4.2.4. Interactions of Piperine with Medicinal Products and Other Substances

The pharmacokinetic interactions of piperine with various chemically and pharmacologically diverse drugs have been observed in both human and animal studies. In most instances, interactions of piperine with drugs resulted in a better bioavailability of the investigated drugs, exemplified by increases in the maximum plasma/serum concentrations (Cmax) and/or increased AUC values (AUC = area under the curve) for the respective drugs. These effects are in line with the purported "bio-enhancing" activity of piperine.

In human studies, oral administration of 20 mg piperine/day (~ 0,29 mg piperine/kg bw based on a body weight of 70 kg) for one or several days resulted in improved bioavailability (elevated serum/plasma concentrations and/or elevated AUC values) of the following drugs: propranolol (antihypertensive drug), theophylline (bronchodilatory drug), phenytoin, carbamazepine (antiepileptic drugs), nevirapine (HIV-1 reverse transcriptase inhibitor), chlorzoxazone (muscle relaxant), diclofenac (non-steroidal anti-inflammatory drug) and fexofenadine (antihistaminic drug). In the cases involving administration of 20 mg of piperine/day, the increases in the drug Cmax and AUC values were approximately 1.07- to 2.2-fold (Cmax) and 1.09- to 2.7-fold (AUC values), depending on the drug and drug dosage. Regarding rifampicin (antibiotic drug), such interactions were reported in the context of concomitant administration of 50 mg piperine/day (for details, see Table 2) [72–77,79,120–122].

For midazolam (sedative), prolonged and increased pharmacological effects of the drug (prolonged duration of sedation, increased number of individuals with amnesia) were observed with piperine administration of 15 mg/day for 3 days and subsequent midazolam administration on the 4th day [78].

For the substances β-carotene and coenzyme Q10, increased bioavailability was already observed in the case of combined administration of 5 mg piperine/day with 15 mg βcarotene/day for 14 days or with 120 mg coenzyme Q10/day, respectively, for 21 days [123,124]. Regarding curcumin, the Cmax or AUC values were approx. 30 or 20 times higher, respectively, when administered together with 20 mg piperine/day [125].


**Table 2.** Effects of piperine on the bioavailability of drugs in human studies.

In these human studies referred to above, piperine was administered once daily as a bolus and the piperine and/or drug administration was performed for one or more days according to different administration schedules. The effects of piperine were influenced by the dose of the administered drug and/or the duration of the piperine or the drug administration and/or possibly by the investigated population group, i.e., healthy individuals or individuals suffering from certain diseases (for details, see Table 2 and below). With the exception of the studies on β-carotene and coenzyme Q10, currently no published human

study could be identified with simultaneous longer-term piperine and drug/substance administration to achieve steady-state levels of piperine and the drug/substance.

In animal studies (rodents or rabbits), higher piperine doses were generally used (2.1–30 mg/kg bw/day), as compared to the doses employed in human studies in which interactions were investigated. The interactions described for the animal studies in most cases also resulted in increased bioavailability (increased serum/plasma Cmax and/or increased AUC values) of the investigated drugs. However, interactions in animal studies were observed with several additional drugs, such as ibuprofen, nimesulide, oxyphenylbutazone (non-steroidal anti-inflammatory drugs); amoxycillin, ampicillin, norfloxacin, marbofloxacin, metronidazole (antibiotic drugs); glimepiride, nateglinide (antidiabetic drugs); simvastatin, rosuvastatin (lipid-lowering drugs); verapamil, diltiazeme (calcium channel blockers); sodium valproate (antiepileptic drug); darunavir ethanolate (HIV protease inhibitor); losartan (angiotensin II receptor type 1 antagonist); domperidone (antiemetic drug); almotriptan (anti-migraine drug); and fexofenadine (antihistaminic drug) [126–144].

Increased bioavailability in animal studies has also been observed for a number of other substances (for example for puerarin, resveratrol, emodin, linarin or cannabidiol).

In the case of the drugs nimesulide, oxyphenylbutazone, ibuprofen or nateglinide, the piperine-mediated elevated bioavailability was reported to also be associated with increased drug efficacy. Moreover, increased pharmacological effects, without concomitant measurement of drug levels, have been observed for pentobarbitone (short-acting barbiturate), pentazocine (opioid), sertraline (selective serotonin reuptake inhibitor), midazolam or diazepam [126,134,140,141,145,146].

By contrast, animal studies were also identified in which no improved drug bioavailability or a decreased bioavailability was observed for the combined administration of piperine with drugs, i.e., with cefadroxil, carbamazepine, warfarin or diltiazem [127,147–149].

For diltiazem, a reduced drug bioavailability was observed with administration of 10 or 20 mg of piperine/kg bw/day for 14 days and diltiazem administration on Day 15 (i.e., one day after termination of piperine administration). The repeated dosing of piperine in this study led to induced gene expression of the multidrug transporter P-glycoprotein, which may have played a role in limiting the bioavailability of diltiazem [147].

As a further example, co-administration of warfarin (2 mg/kg bw) with piperine (10 mg/kg bw) to rats resulted in a decrease in the warfarin Cmax and AUC values by 32 and 20%, respectively, combined with a reduced clinical anti-coagulant effect of warfarin [149].

The influence of the study conditions (investigated species, investigated human population, duration of drug treatment and/or schedule of piperine application and piperine dose) on the results of available interaction studies is illustrated by data obtained with carbamazepine. In healthy humans receiving a single dose of carbamazepine (200 mg) on Day 11 after 10 days of piperine administration (20 mg/day), significant increases in Cmax (1.7 fold) and AUC (1.5 fold) values were observed compared to the control group receiving carbamazepine only, which were attributed to piperine-mediated inhibition of the CYP3A4 enzyme [73]. In epilepsy patients, with long-time carbamazepine monotherapy and steady-state carbamazepine plasma levels, small increases in Cmax (1.1 fold) and AUC (1.1) values were observed after administration of a single dose of 20 mg piperine combined with 500 mg carbamazepine [121]. In an animal study with rats receiving a single combined dose of carbamazepine with 3.5 or 35 mg piperine/kg bw, no significant changes in plasma levels (Cmax or AUC-values) of carbamazepine and the major carbamazepine metabolite, carbamazepine-10,11-epoxide, were seen compared to the control group receiving carbamazepine only. Rats concomitantly receiving the same carbamazepine and piperine doses for 14 days showed significantly decreased plasma levels of carbamazepine (plasma Cmax 25–39% reduced; plasma AUC 29–37% reduced). The plasma levels of carbamazepine-10,11-epoxide and brain levels of carbamazepine were also reduced in both piperine-treated groups, but reached statistical significance only in the high-dose group. Carbamazepine-10,11-epoxide brain levels were also reduced but reached no statistical significance. A decreased plasma concentration of carbamazepine was observed in the high-dose piperine

group, despite decreased CYP3A2 protein expression in rat liver. The reduced carbamazepine drug and metabolite levels were mainly attributed to a reduced carbamazepine absorption (animals displayed increased defecation and wet faeces with piperine) and a decreased brain penetration of carbamazepine caused by piperine [148].

In addition to the abovementioned study conditions, it can be assumed that the concomitant administration of other bioactive substances together with piperine might also influence the effects of piperine on the bioavailability of the investigated drugs.

Concerning the improved bioavailability seen with several drugs, different underlying mechanisms have been discussed, such as improved drug absorption and/or inhibition of degradation or elimination. In this context, depending on the drug or investigated substance, different mechanisms for influencing absorption and different molecular targets relating to the metabolism of xenobiotics are at the center of discussion, viz. the unspecific gastrointestinal effects (increased splanchnic blood flow), altered membrane dynamics, inhibition of cytochrome P-450 enzymes (CYP3A4, CYP2E1, CYP2C9, etc.), inhibition of multispecific efflux transporters, such as P-glycoprotein, or other mechanisms [1,15,73–78,120,124,150].

It is noted that for the reduced bioavailability of diltiazem observed in an animal study, induction of P-glycoprotein has been considered as an underlying principle [147].

In summary, single or short-term administration of piperine bolus doses combined with several chemically and pharmacologically diverse drugs resulted in interactions, which in most cases led to increased drug bioavailability. However, it can be assumed that such interactions with piperine may also occur with other drugs that have not be tested in this respect so far. Such interactions may vary over time and depend on the used drug, drug dosage and piperine dosage, and may also possibly vary depending on concomitant administration of further bioactive substances. Appropriate investigations in humans are required with the drugs in question to further clarify this issue. Interactions (increased bioavailability of drugs) in humans were observed with several drugs at bolus doses of 20 mg piperine/day. For one drug (midazolam), increased clinical efficacy was reported with bolus administration of 15 mg piperine/day. The findings with β-carotene and coenzyme Q10 indicate that already bolus doses of 5 mg piperine/day may possibly cause interactions with certain substances or drugs.

#### **5. Discussion**

This review focuses on the evaluation of the safety of isolated piperine used as a single ingredient in food supplements, i.e., in bolus form, in adult individuals. Children and adolescents were not included in the consideration since adult persons constitute the prime target population for food supplements containing piperine.

Piperine is a natural ingredient of *Piper nigrum* (black pepper) and some other *Piper* species, e.g., *Piper longum*. Due to the use of peppercorns as a spice, it is a common component of the human diet. Apart from this source, piperine may be used in isolated form as a flavouring agent in food production.

When pepper is used for food seasoning purposes, piperine is added to food in combination with all other components of the peppercorns (which may influence observed biological effects) and in varying degrees of comminution (which may influence its bioavailability). In this context, it is usually consumed together with large quantities of food and in several portions throughout the day. By contrast, when piperine is used in food supplements, the piperine supply differs greatly in that piperine is ingested in isolated or highly concentrated form and as a bolus (usually in 1–3 portions per day), without any substantial amounts of other pepper components. These differences may influence the bioavailability and biological effects of piperine ingested as a bolus and in isolated form compared to its usual intake via food seasoning.

Currently available human studies with oral piperine administrations provide no adequate scientific basis for the final assessment of the potential health risks associated with intakes of isolated piperine used as a single ingredient and ingested in bolus form. This is a situation also frequently found with other substances used as ingredients of

dietary supplements. Although hardly any adverse effects (apart from potentially undesirable interactions of piperine with various drugs) were reported in available human studies, it is noted that in most cases safety issues were only marginally addressed or reported. In addition, is to be considered that in many studies piperine was administered in combination with other substances and that the combined administration complicates data interpretation.

However, some currently available human and animal studies already provided indications of potential health risks of bolus doses of isolated piperine.

In human and animal studies with single or short-time application, piperine was shown to interact with various drugs. In most cases, these interactions led to an improved bioavailability of the investigated drugs and are in line with the purported "bio-enhancing" activity of piperine. It can be assumed that such interactions may also occur with other drugs, and can vary, depending on the individual drug, drug dosage, piperine dosage, duration of intake and time span between piperine and drug intake. Additionally, interactions may also possibly vary depending on the concomitant administration of further bioactive substances. For clarification, appropriate investigations in humans are required with the drugs in questions, involving in particular studies with repeated drug and piperine application to reach steady-state levels.

Improved bioavailability can offer advantages in drug therapy if this is performed under medical supervision. Without adequate medical supervision, however, depending on the drug, piperine-based drug interactions may carry the risk of unintended and/or deleteriously increased medicinal drug effects or of the occurrence of adverse drug effects, especially in the case of drugs with a narrow therapeutic range. Increased bioavailability of certain drugs has been observed in several human studies with bolus doses of 20 mg/day (= LOEL), and findings with β-carotene and coenzyme Q10 suggest that in some cases bolus doses of 5 mg piperine/day might also cause such interactions. Taken together, against the background of these data, it seems advisable that individuals taking medicinal products, especially drugs with known piperine interactions or drugs for which no interaction data are available, should consult a physician prior to the use of isolated piperine as a food supplement.

Based on animal data, further potential health risks or potential risk groups, respectively, of bolus doses of isolated piperine can be identified. In four animal studies with juvenile and young adult rats, largely consistent paternal reproductive toxic effects (see Table 1) were observed at piperine bolus doses of 10 mg/kg bw/day [16,58–64].

In young adult rats, male reproductive toxic effects, which were significantly weaker and only partly statistically significant, were observed by Malini et al., already at bolus doses of 5 mg/kg bw/day [58,59]. On the other hand, a NOAEL of 1 mg piperin/kg bw/day (as bolus) can be identified from the study by D'Cruz and co-workers, who used a large spacing between the tested piperine doses (factor 10) [60,61]. Consistently clearer adverse paternal reproductive effects, i.e., disturbed spermatogenesis and accompanying adverse male reproductive effects, however, were observed with doses of 10 mg/kg bw/day.

It is to be acknowledged that these studies are afflicted with certain limitations; however, taken as a whole, the aggregated study findings point in the same direction and paternal toxicological reproductive effects, i.e., disturbed spermatogenesis, are corroborated by findings at different levels (i.e., histopathology, sperm parameters, hormonal changes and other parameters).

The reason for the different findings in terms of paternal reproductive toxic effects of these four studies compared to a sub-chronic 90-day animal toxicity study [29] remains elusive. The major difference seems to lie in the method of piperine administration: in the sub-chronic 90-day animal study, piperine was administered via feed, resulting in multiple intakes of small piperine quantities spread throughout the day. In the case of the other four abovementioned animal studies, piperine administration as a bolus can be assumed, possibly resulting in higher peak blood or tissue levels or otherwise increased bioavailability.

Regarding the reversibility of the disturbed spermatogenesis observed by Chinta and colleagues (see above) [16,62], it remains to be investigated to what extent such reversibility may still be present with piperine administrations lasting longer than 60 days. In addition, it remains to be clarified to what extent these findings may be extrapolated to humans. Even taking into account these findings of Chinta and co-workers with respect to the potential reversibility of the perturbation of spermatogenesis under certain conditions, the consistent paternal reproductive effects observed with bolus doses of 10 mg/kg bw/day should be classified as being toxicologically relevant and of potential concern.

Human studies that included adequate investigations to clarify the possible effects of bolus doses of isolated piperine on the male reproductive system could not be identified in the course of preparation of the current review. Therefore, as long as this knowledge gap has not been resolved and against the background of the adverse male reproductive effects observed in animal studies with high bolus intakes of isolated piperine, it seems advisable to maintain an adequate margin of exposure between those bolus doses of isolated piperine for which adverse male reproductive effects in animal studies were reported and the daily amounts of isolated piperine used in food supplements.

By using a dose of 10 mg piperine/kg bw/day for which adverse male reproductive effects were reported in several animal studies as a point of departure, the application of an uncertainty factor of 3 seems warranted for extrapolation to a NOAEL from this intake level. In addition, for deriving health-based guidance values from animal data, EFSA recommends to use an overall default assessment factor of 100 to account for inter-species and intra-human variability (10 for inter-species variability x 10 for inter-human variability) in the absence of chemical-specific data on the kinetics and/or dynamics [151]. Based on an assumed body weight of 70 kg, this approach would lead to a health-based guidance value of 2.3 mg/day (10 mg/kg bw/day × 70 kg bw/(3 × 100)) for piperine when used in isolated form via bolus administration, i.e., as a food supplement. This intake seems to be low compared to estimates of daily piperine intakes resulting from the usual culinary use of pepper in normal human diet. However, it should be taken into account that the intake of piperine in isolated form may not be directly comparable to its intake in conjunction with all other pepper ingredients and various degrees of comminution of the peppercorn. On the other hand, the effect of pepper consumption (in particular of high bolus doses with a high degree of comminution) on human male reproductive capacity remains to be clarified.

The health-based guidance value of 2.3 mg/day, as calculated above, which would currently be expected to provide an adequate level of protection with respect to potential male reproductive toxicity, would also be below the piperine bolus doses for which interactions with concomitantly administered drugs/substances have been observed.

In animal studies, maternal reproductive toxicity and embryotoxic effects were observed with piperine bolus doses of 10–25 mg/kg bw/day, which varied, depending on the time of piperine administration with regard to the day of mating, the duration of time covered during gestation and the piperine dose (10 mg/kg bw/day: reduced fertility index, reduced implantation rates; 25 mg kg bw/day: reduced implantation rates, abortive effects, delayed labour and increased foetal mortality) [54,68]. It should be noted that the observed adverse effects occurred in these studies at the lowest piperine bolus doses tested and that certain adverse effects were only investigated at 25 mg/kg bw/day. From the available animal studies, a LOAEL of 10 mg/kg bw/day, but no NOAEL regarding maternal reproductive toxicity and embryotoxic effects could be identified. It remains to be clarified whether the use of DMSO in the oral piperine application by Piyachaturawat et al. (1982) might have had an influence on the bioavailability or observed effects of piperine. However, adverse maternal reproductive and embryotoxic effects were also observed in a second study [54].

With regard to the potential health risks for which there are indications when using isolated piperine in bolus doses (drug interactions, disturbed spermatogenesis, adverse maternal reproductive and embryotoxic effects), it remains to be clarified whether these risks may be influenced by other bio-active, additional ingredients in food supplements

ingested together with piperine. In principal, the same considerations apply to highly piperine-enriched pepper extracts, although the difference between some highly piperineenriched extracts and the piperine preparations supplied from chemical companies (purity: ≥ 97%), used for instance in the cited studies of Chinta et al. [16,62] or D'Cruz et al. [60,61], does not seem to be substantial. However, in cases involving either other bio-active ingredients and/or highly piperine-enriched pepper extracts, any claimed mitigation or elimination of the adverse effects described above should be supported by adequate scientific investigations and data.

In conclusion, human and animal studies with single or short-term application of isolated piperine used in bolus form revealed interactions of the substance with several drugs, which can give rise to potential health risks, and based on which individuals taking medications can be identified as a potential risk group. For individuals taking medicinal products (especially drugs with known piperine interactions or drugs for which no interaction data are available), it seems advisable to consult a physician prior to the use of isolated piperine as a food supplement.

Animal studies with higher daily piperine bolus doses provide indications for further potential health risks (disturbed spermatogenesis; adverse maternal reproductive and embryotoxic effects), for which no adequate human data are currently available.

Considering that a distinct NOAEL for maternal reproductive and embryotoxic effects could currently not be identified from the available animal studies, it seems advisable for pregnant women to abstain from the use of food supplements containing isolated piperine. Moreover, the reduced implantation rates that were reported in animal studies may also be of relevance for women who wish to become pregnant.

Regarding the observed adverse paternal reproduction effects in animal studies and the lack of information on the effects of bolus doses of isolated piperine on the human male reproductive system, it appears prudent to maintain an adequate margin of exposure between those bolus doses that produced adverse paternal reproductive effects in animal studies and the maximum daily amounts of isolated piperine in food supplements.

Considering the uncertainties outlined above, the importance of addressing the existing knowledge gaps regarding effects of bolus doses of isolated piperine on human male reproductive capacity in future human intervention studies, by including specific investigations into this endpoint in the study design, is emphasised.

**Author Contributions:** Conceptualization, R.Z.; writing—original draft preparation, R.Z.; writing review & editing, R.Z., K.H., A.L. and K.I.H.-E. All authors have read and agreed the published version of the manuscript.

**Funding:** This research received no external funding.

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** Not applicable.

**Acknowledgments:** The authors would like to thank the Dietary Exposure and Aggregated Exposure Unit of the German Federal Institute for Risk Assessment's (BfR), in particular Christine Sommerfeld, for conducting the estimation of the daily pepper intake of the German population (14–80 years) and Britta Nagl and Carolin Schopf for technical assistance in the preparation of the manuscript.

**Conflicts of Interest:** The authors declare no conflict of interest.
