate 80 (Tween 80) – polyoxyethylene-sorbitanmonooleate 80 | 5–10% (oral, i.v.) |
| Cremophor EL – polyoxyl-35 castor oil | 5–10% (oral, i.v.) | |
| Cremophor RH40 – polyoxyl 40 hydrogenated castor oil | 5–10% (oral, i.v.) | |
| Sodium cholate | 10–20% (oral, i.v.) | |
| Pluronic F68 – or Poloxamer 188: 81% polyethylene glycol and 19% of polypropylene glycol | 20–50% (oral, i.v.) | |
| Solutol HS-15 – macrogol-15-hydroxystearate | 20–50% (oral, i.v.) | |
| VitE-TPGS 1000 – d-α-tocopheryl polyethyl glycol 1000 succinate | 20–50% (oral) | |
| Gelucire 44/14 – lauroyl macrogol-32 glycerides | 20–50% (oral) | |
| Labrasol – caprylocaproyl macrogol-8-glycerides | 40–60% (oral); 20–40% (i.v.) | |
| Lecithin – phosphatidylcholin | 20–50% (oral, i.v.) | |
| Lipid | Soybean oil | 50–100% (oral) |
| Miglyol 812 – mid-chain triglyceride of caprylic/caprolic acid | 60–100% (oral); 20–40% (i.v.) | |
| Labrafil 1944CS – polyxoyethyllated oleic glycerides | 30–60% (oral, i.v.) | |
| Capmul MCM – medium chain mono- and diglycerides | 30–60% (oral, i.v.) |
- * Most of the cosolvents, surfactants or cyclodextrins can be used in combination with pH adjustment in weak base/acid drugs for synergistic solubility enhancement.
- # The concentration range is based on mouse or rat model; administration volume is approx. 10 mL/kg for a single dosing.
References
[1] Developing early formulations: Practice and perspective
[2] IQ-CRO Recommended Dose Volumes for Common Laboratory Animals
[3] 动物实验溶剂大 Battle！
[4] PEG300 (Polyethylene glycol 300，聚乙二醇)
[5] Administration Of Drugs and Experimental Compounds in Mice and Rats (IACUC)