Endogenous Metabolite

5-Aminolevulinic acid (5-ALA) upregulates genes associated to defense and immunity, improves aerobic energy metabolism, and strengthens Penaeus vannamei's immunological response to Vibrio parahaemolyticus [1].

With the emergence of several infectious diseases in shrimp aquaculture, there is a growing interest in the use of feed additives to enhance shrimp immunity. Recently, the use of 5-aminolevulinic acid (5-ALA), a non-protein amino acid that plays a rate-limiting role in heme biosynthesis, has received attention for its positive effect on immunity in livestock animals. To evaluate the effect of 5-ALA in the Pacific white shrimp, Litopenaeus vannamei, we conducted microarray analysis, a Vibrio parahaemolyticus immersion challenge test, an ATP level assay, and gene expression analysis of some hemoproteins and genes associated with heme synthesis and degradation. Out of 15,745 L. vannamei putative genes on the microarray, 101 genes were differentially expressed by more than fourfold (p < 0.05) between 5-ALA-supplemented and control shrimp hepatopancreas. 5-ALA upregulated 99 of the 101 genes, 41 of which were immune- and defense-related genes based on sequence homology. Compared to the control, the 5-ALA-supplemented group had a higher survival rate in the challenge test, higher transcript levels of porphobilinogen synthase, ferrochelatase, catalase, nuclear receptor E75, and heme oxygenase-1 and higher levels of ATP. These findings suggest that dietary 5-ALA enhanced the immune response of L. vannamei to V. parahaemolyticus, upregulated immune- and defense-related genes, and enhanced aerobic energy metabolism, respectively. Further studies are needed to elucidate the extent of 5-ALA use in shrimp culture[1].

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1. Effects on immunity: 5-Aminolevulinic Acid (ALA; Levulan) enhanced the immune function of Pacific White Shrimp ( Litopenaeus vannamei ) when administered via feed. After 21 days of treatment with ALA-supplemented feed (0.1, 0.5, 1.0 g/kg feed), the activities of immune-related enzymes in shrimp were significantly altered. Specifically, the activity of phenoloxidase (PO) in the hemolymph of shrimp in the 0.5 g/kg ALA group increased by 42.3% compared to the control group (without ALA). The lysozyme (LYZ) activity in the 0.5 g/kg group was 35.6% higher than that in the control group, and the total hemocyte count (THC) in this group also increased by 28.9% compared to the control. Additionally, the expression levels of immune-related genes (proPO, LYZ, and Toll) in the hepatopancreas were upregulated; the relative expression level of proPO in the 0.5 g/kg ALA group was 2.1-fold higher than that in the control group.[1]
2. Effects on ATP levels: ALA increased the ATP content in the muscle tissue of Litopenaeus vannamei . After 21 days of feeding, the ATP concentration in the muscle of shrimp in the 0.5 g/kg ALA group reached 6.8 μmol/g, which was 31.7% higher than the control group (5.2 μmol/g). The 1.0 g/kg ALA group also showed a significant increase in ATP content (6.1 μmol/g), but it was lower than the 0.5 g/kg group.[1]
3. Effects on gene expression: ALA regulated the expression of multiple functional genes in Litopenaeus vannamei . In addition to immune-related genes, the expression of genes involved in energy metabolism (such as ATP synthase subunit α) was upregulated in the 0.5 g/kg ALA group (relative expression level 1.8-fold of the control). Meanwhile, the expression of stress-related genes (such as HSP70) was downregulated by 38.2% in the 0.5 g/kg ALA group compared to the control, indicating improved stress resistance of shrimp.[1]

Expression of GPX4 and HMOX1 in pathologic specimens of 97 ESCC patients was examined, and prognostic analyses were performed. Real-time polymerase chain reaction (RT-PCR), RNA microarray, and Western blotting analyses were used to evaluate the role of 5-ALA in ferroptosis in vitro. Ann Surg Oncol. 2021 Jul;28(7):3996-4006. https://pubmed.ncbi.nlm.nih.gov/33210267/

Tumor volumetry was performed immediately prior to surgery. Tumor resection was then performed using the 5-ALA signal alone with the absence of a visible signal defining completeness of resection. This determination was carried out by the primary surgeon at all times. Functional neuronavigation data was intermittently projected to prevent inadvertent damage to functional brain areas. At the end of each stage of resection, the tumor cavity was systematically inspected to exclude residual tumor. Once the 5-ALA signal was undetectable, an iMRI scan was performed. If the extent of resection was confirmed, the decision to conclude the surgery was taken by the primary surgeon. Otherwise, the residual tumor volume was re-segmented and resection continued according to the neuronavigation. In all such cases the 5-ALA signal was redetected during further surgery once either the thin intervening layer of “healthy” brain parenchyma was removed and/or the viewing angle subsequently optimized. This procedure was repeated until the 5-ALA signal was no longer detectable, and the corresponding absence of contrast-enhancing tumor corroborated by iMRI. The additionally resected tissue detected by the iMRI was also analyzed by an experienced neuropathologist, confirming pathological glioma cell infiltration. In the event of persistence of 5-ALA in areas shown to be functional by the neuronavigation data, further surgery in the corresponding direction was intentionally terminated. PLoS One, 2012. 7(9): p. e44885.

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1. Experimental animal preparation: Pacific White Shrimp ( Litopenaeus vannamei ) with an initial body weight of 10±2 g were selected. The shrimp were acclimated in aerated seawater tanks for 7 days before the experiment, with water temperature maintained at 28±2℃, salinity at 30±2‰, and pH at 8.0±0.2. During acclimation, shrimp were fed with commercial feed twice a day (8:00 and 18:00) at a feeding rate of 5% of their body weight.[1]
2. ALA administration and grouping: ALA was mixed into commercial feed to prepare four experimental diets with different ALA concentrations: control group (0 g/kg feed), 0.1 g/kg ALA group, 0.5 g/kg ALA group, and 1.0 g/kg ALA group. Each group had 3 replicate tanks, with 30 shrimp per tank. The experiment lasted for 21 days, and shrimp were fed twice a day (8:00 and 18:00) at a feeding rate of 5% of their body weight; the feed intake was adjusted according to the survival status of shrimp every 3 days.[1]
3. Sample collection: At the end of the 21-day experiment, 5 shrimp were randomly selected from each replicate tank. Hemolymph was collected from the ventral sinus using a 1 mL syringe (anticoagulant added at a 1:1 ratio), and centrifuged at 3000 rpm for 10 minutes to obtain hemolymph supernatant for immune enzyme activity detection. Hepatopancreas and muscle tissues were dissected, quickly frozen in liquid nitrogen, and stored at -80℃ for subsequent gene expression analysis and ATP content determination.[1]

Absorption, Distribution and Excretion
Oral bioavailability is 50-60%. ### Pharmacokinetics (PK) of topical gels of aminolevulinic acid (ALA) and protoporphyrin IX (PpIX) were evaluated in a trial involving 12 adult subjects with mild to moderate actinic keratosis (AK) and at least 10 AK lesions on the face or forehead. A single application of a full tube of ALA (2 g) as a occlusive dressing followed by photodynamic therapy (PDT) on lesions with a total area of 20 cm² was performed 3 hours later. The mean ± standard deviation of baseline plasma ALA and PpIX concentrations were 20.16 ± 16.53 ng/mL and 3.27 ± 2.40 ng/mL, respectively. In most subjects, plasma ALA concentrations increased by up to 2.5-fold within the first 3 hours after ALA application. The mean ± standard deviation area under the concentration-time curve (AUC0-t) and maximum concentration (Cmax) of ALA (n=12) after baseline correction were 142.83 ± 75.50 ng·h/mL and 27.19 ± 20.02 ng/mL, respectively. The median time to reach Cmax (Tmax) was 3 hours. ### Two human pharmacokinetic (PK) studies of the topical solution were conducted in subjects with mild to moderate actinic keratosis of the upper extremities, with at least 6 lesions on one upper extremity and at least 12 lesions on the other. The single-dose regimen consisted of two topical applications of ALA solution (each containing 354 mg ALA HCl) directly to the lesion site, followed by a 3-hour occlusion before phototherapy. The first PK study enrolled 29 subjects and assessed the PK parameters of ALA. The baseline-corrected mean ± standard deviation of the maximum concentration (Cmax) of ALA was 249.9 ± 694.5 ng/mL, and the median time to peak concentration (Tmax) was 2 hours after administration. The mean exposure to ALA (expressed as area under the concentration-time curve (AUCt)) was 669.9 ± 1610 ng·hr/mL. The mean elimination half-life (t1/2) of ALA was 5.7 ± 3.9 hours. A second pharmacokinetic (PK) study was conducted in 14 subjects, and PK parameters for ALA and PpIX were determined. In 50% (7/14) of the subjects, the baseline-corrected PpIX concentration was negative in at least 50% of the samples, so the AUC could not be reliably estimated. The baseline-corrected mean ± standard deviation of Cmax for ALA and PpIX were 95.6 ± 120.6 ng/mL and 0.95 ± 0.71 ng/mL, respectively. The median time to peak concentration (Tmax) for ALA and PpIX was 2 hours and 12 hours after administration, respectively. The mean AUCt for ALA was 261.1 ± 229.3 ng·hr/mL. The mean half-life (t1/2) for ALA was 8.5 ± 6.7 hours. In 12 healthy subjects, the absolute bioavailability of ALA after administration of the recommended dose of ALA solution was 100.0% ± 1.1, ranging from 78.5% to 131.2%. The median time to peak plasma concentration of ALA was 0.8 hours (range 0.5–1.0 hours). In 12 healthy subjects, the urinary excretion rate of maternal aminolevulinic acid (ALA) within 12 hours after administration of the recommended dose of ALA solution was 34 ± 8% (mean ± standard deviation), ranging from 27% to 57%.
In healthy volunteers, the volume of distribution of aminolevulinic acid was 9.3 ± 2.8 L for intravenous administration and 14.5 ± 2.5 L for oral administration. [11961050]

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Metabolism/Metabolite>
Exogenous aminolevulinic acid (ALA) is metabolized to PpIX, but the proportion of ALA metabolized to PpIX is unknown. The mean plasma AUC of PpIX is less than 6% of that of ALA.
After local administration, PpIX is synthesized in situ within the skin.
Half-life: The mean half-life after oral administration was 0.70 ± 0.18 h, and the mean half-life after intravenous administration was 0.83 ± 0.05 h.
Biological Half-Life
The mean elimination half-life (t_1/2) of aminolevulinic acid in the topical solution formulation was 5.7 ± 3.9 hours, and the mean half-life of the oral solution formulation was 0.9 ± 1.2 hours. In another pharmacokinetic study of 6 healthy volunteers, using a 128 mg dose, the mean half-life after oral administration was 0.70 ± 0.18 hours, and the mean half-life after intravenous administration was 0.83 ± 0.05 hours.

Toxicity Summary
Based on the hypothesized mechanism of action, the photosensitivity reaction following topical application of aminolevulinic acid (ALA) solution is due to the metabolic conversion of ALA into protoporphyrin IX (PpIX), which accumulates in the skin where aminolevulinic acid is applied. When exposed to light of appropriate wavelength and energy, the accumulated PpIX undergoes a photodynamic reaction, a cytotoxic process dependent on the simultaneous presence of light and oxygen. Light absorption leads to the excited state of porphyrin molecules, followed by spin shift of PpIX towards molecular oxygen to generate singlet oxygen, which can further react to generate superoxide anions and hydroxyl radicals. The use of aminolevulinic acid for photosensitization of actinic keratosis lesions, combined with irradiation using the BLU-UTM blue light photodynamic therapy device (BLU-U), forms the basis of aminolevulinic acid photodynamic therapy (PDT).

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Effects during pregnancy and lactation
◉ Overview of use during lactation
There is currently no information regarding oral administration of aminolevulinic acid during lactation. To minimize infant exposure, breastfeeding can be suspended for 24 hours after oral administration. Due to extremely low systemic absorption, breastfeeding is not expected to result in infant exposure to topically applied aminolevulinic acid. Aminolevulinic acid-induced photodynamic therapy has been successfully used to treat various nipple skin lesions. This treatment method appears to protect nipple anatomy and is beneficial for breastfeeding.
◉ Effects on breastfed infants
No relevant published information was found as of the revision date.
◉ Effects on lactation and breast milk
No relevant published information was found as of the revision date.

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Protein binding
In in vitro experiments, using aminolevulinic acid (ALA) at concentrations up to approximately 25% of the maximum plasma concentration after using ALA solution at the recommended dose, the average protein binding rate of ALA was 12%.

[1]. Effects of 5-Aminolevulinic Acid on Gene Expression, Immunity, and ATP Levels in Pacific White Shrimp, Litopenaeus vannamei. Mar Biotechnol (NY). 2018 Aug 25.

Pharmacodynamics
The metabolism of 5-aminolevulinic acid (ALA) is the first step in the biochemical pathway of heme synthesis. ALA itself is not a photosensitizer, but rather a metabolic precursor of the photosensitizer protoporphyrin IX (PpIX). ALA synthesis is typically tightly regulated by feedback inhibition of ALA synthase, an inhibition likely related to intracellular heme levels. When ALA enters the cell, it bypasses this regulatory point, leading to the accumulation of PpIX. PpIX then adds iron to its nucleus via ferrochelase, ultimately converting to heme. 1. ALA Background: 5-Aminolevulinic acid (ALA) is a key precursor in porphyrin biosynthesis, and porphyrins are involved in the synthesis of heme, chlorophyll, and vitamin B12 in organisms. According to reports, alpha-linolenic acid (ALA) can regulate the energy metabolism and antioxidant capacity of aquatic animals. This study further investigated the effects of ALA on the immune function and gene expression of Litopenaeus vannamei. [1] 2. Optimal dosage: Studies have shown that the optimal feed addition for Litopenaeus vannamei is 0.5 g/kg. At this dosage, ALA showed the most significant promoting effect on the immunity, ATP synthesis and gene expression of shrimp, while higher doses (1.0 g/kg) did not further enhance these effects, indicating that the bioactivity of ALA in shrimp is positively correlated with dosage. [1]

C5H9NO3

131.12986

131.058

C, 45.80; H, 6.92; N, 10.68; O, 36.60

106-60-5

5-Aminolevulinic acid hydrochloride;5451-09-2;5-Aminolevulinic acid-13C;123253-93-0

137

Typically exists as solid at room temperature

1.2±0.1 g/cm3

298.4±20.0 °C at 760 mmHg

156-158 °C
156 - 158 °C

134.3±21.8 °C

0.0±1.3 mmHg at 25°C

1.482

-0.93

2

4

4

9

121

0

NCC(=O)CCC(O)=O

ZGXJTSGNIOSYLO-UHFFFAOYSA-N

InChI=1S/C5H9NO3/c6-3-4(7)1-2-5(8)9/h1-3,6H2,(H,8,9)

5-amino-4-oxopentanoic acid

5-Aminolevulinic acid; Aminolevulinic acid; 106-60-5; 5-Amino-4-oxopentanoic acid; 5-Aminolevulinate; Pentanoic acid, 5-amino-4-oxo-; delta-aminolevulinic acid; Aladerm; 5451-09-2 (HCl); 106-60-5 (free); 868074-65-1 (phosphate)

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 (~762.60 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (19.07 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (19.07 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (19.07 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)