Technology Readiness Level Roadmap for Developing Innovative Herbal Medicinal Products

Abstract

Despite the vast global botanical diversity, the pharmaceutical development of herbal medicinal products (HMPs) remains underexploited. Of over 370,000 described plant species, only a few hundred are utilized in HMPs. Most of these have originated from traditional use, and only a minority come from megadiverse countries. Exploiting the pharmacological synergies of the hundreds of compounds found in poorly studied plant species may unlock new therapeutic possibilities, enhance megadiverse countries’ scientific and socio-economic development, and help conserve biodiversity. However, extensive constraints in the development process of HMPs pose significant barriers to transforming this unsatisfactory socio-economic landscape. This paper proposes a roadmap to overcome these challenges, based on the technology readiness levels (TRLs) introduced by NASA to assess the maturity of technologies. It aims to assist research entities, manufacturers, and funding agencies from megadiverse countries in the discovery, development, and global market authorization of innovative HMPs that comply with regulatory standards from ANVISA, EMA, and FDA, as well as WHO and ICH guidelines.

Full text

Citation: Pagani, E.; Ropke, C.D.;
Soares, C.M.; Perez, S.A.C.; Benevides,
P.J.C.; Barbosa, B.S.; Carvalho, A.C.B.;
Behrens, M.D. Technology Readiness
Level Roadmap for Developing
Innovative Herbal Medicinal Products.
Pharmaceuticals 2024,17, 703. https://
doi.org/10.3390/ph17060703
Academic Editor: Ilkay Erdogan
Orhan
Received: 19 April 2024
Revised: 15 May 2024
Accepted: 17 May 2024
Published: 29 May 2024
Copyright: © 2024 by the authors.
Licensee MDPI, Basel, Switzerland.
This article is an open access article
distributed under the terms and
conditions of the Creative Commons
Attribution (CC BY) license (https://
creativecommons.org/licenses/by/
4.0/).
pharmaceuticals
Article
Technology Readiness Level Roadmap for Developing
Innovative Herbal Medicinal Products
Eduardo Pagani 1, 2, * , Cristina Dislich Ropke 2, Cristiane Mota Soares 3, Sandra Aurora Chavez Perez 3,
Paulo JoséCoelho Benevides 4,
Barbara Sena Barbosa
2
, Ana Cecilia Bezerra Carvalho
5
and Maria Dutra Behrens
6,
*
1Medical Department, Azidus Brasil, Valinhos 13271-130, SP, Brazil
2Centroflora Group, Innovation Department, Campinas 06460-040, SP, Brazil
3Project Management Office, Vice Direction of Education, Research and Innovation, Institute of Drug
Technology Farmanguinhos, Oswaldo Cruz Foundation, Rio de Janeiro 21041-250, RJ, Brazil
4Business Office, Oswaldo Cruz Foundation, Campo Grande 79081-746, MS, Brazil
5GMESP, Brazilian Health Regulatory Agency, Agência Nacional de Vigilância Sanitária (ANVISA),
Brasília 71205-050, DF, Brazil
6Natural Products Department, Vice Direction of Education, Research and Innovation, Institute of Drug
Technology Farmanguinhos, Oswaldo Cruz Foundation, Rio de Janeiro 21041-250, RJ, Brazil
*Correspondence: eduardo.pagani@azdcro.com.br (E.P.); maria.behrens@fiocruz.br (M.D.B.)
Abstract: Despite the vast global botanical diversity, the pharmaceutical development of herbal
medicinal products (HMPs) remains underexploited. Of over 370,000 described plant species, only
a few hundred are utilized in HMPs. Most of these have originated from traditional use, and
only a minority come from megadiverse countries. Exploiting the pharmacological synergies of
the hundreds of compounds found in poorly studied plant species may unlock new therapeutic
possibilities, enhance megadiverse countries’ scientific and socio-economic development, and help
conserve biodiversity. However, extensive constraints in the development process of HMPs pose
significant barriers to transforming this unsatisfactory socio-economic landscape. This paper proposes
a roadmap to overcome these challenges, based on the technology readiness levels (TRLs) introduced
by NASA to assess the maturity of technologies. It aims to assist research entities, manufacturers,
and funding agencies from megadiverse countries in the discovery, development, and global market
authorization of innovative HMPs that comply with regulatory standards from ANVISA, EMA, and
FDA, as well as WHO and ICH guidelines.
Keywords: phytotherapy; technology readiness level; herbal substance; herbal preparation; herbal
medicinal products; drug discovery; drug development
1. Introduction
Natural products (NPs) have historically been the primary source of lead compounds
in drug discovery [
1
]. Living organisms have developed metabolites for hundreds of
millions of years to improve their survival capacity, thus creating chemical diversity [
2
].
NP collections have increased chemical and steric complexity (e.g., more heterocycles and
chiral centers) and display better drug-like properties than their synthetic counterparts,
making them more useful for early-stage screenings [
3
–
5
]. More than 50% of all marketed
therapeutic small molecules are either NPs, NP derivatives, or synthetic molecules whose
pharmacophoric center was inspired by NPs, including some with traditional use [6,7].
Herbal preparations (HP) comprise hundreds of phytochemicals [
8
,
9
] that could
address medical needs through multitarget mechanisms [
10
–
13
]. The possible synergy of
multiple compounds, within a phytocomplex proposed decades ago, has been substantiated
by advancements in systems biology and “-omics” [14].
Herbal medicinal products (HMPs) are a niche of NPs [
15
]. It is estimated that up
to 80% of the world’s population relies on herbal medicines and medicinal plants as a
Pharmaceuticals 2024,17, 703. https://doi.org/10.3390/ph17060703 https://www.mdpi.com/journal/pharmaceuticals

Pharmaceuticals 2024,17, 703 2 of 23
primary source of healthcare [
16
], which can be beneficial in several non-communicable
diseases [
17
]. The global herbal market was USD 60 billion in 2010 and is expected to reach
USD 5 trillion by 2050 [
18
]. If properly managed, this market growth can generate income
and convert local populations into conservation agents [19].
The world has currently 377,749 accepted plant species, mostly occurring in megadi-
verse countries [
20
]. Only 15% have been characterized phytochemically and 6% pharma-
cologically [
21
,
22
]. The number of species used in HMPs registered by the Food and Drug
Administration (FDA), European Medicines Agency (EMA), and the Brazilian Health Reg-
ulatory Agency (ANVISA) is just a few hundred, and most like Ginkgo biloba,Panax ginseng,
and Echinacea purpurea have been known for centuries. This occurs despite regulations
that have existed for decades in Brazil, the EU, and the USA permitting the registration of
innovative HMPs.
Among the 17 megadiverse countries, only two, namely the USA and China, have a
consistent track record of pharmaceutical innovation. Encouraging the development of
innovative HMPs originating from other countries could potentially stimulate their scien-
tific and socio-economic progress, safeguard biodiversity, and introduce novel therapeutic
options for medical needs [23–25].
The absence of innovation may partly stem from legal challenges. Sanitary regulations
differ across countries [
26
,
27
], and there exist legal uncertainties in the operationalization
of the Nagoya Protocol [
28
,
29
]. Brazil has addressed both issues. ANVISA has issued
a regulatory framework for HMPs aligned with EMA [
30
]. The Brazilian Biodiversity
Law [
31
] aligns with the Nagoya Protocol, offering legal certainty for the assessment of
genetic resources and traditional knowledge, and benefit sharing [
28
]. Also, attempts by
the European Community (EC) and the World Health Organization (WHO) provide models
for multinational harmonization [32].
However, despite these advancements, innovations from Brazil and other biodiverse
countries remain scarce, indicating that a significant factor may still be missing. The devel-
opment of HMPs is a lengthy and intricate process that relies on the effective management
of various capabilities and long-term funding. Furthermore, adaptations to previously
planned tasks are often necessary. Given these challenges, we believe that a comprehensive
yet adaptable roadmap outlining this process could assist research institutions, pharma-
ceutical companies, and funding bodies in establishing consistent, long-term partnerships
aimed at fostering innovation.
NASA proposed the technology readiness levels (TRLs) decades ago to guide man-
agers in assessing the aeronautical technology’s readiness and risks at specific development
key points [
33
,
34
]. The concept has spread over many innovation fields and is currently
used in pharmacology by development-fostering agencies [
35
–
39
]. TRLs fit very well in
the proposal of a roadmap providing hierarchized development concepts and enabling a
step-by-step approach.
Almost no peer-reviewed publication specifically covering TRLs in pharmaceutical
development is available, either for small molecules, HMPs, or biotechnological products.
Moreover, most published TRL scales have general concepts and examples, but lack descrip-
tors defining boundaries, leaving room for inconsistent subjective choices that might vary
among projects. Such a problem was detected and addressed in the chemical industry [
40
].
To face these challenges, we propose a TRL-guided HMP development roadmap
applicable to Brazilian innovations aiming at worldwide marketing. It is organized in a
step-by-step set of actions grouped into six domains, defined by clear boundaries, and
compliant with international guidelines.
This proposal applies to HMPs, based on herbal preparations (HPs) as active phar-
maceutical ingredients (API), and not to isolated or purified substances. It also applies to
plants not included in official monographs, regardless of previous traditional use. For the
development of HMPs based on traditional or well-established use, we refer the reader to
the following references [
41
–
47
]. Moreover, for the innovations addressed here, previous
ethnopharmacological information is helpful, but not mandatory.

Pharmaceuticals 2024,17, 703 3 of 23
2. Results and Discussion
Following the original Mankins style [
33
,
48
,
49
], we discuss each technology readiness
level (TRL) main attribute, citing examples and providing brief descriptions of associated
costs and funding sources. Table 1outlines the primary deliverables for each level defined
by Mankins, alongside our proposed adaptations for herbal medicinal products (HMPs).
Additionally, Table 1provides a visual summary of the key concepts and tasks associated
with each TRL as applied to the development of HMPs.
Table 1. Technology readiness level roadmap according to Mankins [
23
,
34
,
40
] and its adaptation to
herbal medicinal product development (HMP).
Mankins Basic principles observed and reported
TRL-1 HMP Pharmacological effect of an herbal preparation observed and reported
Mankins Technology concept and/or application formulated
TRL-2 HMP Technological application formulated and proof of concept planned
Mankins Analytical and experimental critical function and/or characteristic proof of concept
TRL-3 HMP Proof of concept of herbal preparation (API candidate)
Mankins Component and/or breadboard validation in a laboratory environment
TRL-4 HMP Optimization of herbal preparation in bench-scale and non-GLP validation
Mankins Component and/or breadboard validation in a relevant environment
TRL-5 HMP Optimization of herbal preparation in pilot-scale and GLP validation
Mankins System/subsystem model or prototype demonstration in a relevant environment (ground or space)
TRL-6 HMP Herbal medicinal product prototype completes phase I clinical trial
Mankins System prototype demonstration in a space environment
TRL-7 HMP Herbal medicinal product completes phase II clinical trial
Mankins Actual system completed and “flight qualified” through test and demonstration (ground or space)
TRL-8 HMP Herbal medicinal product completes phase III clinical trial, and the dossier is approved by one
regulatory authority
Mankins Actual system “flight-proven” through successful mission operations
TRL-9 HMP Herbal medicinal product pharmacovigilance and phase IV clinical trials are performed
2.1. TRLs for Herbal Medicinal Products
2.1.1. TRL-1
TRL-1 was described as “basic science envisioning an application” [
33
,
49
–
51
]. For
chemicals, it was summarized as a “concept” [
40
]. BARDA [
38
] describes it as “active
monitoring of scientific knowledge database”.
For HMPs, the starting point is the identification and reporting of an herbal substance
(HS) or herbal preparation’s (HP) pharmacologically relevant effect. This comes either
from an ethnopharmacological observation or laboratory experimentation. Both demand
envisioning a connection between an effect and a medical need [
35
]. Examples of these
two independent possibilities are: 1- an ethnopharmacological description of a plant
infusion with psychic effects, and 2- an experimental finding of vasodilating properties
of a plant extract. The associated costs typically involve academic research expenditures,
encompassing both experimental work and field-based ethnopharmacological surveys.
Funding for this stage is commonly provided by basic science funding agencies.
2.1.2. TRL-2
TRL-2 “formulates the technology concept or application, with no experimental proof
or detailed analysis supporting the conjecture” [
33
,
49
,
52
]. The R&D fostering agencies
agree on “developing hypotheses and experimental designs” [
35
–
38
]. Therefore, TRL-2
creates a project focused on a product.
For HMPs, we propose that TRL-2 devises a technological application and delivers a
proof-of-concept (PoC) research project based on the literature and experimental data. It
starts addressing issues on the HS and/or HP and chooses at least one experimental model
with translational capability. The literature review must be comprehensive and preferably
include some data produced by the responsible research group. This difference from the

Pharmaceuticals 2024,17, 703 4 of 23
original NASA concept is due to the higher dependence of pharmaceutical development
on previous experimental data to design a reliable PoC.
The progression of the two examples mentioned at TRL-1 involves the formulation of
research projects aimed at testing the antidepressant potential of the plant with psychologi-
cal effects or the antihypertensive potential of the extract with vasodilating properties. The
associated costs typically fall within the scope of small- or medium-sized academic projects
sponsored by basic-science funding agencies.
2.1.3. TRL-3
TRL-3 delivers an “analytical and experimental critical function and/or characteristic
proof of concept” [
33
,
49
,
52
]. The R&D fostering agencies agree on the “proof of concept”,
and BARDA adds identifying a target or candidate and demonstrating relevant
in vitro
activity [35–38].
For HMPs, we propose that TRL-3 performs the R&D fostering agencies (PoC) on a
bench scale according to the research plan described in TLR-2. The results must justify the
progression to TRL-4 and may demand more than one assay. The first example progresses
to the performance of an
in vitro
test for serotonin reuptake and/or an animal model for
depression [
53
]. The second could be an
in vitro
vasodilation test and/or an animal model
for systemic arterial hypertension [
54
]. As mentioned above, the associated costs typically
fall within the scope of small- or medium-sized academic projects sponsored by basic
science funding agencies.
2.1.4. TRL-4
TRL-4 goes further in the validation. It must support the concept formulated earlier
and be consistent with the requirements of potential system applications [
33
,
49
,
52
]. Moor-
house also adds “credible for design and performance conditions” [
55
]. The R&D fostering
agencies focus on testing in a defined laboratory or animal model, and BARDA specifically
mentions “optimization” [35–38].
For HMPs, we propose HP optimization on a bench scale and broad validation through
several independent assays. TRL-4 is the most laborious and complex stage up to this
point and may involve the work of different groups for years. It holds the first major
risk inflection point by testing optimized HPs in the most translational disease models.
Here, both examples converge to a chemically defined HP with characterization and
quantification of constituents with therapeutic activity or marker compound(s) and several
in vitro
and
in vivo
tests. The costs and timeliness rise substantially. The challenges
might be faced by academic centers alone, or in collaboration with manufacturers and/or
contract research organizations (CROs). The costs might be public-sponsored by large-
sized academic research projects and may start involving private resources from investors
and manufacturers.
2.1.5. TRL-5
TLR-5 performs “component or prototype validation in a development-relevant envi-
ronment” [33,34,49,50,52,55].
For HMPs, we define “component” as a candidate HP produced through optimized
and standardized processes. “Validation in a development-relevant environment” is
equated with conducting “Good Laboratory Practice (GLP) non-clinical studies” [
35
–
38
].
GLP testing commences only when the phytochemical profile of the HP candidate is well
defined, achieved through standardized preparation procedures and bio-guided studies.
Hence, TRL-5 encompasses two primary tasks: scaling up from the bench to the
pilot scale and conducting GLP animal testing. These activities are usually carried out in
specialized facilities, including CROs. Costs associated with TRL-5 are generally within the
same range as those of TRL-4. While some public funding sources may still be available,
the reliance on investors and manufacturers tends to increase.

Pharmaceuticals 2024,17, 703 5 of 23
2.1.6. TRL-6
TRL-6 refers to a “system/subsystem model or prototype demonstration in a relevant
environment (ground or space)” [33,34,49,50,52,55].
For both drugs and HMPs, the “relevant environment” corresponds to the “first-in-
human” testing in phase I clinical trials conducted under Good Clinical Practices (GCP),
typically conducted in a medical facility [
35
–
38
]. Costs escalate significantly due to the
manufacture of the candidate HMP for human use and GCP testing. While some public
funding sources may still be available, the reliance on investors, manufacturers, and banks
further increases.
2.1.7. TRL-7
TRL-7 is the “system prototype demonstration in the expected operational envi-
ronment” [
33
,
34
,
49
–
52
,
55
]. There is a clear parallel between the “expected operational
environment” and the phase II clinical trials conducted with patients with the condition
potentially treated by the HMP candidate [35–38].
This TRL holds the second major risk inflection point because, in the case of success,
the risks of failure in further stages are reduced. In case of failure, the project shall be
aborted or deeply revised. The costs keep rising. Because the technological risks are still
high, some public funding might be expected, along with a higher amount of private capital
from investors, manufacturers, and banks.
2.1.8. TRL-8
In the TLR-8, the “actual system is completed and qualified through test and demon-
stration” [
33
,
34
,
49
–
52
,
55
]. For pharmaceutical development, it means that an Investiga-
tional New Drug (IND) is reviewed and approved by the regulatory agency.
The parallel with phase III clinical trials is evident. This stage is designed to generate
statistically significant evidence of HMP efficacy and safety in large patient populations
with the target disease [
35
–
38
]. The HMP used is that intended for marketing authorization.
Costs associated with TRL-8 are the highest of the development process. TRL-8 alone can
incur expenses equivalent to those of TRLs 1 to 7combined. However, risks could be lower
compared to TRL-7. Costs are typically privately sponsored by manufacturers, along with
contributions from investors and banks.
2.1.9. TRL-9
TRL-9 is an “actual system flight-proven through successful mission
operations” [
33
,
34
,
49
–
52
,
55
]. This was translated by “post-marketing studies and surveil-
lance of the HMP after approval for use” [
35
–
38
]. By definition, all technologies that
succeed in being applied in actual systems go eventually to TRL-9.
In certain instances, new formulations may be developed with higher or lower concen-
trations, slow-release properties, or tailored for pediatric use, necessitating additional clini-
cal trials (phase IV or post-marketing). Despite the potential inclusion of phase IV clinical
trials, costs typically decrease substantially and are usually sponsored by
the manufacturer.
2.2. TRL Domains and Boundaries
Given the complexity of HMP development, we established six technological domains
whose progression facilitated the delineation of the boundaries for technology readiness
levels (TRLs): herbal substance, herbal preparation, herbal medicinal product, analytical
development, non-clinical assays, and clinical trials. Table 2summarizes the deliverables
and main regulations applicable for HMPs at each level.

Pharmaceuticals 2024,17, 703 6 of 23
Table 2. Main deliverables of each TRL for each HMP development domain with main regulations
applicable.
HS HP HMP AD NCA CT
TLR1 Initial literature review and data collection
TLR-2
Addressing
botanical
inconsistencies
Comprehensive
literature review
Compatibility
with vehicles
Initial
parameters’
proposal
Comprehensive
literature
review
Comprehensive
literature
review
TLR-3
Certified
botanical
identification
Best extract under
current conditions
Advance
solubility and
compatibility
issues
Best characteri-
zation under
current
conditions
Optimal PoC
under current
conditions
TLR-4
Full traceability
supply-chain
development
Bench-scale
optimization and
semi-industrial
scale initiation
Advances
formulation for
pre-clinicals
Consistent char-
acterization
Efficacy and
safety
confirmation
and bio-guided
HP
optimization
Update
literature
review
TLR-5 Supply chain
consolidation
Semi-industrial
scale optimization
Advance
formulation for
human use
Validated
methods for HP
GLP tests for
phase I CT
authorization
Update
literature
review
TLR-6 Stability
assessment
GMP-compliant
batches
GMP-
compliant
batches
Validated
methods for HP
and HMP
GLP tests for
phase II CT
authorization
GCP phase I
clinical trial
TLR-7
Industrial scale
and GACP im-
plementation
GMP-compliant
industrial batches
GMP-
compliant
batches
Validated
methods for HP
and HMP
GLP tests for
long-term use
GCP phase II
clinical trials
TLR-8 GACP
compliance
GMP-compliant
industrial batches
GMP-certified
batches
Validated
methods for HP
and HMP
GLP tests for
long-term use
GCP phase III
clinical trials
TLR-9 GACP certified
GMP-compliant *
or certified
industrial batches
GMP-certified
batches
Validated
methods for HP
and HMP
GCP phase IV
trials and
pharmaco-
vigilance
HS: herbal substance; HP: herbal preparation; HMP: herbal medicinal product; AD: analytical development; NCA:
pre-clinical assays; CT: clinical trials; GACP: Good Agricultural and Collection Practices; GLPs: Good Laboratory
Practices; GCP: Good Clinical Practices; The Nagoya Protocol applies to the HSs and HPs in different forms in
different countries. * The demand for GMP-compliant or GMP-certified HP at TRL-8 varies in different countries.
2.2.1. Herbal Substance
HMPs’ TRLs 1 and 2rely on a comprehensive literature review, eventually supple-
mented by any original data provided by the responsible researcher. Older publications
can be valuable if they contain human data. However, these reports may suffer from miss-
ing, outdated, or unreliable botanical data, inconsistencies in subspecies, or phenotypical
variations. Researchers must diligently address these inconsistencies, striving to include all
potentially relevant information while highlighting sources of confusion and detailing how
these issues were resolved to ensure reliable botanical identification. Additionally, data on
species within the same genus may be useful in predicting the expected phytochemical and
biological profiles of the chosen plant [
56
]. TRL-2 also involves gathering information on
genetic resources and assessments related to traditional knowledge, in compliance with the
Nagoya Protocol [29].
For TRL-3 proof of concept (PoC), botanical identification must be conducted by a
certified botanist using a voucher sample, ideally including the flower, which should be de-
posited in an official herbarium. The collection process must be meticulously documented,
including GPS location, date, time, and weather conditions (preferably dry) at the time

Pharmaceuticals 2024,17, 703 7 of 23
of collection, as well as phenological information and the quantity collected. Records of
conservation conditions (light and humidity) are of utmost importance. The quantity stored
must be sufficient to facilitate counter-proof tests until the completion of TRL-4. Henceforth,
the species should not change, except for official naming updates. If there is a change in
the species, an impact analysis must be conducted to determine which phytochemical and
biological tests need to be repeated.
TRL-4 needs HS full traceability, following the procedures outlined for TRL-3. To
optimize conditions, gathering various batches of HSs from different locations and during
different seasons is essential. This process is time-consuming and progresses from TRLs
4to 7, making an early start crucial. Each batch must be of sufficient quantity and stored
appropriately to facilitate repeated assessments for counter-proof tests until the conclusion
of TRL-7.
TRL-4 also involves addressing the HS supply chain. Sustainable collection practices
are viable if the plant is abundant in the wild and collection methods do not harm the
environment. This approach may even yield environmental benefits by engaging rural
populations as conservation agents [
57
]. Agroecological approaches and organic agriculture
offer viable options as well. Strains can be selected, domesticated, and cultivated on a
small scale to initiate the development of a cultivar. Plant strains exhibiting the desired
metabolite profile and biological activity can be chosen for pilot-scale cultivation, typically
encompassing less than one hectare [58,59].
TRL-5 achieves consistency in production type (collection or cultivation), selects the
optimal strain, collection time, and implements effective conservation practices which are
paramount for scalability. This stage also initiates the development of Good Agricultural
and Collection Practices (GACP) and the qualification of suppliers. Rigorous control of
production’s initial stages is imperative, encompassing GACP and Good Herbal Processing
Practices (GHPP) [
60
,
61
]. From TRL-5 onwards, the HS must undergo quality controls to
ensure plant identification, chemical profiling, and detection of impurities (including other
materials and plants). Any deviations in the supply chain or composition need an impact
analysis on the product. Actions to increase the production scale commence at this stage,
continuing through TRL-7 or 8. Initial non-clinical GLP tests can be conducted with pilot
batches, while long-term pivotal tests and clinical trials are preferably performed using
industrial batches.
At TRL-6, the focus shifts to consolidating the supply chain and finalizing composition
tests. By TRL-7 the HS production scales up to industrial batches, ranging from hundreds of
kilograms to tons, depending on the equipment used. A qualification master plan becomes
necessary for phase II clinical trials [
62
,
63
], and Good Agricultural and Collection Practices
(GACP) must be implemented to provide HS batches for use in phase II and III clinical
trials [61].
At TRL-8, the raw materials utilized must be those intended for HMP registration.
Proof of compliance with GACP guidelines is essential. While formal GACP certification is
not mandatory in Brazil, it is required in the European Community to simplify assessment
and provide assurance to regulatory bodies [
61
]. The FDA mandates compliance with the
full GACP standard for an IND PH3 Investigational New Drug, as outlined in 21 CFR
321.23. Certification is only necessary for the New Drug Application phase [64].
2.2.2. Herbal Preparation
TRL-1 ideally provides some information on the extractive methods and the extract’s
composition. At TRL-2, a thorough literature review should encompass all available
information from scientific and patent databases concerning methodological variations and
hypotheses regarding active, toxic, and inert components. Initiating a bio-guided study
aimed at isolating and identifying or characterizing constituents with therapeutic activity
or markers is advisable at this stage. If conducted, high-throughput and high-content
screenings (HTS, HCS) of extracts or fractions commence here and continue throughout
TRL-3 [22,65].

Pharmaceuticals 2024,17, 703 8 of 23
At TRL-3, a PoC must use the best possible extract, considering the literature review,
the investigators’ capabilities, and the budget available. The quantity produced shall be
sufficient for repetitions and comparisons until the conclusion of TRL-4. The extractive
method must be thoughtfully described and justified. Preferably, the solvents should be
compatible with a future industrial scale-up; otherwise, some tests may need to be repeated.
HP candidates must undergo chemical characterization to the fullest extent possible with
the available resources. The bio-guided study should identify constituents with known
therapeutic activity, markers, or at least the active phytochemical classes.
At TRL-4, the focus is on optimizing extractive conditions at a bench scale to maximize
active components while minimizing inert or potentially toxic ones [
66
,
67
]. This includes
testing various equipment, solvents, techniques, and drying methods [
68
–
70
]. The extracts
undergo evaluation through a series of biological assays to confirm their pharmacological
activity [
69
,
71
]. TRL-4 is considered complete only when the chemical profile of the herbal
preparation (HP) candidate is optimized at the bench scale through bio-guided studies.
Additionally, attention is given to HP stability. Adequate samples must be stored for
repeated testing until TRL-7 or 8. The entire process must be thoroughly described, and
storage conditions must be traceable and auditable.
At TRL-5, the HP candidate production is scaled up to pilot or semi-industrial quan-
tities. The challenge lies in maintaining or improving the characteristics selected in the
previous TRLs. The pilot process must be robust, reproducible, and capable of delivering a
stable product. This lays the foundation for implementing Good Manufacturing Practices
(GMP) for the HP, which are required for regulatory-compliant Good Laboratory Practice
(GLP) preclinical studies. Typically, this process is carried out by companies specialized
in HP manufacture and should adhere to GMP, or in some countries, be GMP-certified.
Development of the extractive method is finalized with at least three pilot batches, each
comprising at least five kilograms of dry extract, accompanied by the respective analytical
reports [16,47,72].
The industrial batches are developed during TRL-6 and TRL-7. The phases I and II
clinical trials can be run with either pilot or industrial batches manufactured in compliance
with GMP, with validation and certification processes initiated [
38
,
73
]. The TRL-8 phase III
clinical trial must be performed with GMP-certified industrial batches identical to those
intended for licensing [
74
]. Information on batch-to-batch variations and the demonstration
that the API (HP) is safe and effective within the tolerated variability is required [74].
Herbal Preparation Concepts in Europe and Brazil
In Europe and Brazil, extracts can be classified into three categories, according to
the European Pharmacopoeia 11th edition [
41
,
75
] and the Brazilian Pharmacopoeia 6th
edition [76]:
Standardized extracts (i), in which constituents with therapeutic activity are known.
The amount of native or genuine (without excipients) extract is variable, adjusted within the
range defined for the active component—the adjustment is carried out with inert excipients
or by blending production batches with a higher or lower content of active constituents,
resulting in a variable amount of native HP (e.g., Atropa belladonna). Quantified extracts (ii),
which are adjusted within a defined range of compounds (active markers), whose relation to
the activity is proved through clinical trials, and, therefore, generally accepted to contribute
to the therapeutic activity. The amount of native (genuine) extract is constant and the
adjustment can only be achieved by blending extract batches with the same specification
and based on a constant amount of native extract (e.g., Ginkgo biloba). Other extracts
(iii) are mainly defined by their manufacturing process (state of the drug to be extracted,
solvent, extraction conditions) as well as their specification. There is no adjustment for a
constituent or group of constituents since the active substance is the native extract, whose
amount is constant. The constituents with known therapeutic activity or markers are not
known. Therefore, analytical markers are indicated, and their contents are batch-specific,

Pharmaceuticals 2024,17, 703 9 of 23
informative, or recommended to have a minimum content referred to (e.g., Valeriana
officinalis). In the case of traditional use, the extraction process should not change.
2.2.3. Herbal Medicinal Product
This domain encompasses administration route, formulation, intellectual property
(IP), and partnerships. TRL-2 and TRL-3 focus on HP solubility and compatibility with
vehicles for both
in vitro
and
in vivo
testing. In addition to academic sources, conducting a
preliminary review of patent databases is advisable to assess freedom to operate.
At TRL-4, the physical form and administration route of the HP are defined, and
formulation development commences after gathering information on API solubility and
stability [
77
]. Seeking professional advice on intellectual property is advisable to ensure
freedom to operate and explore opportunities for patenting formulations that enhance API
delivery and stability. For academic projects, this is an opportune time to explore potential
partnerships with manufacturers.
At TRL-5, GLP tests are conducted with a formulation as close as possible to that
intended for human use, considering animal testing peculiarities. Solid formulations for
oral use, such as pills and capsules, cannot be directly administered to animals. Instead,
GLP guidelines for oral toxicity studies permit administration via gavage, dissolved in
drinking water, or mixed with food [
78
]. Additionally, Brazilian guidelines allow oral tests
to be conducted with the HP [
79
]. Typically, partnerships with manufacturers are already
in operation, and intellectual property (IP) issues have been addressed.
TRL-6 phase I and TRL-7 phase II clinical trials must be performed with the HMP
manufactured in compliance with GMP and stability tests compatible with the clinical
trials’ duration. The investigational product manufacture happens in a GMP-like envi-
ronment but is not necessarily GMP-certified [
38
,
73
]. Still, there might be variations in
the dose/concentration of the product that goes to the market. If the claim is based on
traditional information, traditional doses and concentrations must be followed.
At TRL-8, phase III clinical trials are conducted with the GMP-certified formulation in-
tended for marketing authorization, ensuring stability throughout the study
duration [16,38,73].
If there were differences between the formulation used in phase II and phase III clinical
trials, the impact on safety and efficacy must be addressed and the modifications justified.
For dossier submission, stability tests must be finalized for at least three HMP batches in
their packaging. The Nagoya Protocol mandates notification of the final product’s genetic
resources and associated traditional knowledge assessment [29].
At TRL-9, the HMP is finalized. However, it is feasible to develop new formulations
for alternative routes, varying concentrations, slow-release, or pediatric use, which typically
require new clinical trials for marketing authorization.
2.2.4. Analytical Development
TRL-2 proposes parameters for HS and HP characterization. For known HSs, the best
sources are official pharmacopoeias or compendia [
45
,
46
,
62
,
76
,
80
–
83
]. Each country has its
list of official legally binding pharmacopoeias. For innovative HS, the pharmacopoeias and
scientific publications provide insights on standards to follow. HS characterization typically
includes descriptions of the plant part(s) used, purity, chemical content, contaminants,
etc. [
84
–
86
]. The HP candidate usually contains substances or chemical classes associated
with therapeutic or toxic effects, which can serve as positive and negative markers for HP
optimization and quality control. It is recommended to perform a bio-guided isolation of
bioactive compounds, especially when commercial standards are unavailable.
At TRL-3, analytical methods must be employed for HS and HP characterization by
techniques such as thin-layer chromatography (TLC), high-performance liquid chromatog-
raphy (HPLC), gas chromatography (GC), and mass spectrometry (MS). A fingerprint
chromatogram with all peaks and times, along with the identification of some components,
is recommended.

Pharmaceuticals 2024,17, 703 10 of 23
TRL-4 requires HPs’ comprehensive qualitative and quantitative characterization
by different methods produced from several HS batches. Bio-guided identification of
active compounds and markers progresses, alongside high-throughput screening (HTS)
or high-content screening (HCS), if applicable. The analysis should identify the major
chromatographic peaks. Similar objectives can be achieved through MS virtual libraries.
Hyphenated methods such as HPLC-PDA, UHPLC-PDA-MS, HS-SPME-GC-MS, or GC-
FID/MS are increasingly used [
87
–
89
]. Knowledge of positive and negative markers should
increase through bio-guided studies aimed at compound isolation, especially for species
lacking traditional use or literature data. All HPs tested thereafter must have analytical
reports in the CTD format. ANVISA and EMA have published guidelines adapted to
HMPs [90–92].
TRL-5 needs protocols for HS and HP quality control. The analytical methods must
be either pharmacopoeic or independently developed and validated addressing speci-
ficity/selectivity, linearity, working range, precision, accuracy, detection/quantification
limits, and robustness [
81
,
93
,
94
]. In Brazilian regulations, these methods can replace the
phytochemical assays for quality control, when technically justified [95].
From TRL-6 onwards, the requirements expand to include the evaluation of formula-
tions for human use, as well as stability testing. At this stage, active ingredients or markers
with known therapeutic effects are identified, and all analytical methods must adhere to
pharmacopoeial standards or be thoroughly validated. Additionally, a comprehensive
chromatographic profile (fingerprint) is established, alongside stringent analysis of po-
tential contaminants, including pesticides, heavy metals, adulterants, microorganisms,
and toxins. These analyses must conform to the regulatory requirements of the target
country for licensing, primarily following official pharmacopoeias. For TRL-8, stability
testing is conducted on products in their final packaging. TRL-9 uses the methodologies
established in previous stages, applying them to every industrial batch. New analytical
methods are developed or adapted as necessary to accommodate any new formulations
eventually introduced.
2.2.5. Non-Clinical Assays
The purpose and number of non-clinical assays for innovative HMPs depend on the
documented human exposure. In all cases, TRL-1 relies on some previous data. TRL-2
performs a comprehensive literature review, including all relevant
in vitro
and
in vivo
data on HSs, HPs, and HMPs from the same species. In some cases, data on other species
from the same genus may also be helpful. The review embraces decades of information in
pharmacopoeias, scientific journals, theses, books, websites, congress abstracts, and others.
If available, it is advantageous to include data produced by the development team.
The PoC experiment characterizes TRL-3. It includes
in vitro
assays of relevant targets
and/or
in vivo
disease models. Appropriated negative and positive controls are mandatory.
The assay(s) choice shall be justified, based on the potential for extrapolation to humans,
and described in detail, including allometric dose transposal. The report(s) must address
data quality and sufficiency to progress to the next level.
At TRL-4, assays increase in quantity, quality, and translational capability. The hy-
potheses on active compounds and MoA advance, helping efficacy and safety assess-
ments [
96
].
In vitro
, multitarget screening helps to elucidate the MoA and discloses off-
target effects [97–99].
Extracts, fractions, and pure compounds must be tested to identify those participating
in the biological effects. The impact of minor and unknown components must be addressed.
A set of tests shall be developed for bio-guiding the HP optimization. These should be easy
to run (preferably
in vitro
) and may evolve as knowledge accumulates. Dose–response
curves for different HPs and active isolated compounds shall be produced. Confirmatory
assays examining the same and other related pathways and disease models in different
species are advisable [
100
–
102
]. Collaboration between groups mastering different methods
is encouraged.

Pharmaceuticals 2024,17, 703 11 of 23
Whenever structural information is available, fast and low-cost in silico tests are also
recommended. Its relevance grows as quantitative structure–activity relationship (QSAR)
modeling and chemical databases improve [
69
,
103
]. Also, some tests integrating responses
of various compounds in mixtures are under development [104–106].
The careful selection, performance, interpretation, and reporting of the best models
are of foremost importance. Animal model predictions are often not confirmed in clinical
trials [
107
–
109
]. Therefore, the potential for extrapolating the conclusions to humans
shall be reviewed and deepened. Experiments must address aspects of efficacy, toxicity,
and capability to reach the target tissues/organs (PK). The report should discuss the
experiments’ strengths and weaknesses for translation to human outcomes. This includes
the relevance of the experimental models, the allometric dose transposal, and comparisons
with the controls.
The authors recommend performing the “killer experiment” as soon as possible after
the PoC, at the end of TRL-3 or the beginning of TRL-4. It aims at “killing” an unviable
technology before significant resources are spent. Its performance builds trust that, once
optimized, the technology will be competitive in the market [110].
TRL-5 focuses on regulatory-demanded GLP animal safety and toxicology
tests [78,79,111].
A peculiarity of HMPs is the possibility of waiving some non-clinical tests if sufficient data
from other sources are available [112,113].
For HMPs with a history of human use, the requirement for GLP toxicology studies is
determined by how closely the new HP matches the one previously used by humans. The
comparison encompasses composition, clinical application, exposure (dose and duration),
and the frequency and severity of any known adverse reactions [
16
,
79
]. Nonetheless,
genotoxicity and reproductive toxicity, whose risks are difficult to detect clinically should
be addressed, but exemptions apply [
79
,
114
,
115
]. The potential interaction with other
medicines also needs to be addressed [116,117].
For innovative HMPs, with no prior human exposure, most regulatory agencies
demand the same set of GLP toxicological tests required for small molecules, starting at
TRL-5 and finishing at TRL-7 or TRL-8 [79,118].
At TRL-5, tests needed for phase I clinical trial authorization include cytotoxic-
ity, genotoxicity, local tolerance (if applicable), acute toxicity, short-term repeated dose
toxicity, and safety evaluations of the cardiovascular, respiratory, and central nervous
systems [16,79,111,119].
TRL-6 encompasses tests necessary for phase II clinical trial authorization, specifically
repeated dose toxicity and reproductive toxicity studies [
62
,
111
]. Additional tests may be
required like the HMP batch-to-batch quality control biological test according to the United
States Pharmacopeia, chapters 1032 and 1033, required by the FDA [
120
,
121
]. Its development
starts at TRL-4 and finishes at TRL-8. Another example is the pharmacokinetic (PK) and
pharmacodynamic data (PD) demanded by the EMA for standardized HPs, but not for
quantified and other HPs. PK and PD data may be difficult to obtain if the effect depends
on mixtures but must be obtained in standardized extracts for at least one constituent with
known therapeutic activity. Also, the FDA states that “if feasible, chemical constituents
of a drug product that contribute to toxicity or pharmacology should be assessed in the
pharmacokinetic/toxicokinetic studies” [
62
]. These PK and PD data are not required for
HMPs by most other regulatory agencies [16,79,92].
If applicable, TRL-7 performs the tests demanded for continuous or long-term use
such as chronic toxicity. Carcinogenicity is required if there is evidence of genotoxicity, of
carcinogenicity in repeated dose toxicity, or if the composition holds a structure related to
a known carcinogen [
79
,
111
,
122
]. These must start before the phase III clinical trials and
have enough interim data to support the duration of these trials. Complete data will be
required for HMP licensing [62].
At TRLs 7 and 8, regulators accept in silico testing for assessing the safety of minor
components in standardized extracts. This method is particularly valuable when the

Pharmaceuticals 2024,17, 703 12 of 23
molecular structure is known and traditional toxicological
in vivo
or
in vitro
tests are
impractical [123–125].
Allometric Dose Conversion
Certain biological effects occur
in vitro
and
in vivo
at doses exceeding those feasible
for human use, making the findings clinically irrelevant. Allometric dose conversion
is employed to achieve comparable target tissue concentrations across different species.
For an accurate calculation, it is advised to adjust the methodologies used for isolated
substances to HMPs [126,127].
Killer Experiment
The “killer experiment” is aimed at stopping a project with low chances of success
before consuming significant resources. If a well-designed and well-conducted killer
experiment does not demonstrate the project’s unfeasibility, this is a compelling argument
for its continuity [110]. Examples of “killer experiments” adapted for HMPs are:
a.
Demonstration of low efficacy in doses suitable for humans after allometric calculation
in the gold-standard model of the target disease.
b.
Demonstration of efficacy/toxicity balance worse than a comparator under develop-
ment or already in the market, even after optimization. Note: if the MoA is different
from the available comparators, the product may still be viable, even if the potency
is lower or the toxicity is higher, because it might work in cases not responsive to
the comparator.
c.
Demonstration that a component responsible for a large part of the effect does not
reach an effective concentration in the target tissue/organ in a dose suitable for
humans after formulation optimization.
d.
Demonstration of active components’ instability, except for unmet medical needs. For
these cases, the search for stable related compounds is advisable.
e. Demonstration of manufacturing costs incompatible with the therapeutic indication.
2.2.6. Clinical Trials
Clinical trials for INDs have robust international regulations, which are out of the
scope of this paper [
118
,
128
]. The specific demands for HMPs depend on the degree of
innovation. In all cases, TRL-2 performs a comprehensive review of all available sources,
looking for traditional use, pharmacovigilance, clinical trials, accidental intoxications, and
other reports on human use. Literature-based comparisons with existing drugs for the
same indication are advisable to reinforce the medical utility of the proposed innovation.
The review must be updated at TRL-5, before starting clinical trials.
For innovative HMPs, with no prior human exposure, most regulatory agencies follow
the IND regulations [
118
]. In cases with reliable data on prior human use, some IND
requirements could be exempt. The sponsor is recommended to contact the regulatory
authority to obtain advice on the clinical development plan [62].
At TRL-6, phase I clinical trials address aspects of safety and PK for the constituents
with known therapeutic activity. The FDA and EMA have specific recommendations
applicable to first-in-human studies which are worth consulting [
73
,
129
]. For HMPs,
whenever enough information is available from traditional use, phase I clinical trials in
normal volunteers are usually waived. A substantial prior human use of the HMP generally
conveys confidence that similar amounts are safe for small numbers of carefully monitored
subjects in phase II trials [
16
,
62
]. However, the FDA recommends getting PK parameters to
achieve the same objectives of clinical pharmacology studies for nonbotanical drugs, if the
major active constituent(s) in a botanical product is(are) known [62].
At TRL-7, phase II clinical trials explore efficacy, safety, and dose-finding in small
populations. If the phase I clinical trial is waived and phase II is the first-in-human test,
consult the FDA and EMA-specific regulations [
73
,
129
]. This is one of the most critical
stages of pharmaceutical development, often referred to as the “valley of death”, where

Pharmaceuticals 2024,17, 703 13 of 23
most clinical failures are seen [
97
]. Although generally regarded as a PoC in humans, the
phase II clinical trials do not necessarily produce statistically significant results. Instead,
they provide information suggestive of efficacy that enables the sample size calculation
for the confirmatory phase III clinical trials. Moreover, TRL-7 must generate dose-ranging
data. Some regulatory agencies demand dose-ranging studies if the scientific literature
does not contain valid data [
16
]. The FDA demands the dose selection rationale to be based
on experimental human PK data, regardless of previous marketing experience [62].
At TRL-8, phase III clinical trials confirm efficacy and safety in broader patient pop-
ulations. These trials are designed to yield statistically significant results for predefined
variables through predetermined statistical methods (estimands) [
130
]. They also expand
the safety evaluations to larger cohorts to uncover rare adverse events [
16
,
113
]. Achiev-
ing marketing authorization from at least one regulatory agency signifies the completion
of TRL-8 criteria. Therefore, it is recommended for sponsors to engage with regulatory
agencies before initiating any phase III trial [
62
]. At TRL-9, the sponsor may conduct
phase IV clinical trials to further elucidate the HMP’s therapeutic profile or to assess
new formulations.
Special Issues on Clinical Trials of Herbal Medicinal Products
Although considered the gold standard for randomized-controlled clinical trials, the
use of a placebo is especially challenging for HMP development [
131
]. The possibilities
for placebos include excipients and other pharmacologically inert ingredients. However,
HMPs usually have typical organoleptic properties such as taste, odor, and appearance that
might be difficult to mask [
132
]. A possibility is using other botanical materials with no
pharmacological activity. However, concerns about an unsuspected activity might always
arise. Another possibility is a double-dummy design that randomizes participants to take
either the active treatment “1” and the placebo of the treatment “2”, or the placebo of the
treatment “1” and the active treatment “2” [
133
]. The FDA understands that it might be
difficult to select an ideal placebo for HMPs and encourages the sponsor to consult with
the agency beforehand [62].
Another challenge is the adjuvant treatment of serious conditions to which the FDA
recommends the “add-on to standard care versus standard care” [
62
]. In Brazil, according
to the Brazilian National Health Council, the placebo may be used only in cases where
no recognized helpful treatment is available. Also, for these cases, the add-on design
is recommended.
To minimize different understandings, the WHO published the guideline “Operational
guidance: Information needed to support clinical trials of herbal products”. This should be
reviewed before designing the trials alongside the national regulations of the country
where the HMP is intended to be licensed [16].
Ethnopharmacological Information, Traditional Use
The research on NPs has historically been guided by ethnopharmacology. The level
of information spans from an anecdotal mention of use to long-term, high-quality phar-
macovigilance documentation with detailed descriptions of the HP, therapeutic effects,
recommended doses, and adverse events. Whenever available, no human data shall be ne-
glected. The FDA, ANVISA, and EMA demand the submission of all information available
on prior human exposure [62].
Ethnopharmacology can be a development starter or accelerator. In some cases, it
starts a research program to confirm and clarify the pharmacological potential. In other
cases, it justifies skipping some non-clinical assays and clinical trials after scaling-up issues
have been addressed. If consistent information on HP use by humans is available, it might
even be unethical to submit animals or humans to IND-directed studies [112,113].
However, handling ethnopharmacological information holds many challenges. The
first is species identification. Many historical records only mention common names, which
can be attributed to multiple plants across different botanical families, complicating ac-

Pharmaceuticals 2024,17, 703 14 of 23
curate identification. The next challenge is the HP description, including the part of the
plant used, extractive methods, route of administration, and doses, among others. Once
these challenges are overcome, the value of the ethnopharmacological source is given by
the richness and reliability of the information on human effects. Traditional communities
may employ terminology vastly different from that of Western medicine, potentially com-
plicating the interpretation of therapeutic effects. However, if clinical manifestations are
documented, it may be feasible to translate these into terms recognizable within Western
medical frameworks [
134
]. This can be the case for more consolidated and documented al-
ternative medical systems such as Ayurveda, Traditional Chinese Medicine, Unani, Siddha,
and other herbal medicines [113].
Moreover, according to the FDA [
62
], “when the rationale for developing certain
botanical drug products is based on prior clinical experience in alternative medical systems
e.g., Ayurveda,
Traditional Chinese Medicine, Unani, Siddha, and other herbal medicine
and pharmacognosy textbooks), the sponsor may propose to incorporate traditional prac-
tices into their clinical protocols. For example, patients may be selected or grouped based
on alternative medical practice and treated with specific botanical regimens accordingly,
or the final dosage form may be prepared by individual patients according to traditional
Chinese or Indian methods”.
3. Materials and Methods
The TRLs for HMPs were based on the original NASA concepts from aeronautical
development [
33
,
34
,
48
–
52
,
55
,
135
]. A search was conducted on the Web of Science, PubMed,
Embase, and Scopus databases using MeSH descriptors and keywords related to technology
readiness levels (TRLs) in pharmaceutical development, identifying
360 references.
After
removing duplicates, 262 articles were analyzed, and 12 were selected for further consid-
eration based on the specific mention of TRL in any biological
field [40,136–146].
None
specifically addressed TRLs in pharmaceutical development, including small molecules,
biotechnological products, and herbal medicinal products (HMPs) for human use. The
closest were in chemistry [
40
], drugs and vaccines for animal health [
145
], and the bioin-
dustry [
146
]. Some R&D agencies’ websites have adapted technology readiness level (TRL)
concepts to pharmaceutical development, serving as both starting points and founda-
tions upon which we built our concepts for HMPs [
35
–
38
]. Drawing from these sources
and our professional experience, we applied technology readiness level (TRL) concepts
to herbal medicinal products (HMPs), creating a list of milestones to be reached at each
level. These milestones were categorized into six domains and aligned with relevant
guidelines from the ANVISA, EMA, FDA, the International Council for Harmonisation of
Technical Requirements for Pharmaceuticals for Human Use (ICH), the Organisation for
Economic Co-operation and Development (OECD), and the World Health Organization
(WHO). Considering the extensive European expertise in HMP regulation, we embraced
the terminology established by the EMA. The correspondence of EMA terminology to that
of the FDA and ANVISA is in Table 3.
Table 3. Terminology correspondence between EMA, FDA, and ANVISA.
EMA Term [31] FDA Term [48] ANVISA Term [63] EMA Definition
Herbal
substance
Botanical drug
substance Droga vegetal
Mainly whole, fragmented, or cut plants, plant parts,
algae, fungi, and lichen in an unprocessed, usually
dried form, but sometimes fresh. Certain exudates
that have not been subjected to a specific treatment are
also considered to be herbal substances. Herbal
substances are precisely defined by the plant part used
and the botanical name according to the binomial
system (genus, species, variety, and author) [114].

Pharmaceuticals 2024,17, 703 15 of 23
Table 3. Cont.
EMA Term [31] FDA Term [48] ANVISA Term [63] EMA Definition
Herbal preparation Botanical drug
preparation Derivado vegetal
Preparations are obtained by subjecting herbal
substances to treatments such as extraction,
distillation, expression, fractionation, purification,
concentration, or fermentation. These include
comminuted or powdered herbal substances, tinctures,
extracts, essential oils, expressed juices, and processed
exudates [115].
Herbal medicinal
product
Botanical drug
product Fitoterápico
Any medicinal product, exclusively containing as
active ingredients one or more herbal substances or
one or more herbal preparations, or one or more such
herbal substances in combination with one or more
such herbal preparations [116].
4. Concluding Remarks
This TRL-based roadmap is primarily designed to facilitate collaboration among
Brazilian scientists, manufacturers, and funding agencies in the development of innovative
herbal medicinal products (HMPs) targeting global markets. However, it could also be
beneficial for developers in other megadiverse countries looking to navigate similar paths.
Biodiversity along with ethnopharmacological information are still very underexploited
assets from megadiverse countries for drug discovery and development [
147
–
150
]. Most mar-
keted HMPs are based on European plants or traditional Chinese
medicine [151,152].
After the
Nagoya Protocol, some large pharmaceutical companies have scaled back their bioprospect-
ing activities [
1
,
22
,
153
]. This presents a potential window of opportunity for research
institutions in megadiverse countries.
The challenges are many. The pharma R&D processes are not mature in tropical
countries’ manufacturers. HMPs lack regulatory harmonization, and some legal uncertain-
ties from the Nagoya protocol still remain. Also, the HMPs’ R&D is more complex than
synthetic small molecules because the HP composition is susceptible to variations in plant
strain, place and season of cultivation/collection, and extraction and storage conditions.
This roadmap proposal is an effort to overcome some of these challenges. It should be
viewed as a general guidance meant to be tailored to each unique development scenario,
acknowledging that every case is distinct and has its own set of specific characteristics.
As previously noted, the established TRL scales lack boundary indicators [
28
,
113
].
This was the pioneers’ proposal and the main reason for TRLs’ universality and application
to many diverse technologies. Nevertheless, this allows the classification of the same project
in different TRLs, depending on subjective judgments. To avoid this ambiguity, besides the
TRLs transposal, we created a list of indicators organized into six domains that progress
with the TRLs. The domains help project management and allow more precise indicators.
Due to the universality of the TRL concepts, the transposal consistently met the HMP’s
R&D milestones. Aeronautical and pharmaceutical developments share some similarities.
Both start with material properties, followed by a proposal of application. However,
one important difference was the requirement of experimental data at TRL-2 to design
the PoC. This was due to the larger dependence of pharmacology on experimentation.
TRL-3 was kept as the PoC proposed by Mankins. This crucial step concerns a low-cost
experiment (or group of experiments) that needs to be convincing enough to justify the
project’s progression.
TRL-4 is very important for the project’s success and its first major risk inflection
point. For HMPs, the “validation in laboratory environment” proposed by Mankins was
translated as the complex, long, expensive, and laborious bench-scale HP optimization.
In our opinion, the lack of innovation in HMPs is mostly due to insufficient optimization.
Several projects have progressed unoptimized to GLP testing or even to clinical trials
and went back to initial stages or were abandoned afterward, with inconsistent results.

Pharmaceuticals 2024,17, 703 16 of 23
Therefore, we equate TRL-4 with the first “valley of death” [
154
]. It is the gap between
the fundamental research and the regulatory-demanded non-clinical and clinical tests that
finalize the innovation process. This gap still endures, despite the availability of non-
refundable funding specifically directed to innovation, perhaps due to the unawareness of
its importance and complexity [155].
The killer experiment is a good managerial principle to avoid spending scarce re-
sources on wrong projects. It reduces technological risk and justifies the scaling-up invest-
ment. In case of failure, it might be necessary to abort or to improve the HP candidate,
meaning that fundamental science is lacking. Therefore, in some cases, it might drive the
interruption of the technological aspect to stimulate fundamental science. It should be
envisioned early on at TRL-2 and executed at TRL-3 to TRL-4. Every innovative project
achieving TRL-4 demands broader pharmacological knowledge. Hereon, the more the
project matures and the closer it gets to the market, the more information will be needed to
minimize the clinical trials’ risks. These studies are generally carried out in research centers
and shall not be confused with the technological project itself but establish a relationship of
reciprocal benefits. In this way, while a technological project progresses to higher TRLs,
phytochemical and pharmacological scientific studies can feed it with valuable information
and receive suggestions that can motivate new research lines.
TRL-5 has two important goals: scaling up and validating the HP. Considering this,
the “relevant environment” proposed by Mankins was translated as the GLP condition. It
is also time to improve the manufacturing standards to face the clinical trials’ demands.
TRL-6 has another remarkable similarity between aeronautic and pharmacological
development—the first flight and the first human test demand substantial preceding
validations. Additionally, the object of development is called by Mankins: “component” at
TRL-5, “system/subsystem” at TRL-6, “prototype” at TRL-7, and “actual system” at TRL-8.
This bears close similarity with the HMP. At TRL-5, the “component” is the HP and, at
TRL-6 and TRL-7, the prototype and system/subsystem correspond to the evolving HMP
formulation to be administered to humans. At TRL-8, the “actual system” corresponds to
the HMP that goes to the market. In either case, models could be considered one-order-of-
magnitude approximations for performance [55].
Mankins also stated that “not all technologies undergo TRLs 6 and 7”, but, by defini-
tion, all go through TRL-8, which completes the “system development” [
52
]. For HMPs,
those projects with previous human data can skip at least TRL-6, and in the case of previ-
ously established doses according to traditional use, TRL-7 could also be skipped, but, by
definition, all successful projects must have approval by at least one regulatory authority,
which characterizes the TRL-8 completion.
TRL-9 holds an important difference between HMPs and astronautic development.
Except for new formulations demanding new clinical trials, no changes to the HMP or
manufacturing chain are allowed.
As a final remark, the TRLs do not provide insight into the uncertainty in pursuing
further maturation levels. Therefore, they are not substitutes for individualized expert
assessments of each project. Instead, they provide tools to guide the expert assessment. For
risk assessment, an additional metric is needed: the R&D degree of difficulty as described
elsewhere [34,52].
Future Implications
This TRL-based roadmap was created to enhance HMP development in megadiverse
countries with scientific rigor and regulatory compliance. It facilitates streamlined manage-
ment and resource allocation, fostering trust among scientists, manufacturers, and funding
agencies in the planning and execution of HMP development projects. We anticipate that it
will promote sustainable practices and catalyze therapeutic breakthroughs from underuti-
lized plant species, thereby increasing the introduction of innovative HMPs. The future
application of this roadmap in concrete cases will reveal areas for potential improvements.

Pharmaceuticals 2024,17, 703 17 of 23
Author Contributions: Conceptualization, E.P., C.D.R., C.M.S., S.A.C.P., B.S.B., A.C.B.C., and M.D.B.;
investigation, E.P., C.D.R., C.M.S., S.A.C.P., P.J.C.B., B.S.B., and M.D.B.; methodology, E.P., C.D.R.,
C.M.S., S.A.C.P., and M.D.B.; validation, E.P., C.D.R., C.M.S., A.C.B.C., and M.D.B.; writing—original
draft, E.P.; writing—review and editing, E.P., C.D.R., C.M.S., S.A.C.P., P.J.C.B., B.S.B., A.C.B.C., and
M.D.B. All authors have read and agreed to 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: The raw data supporting the conclusions of this article will be made
available by the authors on request.
Acknowledgments: The authors wish to thank Henry Suzuki (Axonal) and Ana Claudia Dias
(ABIFINA) for their important contributions to the intellectual property of herbal medicinal products,
Rodrigo Rocha Secioso de Sá(FINEP) for contributions regarding the funding of research and
development projects, Carlos Eduardo Vitor for information about final product development, and
Emerson Queiroz (UniGe) for important contributions about the international herbal medicinal
products R&D scenario.
Conflicts of Interest: The authors declare that the research was conducted in the absence of any
commercial or financial relationships that could be construed as potential conflicts of interest.
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