On 2016 May 04, Simon Young commented:

This review raises a number of issues related to the content and interpretation of articles that are cited.

1. Paragraph 6 on page 347, about side effects of TRP, cites two placebo controlled trials of TRP (Thomson et al and Steinberg et al) and mentions side effects of TRP. However, the article fails to mention that the study by Thomson et al reported no significant differences in side effects between TRP and placebo, and that in the study of Steinberg et al only 1 of 20 side effects measured (dizziness) was significantly greater after TRP group than after placebo. The same paragraph also contains the sentence “In a study in which 3 g TRP daily was administered to participants for 12 weeks, one patient reported diarrhea as a side-effect of TRP intake (Van Praag et al., 1972)”. However, the van Praag article states “The precursor we used was not tryptophan but dl-5-HTP”. 5-Hydroxytryptohan has a different side effect profile from that of TRP.

2. Paragraph 6 on page 347 mentions that the dose of TRP in the study of Steinberg et al was 6g daily for 3 months and suggests that “Such high doses might not be recommendable not only because of such side-effects, but also because the TPH enzyme is likely to already be saturated by a dose of TRP up to 3g”. While it is true that the daily dose was 6g it was given in three doses of 2g each (after breakfast, after lunch, and before bed), which would not be likely to saturate TRP hydroxylase, and explains why only 1 of 20 side effects measured was significantly greater in the TRP group than in the placebo group. Also, the article did not mention that TRP altered social behavior, specifically a decrease in irritability, in the study of Steinberg et al.

3. The article lists the dose of TRP in the study of Volvaka et al as “6g for 3 weeks”. It fails to mention that the daily dose of 6g was given in divided doses, either 3 or 4 times per day. The article lists the dose of TRP in the study of Nemzer et al as 100mg/kg daily for 1 week. TRP was given in divided doses in the morning and afternoon. The article gives the dose of TRP in the study of Cerit et al as “2.8g per day for 6 days”. The TRP was given in 3 doses per day, 0.8g in the morning and afternoon and 1.2g in the evening. These differences are important as a divided dose is likely to increase serotonin synthesis for longer during each day than the same amount given as a single dose, as well as possibly reducing side effects.

4. Five of the articles listed in Table 1 and discussed in the text involve a technique called acute TRP depletion (ATD). The discussion of all these articles lacks important information. In ATD participants are given a mixture of amino acids that is devoid of TRP (T-), or the same amount of amino acids plus the appropriate amount of TRP as in a balanced protein (B mixture), which for the studies described is 2.3g Young SN, 2013. When the T- mixture is given the amino acid mixture induces protein synthesis and TRP in the blood and tissues is incorporated into protein. Over 5 hours TRP levels fall dramatically and the rate of serotonin synthesis in human brain can decline by more than 90% Nishizawa S, 1997. The B mixture, which contains TRP, will not increase brain TRP or serotonin synthesis, for reasons explained in the paragraph 2 of section 1.1 of the article. Sometimes in ATD studies an additional 8g of TRP is added to the B mixture to raise brain TRP (T+). The problems in the discussion of these articles are: (i) The study of Marsh et al compared the T- mixture, the B mixture, and fasting participants. None of these treatments would increase brain TRP so this was an ATD study, not a TRP supplementation study. The study demonstrated that ATD increases aggressive responding relative to the B mixture. This does not necessarily mean that TRP supplementation would have decreased aggression relative to the B mixture. The fact that the B mixture decreased aggressive responding relative to the fasting condition is irrelevant as the B treatment does not increase brain TRP. While the B and T- mixtures were given under blind conditions, the fasting day occurred after both days in which amino acid mixtures were given, and the participant knew they were not receiving an amino acid mixture. Any effect could have been due to lack of blinding or an order effect, issues not mentioned in the article. Also (a minor point) the article mentions “a control condition (i.e. a low monoamine diet)”. The participants were on a low monoamine diet throughout the study, and the control condition was fasting. (ii) The study of Bjork et al (2000) compared ATD treatment with a T+ treatment containing 10.3g TRP, and a fasting condition. The conclusion given in Fig. 1 of the article by Steenbergen et is that TRP supplementation “Decreased aggressive responding”. However, the article by Bjork et al (2000) states that “TRP depletion increased aggression relative to TRP loading in aggressive men”. Any difference could have been due to increased aggression after ATD, or decreased aggression after TRP loading, or both, so this study did not demonstrate an effect of TRP loading. (iii) The study of Cleare and Bond looked at the effect of ATD and a T+ mixture containing 10.3g TRP. As reported by Steenbergen et al TRP supplementation “was found to reduce self-report ratings of angriness, quarrelsomeness, hostility, and annoyance, but only for males with high trait levels of aggression”. These were changes over time after administration of the amino acids and were relatively small (at most 20%). Given the absence of any control group with unaltered TRP levels these changes cannot necessarily be attributed to the effect of TRP supplementation. (iv) The study of Finn et al compared the effect of a B mixture and a T- mixture. As there was no treatment that increased brain TRP this paper should not have been included in the article.

5. Section 3 of the article includes the statement “if the TPH1 enzyme in the gut is very active, more TRP is converted there and less will be available to pass through the BBB and be converted into 5-HT in the brain. Thus TRP might have less impact on social behavior in individuals with highly active TPH1 enzyme.” Under normal circumstances only about 1% of ingested TRP is converted into serotonin by TPH (see section 4, paragraph 5 of the article and SJOERDSMA A, 1956). Only in a rare condition, the carcinoid syndrome, which is due to neuroendocrine tumors metastasizing in the liver and producing large amounts of serotonin Molina-Cerrillo J, 2016, is metabolism via TPH quantitatively significant, with up to 60% of TRP converted to serotonin SJOERDSMA A, 1956. The main catabolic route of TRP is along the kynurenine pathway, and flux along this pathway is increased in patients with major depressive disorder Teraishi T, 2015. However, TRP is an effective antidepressant according to a Cochrane review Shaw K, 2002. Therefore, it is unlikely that increased peripheral catabolism of TRP will mitigate the effect of TRP on social behavior to any great extent.

6. Section 4 of the article states “some of the effects of TRP administration on social behavior might in fact be a result of enhanced sleep and mood”. An effect on sleep cannot apply to the majority of the studies discussed, which were carried out during a single day. When TRP was given for many days TRP was usually given in a divided dose with only a small dose (usually around 1g) given in the evening, sometimes with dinner. When TRP is given as a hypnotic it is usually given shortly before bedtime, as plasma TRP rises quickly after ingestion. Whether the low doses of TRP given in the evening in the studies lasting longer than a day would have had an effect on sleep is not clear. In relation to improved mood as a mediator of more positive social behavior, the study of Moskovitz et al (2001) demonstrated that TRP resulted in more positive social behavior without any change in mood.

On 2016 May 04, Simon Young commented:

This review raises a number of issues related to the content and interpretation of articles that are cited.

1. Paragraph 6 on page 347, about side effects of TRP, cites two placebo controlled trials of TRP (Thomson et al and Steinberg et al) and mentions side effects of TRP. However, the article fails to mention that the study by Thomson et al reported no significant differences in side effects between TRP and placebo, and that in the study of Steinberg et al only 1 of 20 side effects measured (dizziness) was significantly greater after TRP group than after placebo. The same paragraph also contains the sentence “In a study in which 3 g TRP daily was administered to participants for 12 weeks, one patient reported diarrhea as a side-effect of TRP intake (Van Praag et al., 1972)”. However, the van Praag article states “The precursor we used was not tryptophan but dl-5-HTP”. 5-Hydroxytryptohan has a different side effect profile from that of TRP.

2. Paragraph 6 on page 347 mentions that the dose of TRP in the study of Steinberg et al was 6g daily for 3 months and suggests that “Such high doses might not be recommendable not only because of such side-effects, but also because the TPH enzyme is likely to already be saturated by a dose of TRP up to 3g”. While it is true that the daily dose was 6g it was given in three doses of 2g each (after breakfast, after lunch, and before bed), which would not be likely to saturate TRP hydroxylase, and explains why only 1 of 20 side effects measured was significantly greater in the TRP group than in the placebo group. Also, the article did not mention that TRP altered social behavior, specifically a decrease in irritability, in the study of Steinberg et al.

3. The article lists the dose of TRP in the study of Volvaka et al as “6g for 3 weeks”. It fails to mention that the daily dose of 6g was given in divided doses, either 3 or 4 times per day. The article lists the dose of TRP in the study of Nemzer et al as 100mg/kg daily for 1 week. TRP was given in divided doses in the morning and afternoon. The article gives the dose of TRP in the study of Cerit et al as “2.8g per day for 6 days”. The TRP was given in 3 doses per day, 0.8g in the morning and afternoon and 1.2g in the evening. These differences are important as a divided dose is likely to increase serotonin synthesis for longer during each day than the same amount given as a single dose, as well as possibly reducing side effects.

4. Five of the articles listed in Table 1 and discussed in the text involve a technique called acute TRP depletion (ATD). The discussion of all these articles lacks important information. In ATD participants are given a mixture of amino acids that is devoid of TRP (T-), or the same amount of amino acids plus the appropriate amount of TRP as in a balanced protein (B mixture), which for the studies described is 2.3g Young SN, 2013. When the T- mixture is given the amino acid mixture induces protein synthesis and TRP in the blood and tissues is incorporated into protein. Over 5 hours TRP levels fall dramatically and the rate of serotonin synthesis in human brain can decline by more than 90% Nishizawa S, 1997. The B mixture, which contains TRP, will not increase brain TRP or serotonin synthesis, for reasons explained in the paragraph 2 of section 1.1 of the article. Sometimes in ATD studies an additional 8g of TRP is added to the B mixture to raise brain TRP (T+). The problems in the discussion of these articles are: (i) The study of Marsh et al compared the T- mixture, the B mixture, and fasting participants. None of these treatments would increase brain TRP so this was an ATD study, not a TRP supplementation study. The study demonstrated that ATD increases aggressive responding relative to the B mixture. This does not necessarily mean that TRP supplementation would have decreased aggression relative to the B mixture. The fact that the B mixture decreased aggressive responding relative to the fasting condition is irrelevant as the B treatment does not increase brain TRP. While the B and T- mixtures were given under blind conditions, the fasting day occurred after both days in which amino acid mixtures were given, and the participant knew they were not receiving an amino acid mixture. Any effect could have been due to lack of blinding or an order effect, issues not mentioned in the article. Also (a minor point) the article mentions “a control condition (i.e. a low monoamine diet)”. The participants were on a low monoamine diet throughout the study, and the control condition was fasting. (ii) The study of Bjork et al (2000) compared ATD treatment with a T+ treatment containing 10.3g TRP, and a fasting condition. The conclusion given in Fig. 1 of the article by Steenbergen et is that TRP supplementation “Decreased aggressive responding”. However, the article by Bjork et al (2000) states that “TRP depletion increased aggression relative to TRP loading in aggressive men”. Any difference could have been due to increased aggression after ATD, or decreased aggression after TRP loading, or both, so this study did not demonstrate an effect of TRP loading. (iii) The study of Cleare and Bond looked at the effect of ATD and a T+ mixture containing 10.3g TRP. As reported by Steenbergen et al TRP supplementation “was found to reduce self-report ratings of angriness, quarrelsomeness, hostility, and annoyance, but only for males with high trait levels of aggression”. These were changes over time after administration of the amino acids and were relatively small (at most 20%). Given the absence of any control group with unaltered TRP levels these changes cannot necessarily be attributed to the effect of TRP supplementation. (iv) The study of Finn et al compared the effect of a B mixture and a T- mixture. As there was no treatment that increased brain TRP this paper should not have been included in the article.

5. Section 3 of the article includes the statement “if the TPH1 enzyme in the gut is very active, more TRP is converted there and less will be available to pass through the BBB and be converted into 5-HT in the brain. Thus TRP might have less impact on social behavior in individuals with highly active TPH1 enzyme.” Under normal circumstances only about 1% of ingested TRP is converted into serotonin by TPH (see section 4, paragraph 5 of the article and SJOERDSMA A, 1956). Only in a rare condition, the carcinoid syndrome, which is due to neuroendocrine tumors metastasizing in the liver and producing large amounts of serotonin Molina-Cerrillo J, 2016, is metabolism via TPH quantitatively significant, with up to 60% of TRP converted to serotonin SJOERDSMA A, 1956. The main catabolic route of TRP is along the kynurenine pathway, and flux along this pathway is increased in patients with major depressive disorder Teraishi T, 2015. However, TRP is an effective antidepressant according to a Cochrane review Shaw K, 2002. Therefore, it is unlikely that increased peripheral catabolism of TRP will mitigate the effect of TRP on social behavior to any great extent.

6. Section 4 of the article states “some of the effects of TRP administration on social behavior might in fact be a result of enhanced sleep and mood”. An effect on sleep cannot apply to the majority of the studies discussed, which were carried out during a single day. When TRP was given for many days TRP was usually given in a divided dose with only a small dose (usually around 1g) given in the evening, sometimes with dinner. When TRP is given as a hypnotic it is usually given shortly before bedtime, as plasma TRP rises quickly after ingestion. Whether the low doses of TRP given in the evening in the studies lasting longer than a day would have had an effect on sleep is not clear. In relation to improved mood as a mediator of more positive social behavior, the study of Moskovitz et al (2001) demonstrated that TRP resulted in more positive social behavior without any change in mood.